Outgrow your limits
The Nara Institute of Science and Technology (NAIST) was founded
in 1991 as a Japanese national university with the aim of conducting
advanced research and educating scientists and technologists to
support the development of society. NAIST is comprised of the
Graduate School of Science and Technology, which focuses on theareas of information, biological and materials sciences and thedevelopment of their interdisciplinary fields. At present, over 1,000
students—roughly 25% from overseas—are supervised by roughly
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NAIST Research Highlights
ISSN 2189-714X

For further information on the research
published in NAIST Research Highlights or
to arrange an interview with a faculty,
please contact:

NAURA (NAIST URA Team)
Institute for Research Initiatives
Nara Institute of Science and Technology
8916-5 Takayama-cho, Ikoma, Nara 630-0192, JAPANE-mail: uramember@rsc.naist.jp©2023 NAIST All rights reserved.

Aims & Scope

NAURA

NAIST Research Highlights showcases
promising and important researchachievements at NAIST and presents
their current research and coretechnologies to the public. The
publication distills highly technicalresearch papers into short,easy-to-understand articles that appeal
to a global audience of both specialists
and non-specialists. NAIST ResearchHighlights aims to inform readers of
the latest developments in NAIST’s
pioneering research and to stimulate
new and existing international
collaborations.

NAURA (NAIST URA team)publishes NAIST Research Highlights
under the auspices of the “Program for
Promoting the Enhancement ofResearch Universities”, which is fundedby Japan’s Ministry of Education,
Culture, Sports, Science andTechnology (MEXT). NAURA
facilitates pioneering researchdevelopments, international researchcollaboration activities andinterdisciplinary research projects tofurther strengthen NAIST’s researchcapabilities.

NAIST Divisions and Laboratories
NAIST conducts advanced research in the core study areas of information, biological and materials sciences and engages ininterdisciplinary studies to explore and seek solutions in the most challenging areas. NAIST actively addresses important social issues bygenerating international-level research products that will help to build a better society.

Division of Information Science
Computer Science

Media Informatics

Applied Informatics●Computing Architecture●Natural Language Processing●Human Robotics●Dependable System p.6●Ubiquitous Computing Systems●Software Engineering●Augmented Human Communication●Social Computing

 p.7●Robot Learning p.8●Large-Scale Systems Management●Network Systems●Mathematical Informatics●Software Design and Analysis●Interactive Media Design●Imaging-based Computational●Cyber Resilience●Optical Media Interface●Information Security Engineering●Cybernetics and Reality Engineering p.10 p.15

Biomedicine●Computational Systems Biology●Internet Architecture and Systems●Computational Behavioral

 p.18

Neuroscience

Division of Biological Science
Plant Biology●Plant Developmental Signaling p.20●Plant Metabolic Regulation

Biomedical Science

Systems Biology●Molecular Signal Transduction●Microbial Molecular Genetics●Functional Genomics and Medicine  p.29

●Applied Stress Microbiology

 p.34●Plant Growth Regulation

 p.28●Tumor Cell Biology●Environmental Microbiology

 p.36●Plant Stem Cell Regulation and p.22●Molecular Immunobiology●Structural Life Science p.38Floral Patterning●Plant Physiology

 p.24●Plant Immunity●Plant Symbiosis

 p.26●Plant Secondary Metabolism

 p.30●Molecular Medicine and Cell Biology  p.32

●Gene Regulation Research●RNA Molecular Medicine●Systems Neurobiology and Medicine  p.39

●Stem Cell Technology●Bioengineering●Developmental Biomedical Science●Data-driven Biology●Organ Developmental Engineering●Plant Regeneration and Morphogenesis

Division of Materials Science
Core Laboratories●Quantum Materials Science

Core Laboratories(Cooperative)●Functional Organic Chemistry●Bio-Process Engineering p.42●Solid-state Information Physics

 p.46●Photonic Device Science●Biomimetic and Technomimetic

 p.52

Molecular Science●Functional Supramolecular●Information Device Science

Chemistry●Applied Quantum Physics●Complex Molecular Systems●Organic Electronics

 p.48●Nanomaterials and Polymer Chemistry●Photonic and Reactive Molecular p.54●Materials Informatics

Science●Data-driven ChemistryCore Laboratories(Collaborative)●Mesoscopic Materials Science●Sensory Materials and Devices●Functional Polymer Science●Ecomaterial Science●Advanced Functional Materials

As of December 22, 2022
NAIST Research Highlights

1

Contents
1

NAIST Divisions and Laboratories

4

Editorial

8

Social Computing

10

Optical Media Interface

12

Optical Media Interface

14

Optical Media Interface

15

Mathematical Informatics

16

Mathematical Informatics

18

Computational Behavioral Neuroscience

21

Plant Developmental Signaling

22

Plant Stem Cell Regulation and Floral Patterning

23

Plant Stem Cell Regulation and Floral Patterning

24

Plant Physiology

26

Plant Symbiosis

Information Science6

Dependable System

7

Software Engineering

Toward accurate modeling of power
MOSFET electrical characteristics
Software ecosystem:
Everybody needs somebody

COVID vaccine:
Who's searching for reassurance?
Turning the tables into touchscreens
Keeping the light from fading
A technology for embedding data
in printed objects
New model can predict a patient’s responsivenessto fMRI-based mental health treatment
How is the brain programmed forcomputer programming?
Studying the OCD cycle

Biological Science20

2Plant Developmental Signaling

Switching roles: Key proteins evolved from
activators to maintainers in plants

NAIST Research Highlights

A new role of autophagy in plant celldifferentiation revealed
How to beat the heat: Memory mechanism
allows plants to adapt to heat stress

The flower clock: How a small protein helps
flowers to develop right and on time
The circadian clock makes sure plant
cells have the time of their lives
When plants attack: Parasitic plants
use ethylene as a host invasion signal

28

Plant Growth Regulation

34

Applied Stress Microbiology

29

Functional Genomics and Medicine

36

Environmental Microbiology

30

Molecular Immunobiology

38

Structural Life Science

39

Systems Neurobiology and Medicine

40

Plant Cell Function

51

Organic Electronics

52

Biomimetic and Technomimetic Molecular Science54

Photonic and Reactive Molecular Science

31

32

33

Damage control: Plants juggle genome
maintenance and growth by being organized
A single allele deletion in gene encoding
Zbtb38 leads to early embryonic death
Tropical ginger treatment for
blocking inflammation
Molecular Immunobiology

Alveolar macrophages help CD8+
T cells go (anti-)viral
Molecular Medicine and Cell Biology

How cellular fingertips may help
cells “speak” to each otherJapanese sake: The new pick-me-up?
Yeast strain makes fatigue-fighting ornithine
How a bacterium may help solve
the plastic pollution crisis
A protein with an unprecedented fold helps
bacteria uptake thiosulfate as a sulfur source
Shootin1a – The missing link underlying
learning and memory
Hired blade: Anchoring complex in plant cells
recruits its own katana sword

Molecular Medicine and Cell Biology

AI knows where your proteins go

Materials Science42

Bio-Process Engineering

43

Bio-Process Engineering

44

Bio-Process Engineering

45

Bio-Process Engineering

46

Solid-state Information Physics

48

Organic Electronics

50

Antibiotic resistance testing no longer
impeded by time
Feeling the pressure

Separating tiny bacteria by shape:
Simple techs for E. coli sorting
From outside to inside:
A rapid and precise total assessment
method for cells
Old silicon learns new tricks

Easy fabrication of next-generation,
super-flexible electronic circuits
Organic Electronics

Shedding light on polymer solar cells:
Illuminating how solvent additives
improve efficiency

Fill me up: Improved efficiency of
all-polymer solar cells
Gearing up nanoscale machines
Invisible X-rays turn blue

55 Innovative Research and Education Programs
NAIST Research Highlights

3

NARA INSTITUTE of SCIENCE and TECHNOLOGYRESEARCH HIGHLIGHTS 2023

Nurturing
discoveryW

elcome to NAIST Research Highlights 2023. This publication
provides an overview of recent research achievements of
one of Japan’s leading research bodies, the Nara Institute of
Science and Technology. In these pages we present a view of recent
extraordinary advances and breakthroughs, including 36 topics fromthe three divisions.
Of course, at the heart of such outcomes are our researchers. Each year
NAIST attracts more leaders in scientific fields. We look forward tonurturing their important work and the careers of all our researchers in
the atmosphere of cutting-edge discovery that will continue to befostered at NAIST.

On the cover
By analyzing social concerns
regarding COVID-19 vaccination based on search engine
queries, NAIST researchers
assist in improving online
messaging about vaccination.
Page 84NAIST Research Highlights

INFORMATIONSCIENCEDependable System
Prof. Michiko Inoue

Toward accurate modeling of power
MOSFET electrical characteristics
Researchers at NAIST use automatic differentiation to dramatically speed up the calculation ofmodel parameter extraction for metal oxide semiconductor field-effect transistor data, which maylead to more energy efficient power converters.

S

cientists from NAIST have developed the mathematical method
called automatic differentiation to find the optimal fit of experimental data up to four times faster. This research can be applied
to multivariable models of electronic devices, which may allow them
to be designed with increased performance while consuming lesspower.
Wide bandgap devices, such as silicon carbide (SiC) metal-oxide
semiconductor field-effect transistors (MOSFET), are a criticalelement for making converters faster and more sustainable. This isbecause of their larger switching frequencies with smaller energy
losses under a wide range of temperatures when compared with conventional silicon-based devices. However, calculating the parametersthat determine how the electrical current in a MOSFET responds asa function of the applied voltage remains difficult in a circuit simulation. A better approach for fitting experimental data to extract theimportant parameters would provide chip manufacturers the ability
to design more efficient power converters.
Now, a team of researchers led by NAIST has successfully used themathematical method called automatic differentiation (AD) to significantly accelerate these calculations. While AD has been used extensively when training artificial neural networks, the current projectextends its application into the area of model parameter extraction.
For problems involving many variables, the task of minimizing the error is often accomplished by a process of “gradient descent,” in which
an initial guess is repeatedly refined by making small adjustments inthe direction that reduces the error the quickest. This is where AD

can be much faster than previous alternatives, such as symbolic or
numerical differentiation, at finding direction with the steepest “slope”.
AD breaks down the problem into combinations of basic arithmetic
operations, each of which only needs to be done once. “With AD, thepartial derivatives with respect to each of the input parameters are
obtained simultaneously, so there is no need to repeat the model evaluation for each parameter,” first author Michihiro Shintani says. By
contrast, symbolic differentiation provides exact solutions, but uses
a large amount of time and computational resources as the problem
becomes more complex.
To show the effectiveness of this method, the team applied it
to experimental data collected from a commercially available SiC
MOSFET. “Our approach reduced the computation time by 3.5× incomparison to the conventional numerical-differentiation method,which is close to the maximum improvement theoretically possible,”
Shintani says. This method can be readily applied in many other areasof research involving multiple variables, since it preserves the physicalmeanings of the model parameters. The application of AD for theenhanced extraction of model parameters will support new advancesin MOSFET development and improved manufacturing yields.

Overview of the mathematical method developed in this study.
6

NAIST Research Highlights

Reference

Michihiro Shintani, Aoi Ueda, Takashi Sato. 2022. Accelerating parameter
extraction of power MOSFET models using automatic differentiation.
IEEE Transactions on Power Electronics, 37, 2970‒2982.

Software Engineering
Prof. Kenichi Matsumoto

Software ecosystem:
Everybody needs somebody
Researchers at NAIST employ network analysis methods from social science to study volunteer
contributions to open-source software libraries, and find correlations between dependency networks and viability, which may identify libraries about to become dormant.

U

107,242 different libraries. “Peaks in our generated metrics correlate
sing socio-technical techniques, researchers from NAISTwith important ecosystem events,” says senior author Kenichi Matsuhave measured the congruence between the network of
moto.
contributors to open-source programming libraries and theThis research may help keep software running and identify fragile
dependencies of that library within the ecosystem. This work suggests
points in the dependency network, and may ultimately encouragethat the level of matching between the network of contributors anddependency contributions that support the maintenance of interdenetworks of dependencies could be used as an indicator of libraries atpendent third-party libraries used in software development.
risk of becoming inactive.
The modern computer programs that run your favorite apps or
websites can be extremely large, often measured in millions of lines ofcode. This is obviously much more complex than can be handled by
Reference
any one individual. Most programming languages therefore rely on
Supatsara Wattanakriengkrai, Dong Wang, Raula Gaikovina Kula,
specialized modules called third-party libraries to accomplish specific
Christoph Treude, Patanamon Thongtanunam, Takashi Ishio, Kenichitasks. These libraries are often open-source and freely available toMatsumoto. 2022. Giving back: Contributions congruent to libraryanyone who wants to download and use them. For example, programdependency changes in a software ecosystem. IEEE Transactions onmers in JavaScript have access to over one million libraries, while
Software Engineering, in press.
there are more than 300,000 libraries for the Python community. The
libraries themselves often rely on each other, withthe typical library requiring the use of about fiveothers. However, the ecosystem of interconnected
libraries and their dependencies on each other ispoorly understood, which is concerning since a
failure in one could have cascading effects on theentire system. Sustained contributions are crucial,because the dependencies of any one library onothers must be constantly updated in response tochanges. However, maintainers of these libraries areoften overworked and often contribute as unpaid
volunteers.
Now, a team of researchers led by NAIST has
studied these networks by defining a metric called“dependency-contribution congruence” (DC congruence), which measures how closely the networkof library dependencies matches the network ofcontributor changes. The congruence metric islargest when the same contributor makes changes
to both a library and its dependents. “We found that
DC congruence shares an inverse relationship withthe likelihood that a library becomes dormant. Specifically, a library is less likely to become dormant
if the contributions are congruent with upgrading
dependencies,” says first author Supatsara Wattanakriengkrai. The team measured the DC congruence
within the npm ecosystem of JavaScript libraries andLibrary-level DC congruence in npm JavaScript Ecosystem.
analyzed over 5.3 million change commits across
NAIST Research Highlights

7Social Computing

Prof. Eiji Aramaki

COVID vaccine:
Who's searching forreassurance?
Using a vaccine concern index based on internet searches on the COVID-19 vaccine, researchersat NAIST find that “adverse reaction” searches positively correlate with adoption, which may help
online resources encourage vaccination.

8

NAIST Research Highlights

Societal concerns over COVID-19 vaccines as shown by the vaccine concern index that is calculated
from vaccine-related queries searched on Yahoo! JAPAN.

S

ince becoming available, vaccines
against COVID-19 have been vital inpreventing deaths. However, vaccination rates in many developed countries are
below expectations of public health officials,
which is partly due to vaccine hesitancy.
Scientists from Japan have found that searches related to vaccine side effects actually
increased with adoption rates of vaccination.
This work may help public health officials
better understand vaccine hesitancy.
Va c c i ne s ag ai nst C OV I D - 1 9 we reremarkable in their rapid progress throughdevelopment, manufacture, and introduction
to the market. However, the rate of their up-

take among health consumers did not match
this speed. In Japan, a large survey foundthat many people expressed concerns about
adverse reactions and vaccine effectiveness.
Even before the pandemic, Japan had scoredlow on measures of trust in vaccines. Thus,
being able to understand and measurepeople’s concerns could help shape public
relations outreach efforts with the goal ofincreasing vaccine adoption.
A team of researchers led by NAIST has
examined online searches on Yahoo! JAPANto quantify the degree of unease about getting
the COVID-19 vaccine. They worked with
Yahoo! JAPAN to develop a ‘vaccine concern
index’ (VCI) based on the aggregatedsearch counts of vaccine-related queries byprefecture for August and September 2021.
The VCI captured searches of terms that may
indicate vaccine hesitancy (such as adverse
reactions and side effects) out of all vaccine-related searches (including searches for
vaccination centers or appointments). The
team found that the concern index tended
to be lower in more populated areas, buthigher in people in their 20s to 40s, especiallyfemale users. “This could be related to theonline spread of misinformation that thevaccine causes infertility,” says author Shoko
Wakamiya, “and lower concerns among older
people may be because they have limited

familiarity with online searching.”
The team found that the VCI was larger
in prefectures with higher vaccination rates.
This suggests that web searches for vaccine
adverse reactions may be one step taken byhealth consumers before ultimately choosing
to get vaccinated, which highlights the importance of providing complete and accurate
information online. Conversely, individuals
strongly hesitant towards vaccination may
not be looking for online information, or are
looking online less often. “As part of a collectivistic, Confucian culture, Japanese people
often have an interdependent view of self,”
explains senior author Eiji Aramaki. “As a result, ‘greater good’ messaging and guidelines
may be more effective in Japan.” This work
may pave the way for equally cost-effective
examinations of societal attitudes towards
COVID-19 vaccination in similar cultures,and ultimately improving online resources
about these vaccines.

Reference

Makoto Uehara, Sumio Fujita, Nobuyuki Shimizu,
Kongmeng Liew, Shoko Wakamiya, Eiji Aramaki. 2022.
Measuring concerns about the COVID‑19 vaccine
among Japanese internet users through search queries.
Scientific Reports, 12, 15037.

NAIST Research Highlights

9Optical Media Interface

Prof. Yasuhiro Mukaigawa

Turning the tables into
touchscreens
Researchers at NAIST develop a projected touchscreen system that requires
only one camera and one projector, allowing for the production of inexpensive large interactive displays and computer interfaces.

10

NAIST Research Highlights

S

cientists from NAIST have developed
a touchscreen-like interface on a flat
surface using a synchronized projector
and camera. B ecause the camera only
registered the user’s fingers when actuallyin contact with the surface, the need for any
additional cameras, depth sensors, or lightsources was eliminated. This work may leadto portable projection systems that can beused to create large interactive displays nearly
anywhere.
Everyday life is full of handheld devices
that show videos and accept touch input.
These include smartphones, tablets, or eventhe credit card reader at your local store.
However, larger touchscreen interfaces
remain expensive and slow to respond. Projection systems that are currently available
need multiple detectors, because a single

camera usually cannot distinguish depth.
Now, a team of researchers led by NAIST has
created a system that uses just one camera.
“Typical cameras observe a three-dimensional situation as a two-dimensional plane.
Thus, even if the position of a fingertip canbe detected, it is difficult to know whether it
is touching the surface or hovering above it,”senior author Yasuhiro Mukaigawa says.
The researchers took advantage of “slope
disparity gating,” in which the projector
scanning the table is synchronized with thecamera, so that it captures only the regionslightly above the surface. Using a dedicatedimage-processing algorithm, the touch of afinger could be localized with high efficiency.
Because the same light source is used to
produce the projected video as well as detect
touch, the algorithm was not misled by the

projected image.
“The method developed in the study canbe used to produce a touch display on any flat
surface. In the future, we will hope to expand
to include touchless operations or even add
gesture recognition,” Mukaigawa says. Also,
because the method captures only the regionslightly above the surface, privacy concerns
are reduced, as human faces and otheridentifying details are not recorded at all.

Reference

Mayuka Tsuji, Hiroyuki Kubo, Suren Jayasuriya,
Takuya Funatomi, Yasuhiro Mukaigawa. 2021. Touch
sensing for a projected screen using slope disparity
gating. IEEE Access, 9, 106005–106013.

NAIST Research Highlights

11

Optical Media Interface

Prof. Yasuhiro Mukaigawa

Keeping the lightfrom fading
Researchers at NAIST develop a new mathematical method to calibratescans of stained-glass windows to account for changes in natural light,
which may help assist in the accurate preservation of famous artworks.

12

NAIST Research Highlights

S

cientists from NAIST have created a
new approach to compensate for variations in illumination while scanning
cathedral stained-glass windows. This work
may be applied to other objects of cultural
significance to help capture their colors in themost lifelike way.
It’s hard to think of a more inspirational
experience than watching the sun slowly set
through historic stained-glass windows, suchas those found in the cathedrals in Europe.
While the changing light levels over time may
be breathtaking, it also makes high-resolutionscans of the windows more challenging. Thatis, if the scanning process requires minutes
or even hours to complete, variations in thenatural illumination can lead to inconsistent
results.
Now, a team of researchers led by NAIST
has developed a new calibration method
to help compensate for changes in the sun’sillumination over the course of the scan.
“It can take hours to capture thousands ofspectral channels pixel by pixel. Thus, themeasurement can be significantly affectedby the perturbations in natural light,” firstauthor Takuya Funatomi says.
The researchers set out to capture hyperspectral images of the famous stained-glass
windows in the Amiens Cathedral in France.
With some window panels dating back to the13th century, this location which has been
designated as a UNESCO World Heritage
Site. A whisk-broom scanner was used toacquire hyperspectral images. This kind of

sensor uses a movable mirror to slowly scanacross an object. Each pixel is measured one
at a time as its light is reflected onto the single
detector with the sky in the background.
However, when it is applied to outdoor
cultural heritages, temporal illumination
variations become an issue due to the lengthy
measurement time. Hyperspectral scanning
is not limited to the wavelengths of light thatare visible to humans. For this research, theteam used a spectrometer that recorded morethan 2,000 channels over a spectrum ranging
from about 200 nm to 1100 nm, which includes ultraviolet, visible and infrared colors.
An extra single column scan was added
to help calibrate the images. Using matrix
methods, variations in temporal illumination
could be removed. This allowed for much
more accurate results compared with simply
normalizing the total brightness, because
each color might be impacted differently by
the changing light. “Our method provides a
new modality for the digital preservation oflarge cultural assets,” senior author Yasuhiro
Mukaigawa says. This method can be easily
adapted to other situations in which outdoor
scanning has to occur over long time periods.
Reference

Takuya Funatomi, Takehiro Ogawa, Kenichiro Tanaka,
Hiroyuki Kubo, Guillaume Caron, El Mustapha
Mouaddib, Yasuyuki Matsushita, Yasuhiro Mukaigawa.
2022. Eliminating temporal illumination variations
in whisk-broom hyperspectral imaging. International
Journal of Computer Vision, 130, 1310–1324.

Color compensation procedure developed in this study.

NAIST Research Highlights

13

Optical Media Interface
Prof. Yasuhiro Mukaigawa

A technology for embedding data
in printed objects
Researchers at NAIST develop a new method to embed information in a 3D printed object
without modifying the shape of the object and retrieve it using a single image of a commerciallyavailable document scanner.

Dmethod conducts this procedure only with a common document
igital watermarking is a technology that embeds information
scanner, and does not require any special equipment. The FDM printinside digital contents such as image, audio, video, and 3D
ing process naturally produces some layering artifacts that are visiblemodels. Some methods including barcode and QR codein images obtained by a document scanner. These artifacts enable toembed information in a visible way. Other methods embed it covertly,
measure the thickness of the layers and extract the information.
with additional hidden information that is not perceivable by the user.
The new approach makes it possible to embed various types ofSince 2010, the 3D printing technology has increasingly gained popinformation such as an URL that can be linked to Web services, aularity, leading to a growing interest in the watermarking technology
unique ID that can be used for product tracing, and a printer ID andfor 3D printed objects.
printing date for batch quality managements.
In the study published in IEEE Transactions on Multimedia,
researchers from NAIST have developed a new method to embed
information in a 3D printed object and retrieve it using a consumer
document scanner. Information such as a serial ID can be embedded
without modifying the shape of the object, and simply extracted from
Referencea single image of a commercially available document scanner.
Arnaud Delmotte, Kenichiro Tanaka, Hiroyuki Kubo, Takuya Funatomi,
Several technologies have been developed for 3D printing, includYasuhiro Mukaigawa. 2020. Blind watermarking for 3D printed objects bying the Fused Deposition Modeling (FDM) that is most commonly
locally modifying layer thickness. IEEE Transactions on Multimedia, 22,2780–2791.
used. It consists of deposing layers of molten plastic on top of eachother. The desired shape is obtained by
precisely controlling the position andflow of a printing nozzle such that thedeposed plastic layers have a controlled
path and thickness. The plastic flow
is generally controlled to produce a
constant layer thickness.
In the new method developed by
the NAIST researchers, the plastic
flow is modified during the print tolocally change the layer thickness andembed some additional information.
In order to prevent the degradationof the external surface of the object,pairs of vertically adjacent layers are
selected and the ratio of their respective
thicknesses is modified while keeping
constant the sum of the two layer
thicknesses. Since a standard layer
thickness is about 0.2mm, information
can be embedded in a relatively small
area ranging from several millimeters
to a few centimeters.
To retrieve the embedded information, it is necessary to measure
Scanned images of normal (left) and newly developed printing methods (right).
the thickness of the layers. The new

14

NAIST Research Highlights

Mathematical Informatics
Prof. Kazushi Ikeda

New model can predict a patient’s
responsiveness to fMRI-based
mental health treatment
Researchers at NAIST develop a new machine-learning based model that can forecast a person’s
aptitude for neurofeedback training treatments with a high generalization ability.

Ad v anc e me nt s i n me d i c a lscience have allowed the treatment of psychiatric disorders
like major depressive disorder (MDD)with functional magnetic resonanceimaging neurofeedback (fMRI-NF)
training. fMRI-NF training is a typeof treatment that provides a non-invasive way to control and reinforce
brain functions in patients withmental disorders through the use ofreal-time fMRI monitoring. However,
the effectiveness of the treatment is
not universal—it is influenced by aparameter called neurofeedback (NF)
aptitude.
The high generalization ability of the new neurofeedback (NF) aptitude prediction model offers a quick, simple
NF aptitude refers to an indiand non-invasive method to screen candidates in clinical settings for whom fMRI-NF training would be mostvidual’s capacity to respond to NF
beneficial.
training by displaying changes inbrain activity. But NF aptitude varies
which FCs were most effective for predicting NF aptitude.
from individual to individual. Thus, predicting a patient’s NF aptitude
They found that the PLS model could be generalized to the indebecomes important not only for the success of fMRI-NF training,
pendent
dataset from other institutes, i.e., it could successfully predict
but also to reduce the physical and economic burden on the patienttheNF
aptitudeof individuals based solely on resting-state fMRI
and healthcare system. Thus far, NF aptitude prediction models havescanning.
Theyalso
found that a part of the brain called the posterior
focused on specific target regions in the brain, where the NF training
cingulate
cortexwasthe functional hub among the brain regions,
was focused. Now, in a new study published in NeuroImage, a group ofsuggesting
that
it
plays
a major role in NF aptitude. “We believe that
researchers, led by Junichiro Yoshimoto from NAIST, has successfully
ourresearchwill
help
fMRI-NF
training become more popular as adeveloped a mathematical model for the prediction of NF aptitude
non-invasive
treatment
withminimalside effects for patients with
with a high generalization ability.
mentalhealthdisorders,”concludes
Yoshimoto.
Speaking about their research, Yoshimoto says, “We applied
Even though the study focused on MDD, the generalizability ofmachine learning, which is an offshoot of artificial intelligence (AI)themodel developed in this study ensures that it can be applied totechnology, on data obtained from heathy individuals and patientsdifferent
neuropsychological disorders, providing hope to patientswith major depressive disorder to successfully develop a mathematicalsufferingfrommental illnesses and neurological disorders.
model that can predict individual fMRI-NF training aptitude, basedon their pre-recorded brain activity at the resting state.”
To arrive at the model, the scientists first studied fMRI images ofhealthy patients and patients with MDD before fMRI-NF training.
Reference
They then used these images to calculate the resting state functional
Takashi Nakano, Masahiro Takamura, Haruki Nishimura, Maro Machizawa,
connectivity (FC), which describes the correlated or anti-correlated
Naho Ichikawa, Atsuo Yoshino, Go Okada, Yasumasa Okamoto, Shigeto
activities in different areas of the brain. They then applied a technique
Yamawaki, Makiko Yamada, Tetsuya Suhara, Junichiro Yoshimoto. 2021.
called ‘partial least squares regression’ (PLS) to transform the FC
Resting-state brain activity can predict target-independent aptitude inpatterns into participants’ NF aptitude. Furthermore, they determined
fMRI-neurofeedback training. NeuroImage, 245, 118733.

NAIST Research Highlights

15

Mathematical Informatics

Prof. Kazushi Ikeda

How is the brain
programmed forcomputer programming?
Researchers at NAIST analyze the brain activities of computer programmers of different skill level using fMRI and find that several regions of expert
programmers’ brains are fine-tuned for programming, which may provide
better methods and tools for everyone to learn programming.

Countries around the world are seeing
a surge in the number of computerscience students. Enrolment in related
university programs in the U.S. and Canada
tripled between 2006‒2016 and Europe too
has seen rising numbers. At the same time,
the age to start coding is becoming younger
and younger because governments in manydifferent countries are pushing K-12 computer science education. Despite the increasing
popularity of computer programming, little
is known about how our brains adapt to thisrelatively new activity.
A team of researchers led by NAIST
has examined the brain activity of thirty
programmers of diverse levels of expertise,finding that seven regions of the frontal,
parietal and temporal cortices in expert
programmer's brain are fine-tuned for programming. The finding suggests that higher

16

NAIST Research Highlights

programming skills are built upon fine-tuned
brain activities on a network of multiple
distributed brain regions.
“Many studies have reported differences
between expert and novice programmers inbehavioural performance, knowledge structure and selective attention. What we don’tknow is where in the brain these differences
emerge,” says Takatomi Kubo, one of the leadauthors of the study.
To answer this question, the researchers
observed groups of novices, experienced, andexpert programmers. The programmers wereshown 72 different code snippets while under
the observation of functional MRI (fMRI)
and asked to place each snippet into one
of four functional categories. As expected,
programmers with higher skills were betterat correctly categorizing the snippets. A
subsequent searchlight analysis revealed thatthe amount of information in seven brain
regions strengthened with the skill level ofthe programmer: the bilateral inferior frontal
gyrus pars triangularis (IFG Tri), left inferiorparietal lobule (IPL), left supramarginal gyrus(SMG), left middle and inferior temporal gyri
(MTG/IT), and right middle frontal gyrus(MFG).
“Identifying these characteristics in expert
programmers’ brains offers a good starting
point for understanding the cognitive
mechanisms behind programming expertise.
Our findings illuminate the potential set of
cognitive functions constituting programming expertise,” Kubo says.
More sp ecif ically, the left IFG Tri
and MTG are known to be associatedwith natural language processing and, inparticular, semantic knowledge retrieval in agoal-oriented way. The left IPL and SMG areassociated with episodic memory retrieval.
The right MFG and IFG Tri are functionallyrelated to stimulus-driven attention control.
"Programming is a relatively new activity
in human history and the mechanism islargely unknown. Connecting the activity toother well-known human cognitive functions
will improve our understanding of programming expertise. If we get more comprehensive theory about programming expertise, itwill lead to better methods for learning andteaching computer programming," Kubo
says.
Reference

Yoshiharu Ikutani, Takatomi Kubo, Satoshi Nishida,
Hideaki Hata, Kenichi Matsumoto, Kazushi Ikeda,
Shinji Nishimoto. 2021. Expert programmers havefine-tuned cortical representations of source code.
eNeuro, 8, ENEURO.0405-20.2020.

Overview of the study. Multiple brain regions have fine-tuned representation of source code in proportion
to the behavioral performance of program comprehension based on the fMRI data obtained from 30
subjects with different levels of programming expertise while performing the program categorization
task.
NAIST Research Highlights

17

Computational Behavioral Neuroscience
Associate Prof. Saori Tanaka

Studying the OCD cycle
Researchers at NAIST develop a new model of obsessive-compulsive disorder basedon principles of reinforcement learning, which may lead to better treatment for obsessive-compulsive and related disorders.

A

team of researchers led by NAIST has demonstrated thatobsessive-compulsive disorder (OCD) can be understood asa result of imbalanced learning between reinforcement andpunishment. On the basis of empirical tests of their theoretical model,
they showed that asymmetries in brain calculations that link current
results to past actions can lead to disordered behavior. Specifically,
this can happen when the memory trace signal for past actions decays
differently for good and bad outcomes. In this case, “good” means theresult was better than expected, and “bad” means that it was worse
than expected. This work helps to explain how OCD develops.
OCD is a mental illness involving anxiety, characterized byintrusive and repetitious thoughts, called obsessions, coupled withcertain repeated actions, known as compulsions. Patients with OCD
often feel unable to change behavior even when they know that theobsessions or compulsions are not reasonable. In severe cases, these
may render the person incapable of leading a normal life. Compulsive
behaviors, such as washing hands excessively or repeatedly checking
whether doors are locked before leaving the house, are attempts to
temporarily relieve anxiety caused by obsessions. However, hitherto,
the means by which the cycle of obsessions and compulsions becomes
strengthened was not well understood.
Now, the team has used reinforcement learning theory to modelthe disordered cycle associated with OCD. In this framework, an
outcome that is better than predicted becomes more likely (positive
prediction error), while a result that is worse than expected issuppressed (negative prediction error). In implementation of reinforcement learning, it is also important to consider delays, as well as
positive/negative prediction errors. In general, the outcome of a certain choice is available after a certain delay. Therefore, reinforcement
and punishment should be assigned to recent choices within a certain
time frame. This is called credit assignment, which is implemented
as a memory trace in reinforcement learning theory. Ideally, memory
trace signals for past actions decay at equal speed for both positive
and negative prediction errors. However, this cannot be completely
realized in discrete neural systems. Using simulations, the researchers
found that agents implicitly learn obsessive-compulsive behavior
when the trace decay factor for memory traces of past actions related
to negative prediction errors (ν−) is much smaller than that related
to positive prediction errors (ν+). This means that, from the opposite
perspective, the view of past actions is much narrower for negative
prediction errors than for positive prediction errors. “Our model,
with imbalanced trace decay factors (ν+ > ν−) successfully represents
the vicious circle of obsession and compulsion characteristic of OCD”,
say co-first authors Yuki Sakai and Yutaka Sakai.
To test this prediction, the researchers had 45 patients with OCD
and 168 healthy control subjects play a computer-based game with
monetary rewards and penalties. Patients with OCD showed muchsmaller ν− compared with ν+, as predicted by computational characteristics of OCD. In addition, this imbalanced setting of trace decay
factors (ν+ > ν−) was normalized by serotonin enhancers, which are
first-line medications for treatment of OCD. “Although we think that
we always make rational decisions, our computational model provesthat we sometimes implicitly reinforce maladaptive behaviors,” sayscorresponding author, Saori C. Tanaka.
Although it is currently difficult to identify treatment-resistant patients based upon their clinical symptoms, this computational model
suggests that patients with highly imbalanced trace decay factors may
not respond to behavioral therapy alone. These findings may one
day be used to determine which patients are likely to be resistant tobehavioral therapy before commencement of treatment.

Reference

Memory trace imbalance in reinforcement and punishment systems canreinforce implicit choices leading to obsessive-compulsive behavior.
18

NAIST Research Highlights

Yuki Sakai, Yutaka Sakai, Yoshinari Abe, Jin Narumoto, Saori C. Tanaka.
2022. Memory trace imbalance in reinforcement and punishment systems
can reinforce implicit choices leading to obsessive-compulsive behavior.
Cell Reports, 40, 111275.

BIOLOGICALSCIENCEPlant Developmental Signaling
Prof. Keiji Nakajima

Switching roles:
Key proteins evolved from
activators to maintainers in plants
Researchers at NAIST find that the ancestral role of KNOX and BELL transcription factors inplants is zygote activation, later switching to meristem maintenance during plant evolution.

Sometimes in research, just as in other areas of life, answers tofundamental questions can be sitting in plain sight. Scientists
have discovered a key piece of the puzzle of plant evolution
previously overlooked by plant scientists.
In a study published in eLife, a team of researchers led by NAIST
has revealed that an ancestral function of the plant KNOX/BELL
proteins is activation of the zygote (the first diploid cell formed by thefusion of female and male gametes, also known as reproductive cells),
and this role shifted toward the maintenance of organ development
during the evolution of land plants.
One of the central questions of developmental biology is
how parental genomes mix in a zygote and are activated to begin
diploid development. Two proteins, KNOX and BELL, function astranscription factors—proteins that play an essential role in gene
expression. KNOX and BELL activate diploid development in plants
such as unicellular green alga, but in land plants such as angiosperms(flowering plants) they play a part in the maintenance of the shootmeristem—the tissue that generates the whole of the plant that grows
above ground—and the process of organ formation in the later stagesof diploid development.
“It’s unknown if the differing functions of KNOX and BELL wereattained separately in land plants and algae,” says senior author of thestudy, Keiji Nakajima. “Althoughmost ly dismiss ed f rom t he
spotlight by plant biologists
until now, the zygote-activating
functions of algal KNOX/BELLs,
and how they relate to those ofland plants, were the focus of
this study.”
To i nv e s t i g at e t h i s , t h eresearch team looked at a basal
land plant species, the liverwort
Marchantia polymorpha, which
has recently been recognized
as a good model for studying
the evolution of land plants,
especially in sexual reproduction
research. The team found thatgamete-expressed KNOX andB E L L g e n e s a re n e e d e d t obegin zygote development bypromoting nuclear fusion in the zygote in a way very similar to that
previously found in the green alga Chlamydomonas reinhardtii.
“Our results suggest that the ancestral role of KNOX/BELL
transcription factors is zygote activation,” explains Nakajima. “As land
plants evolved, this moved toward meristem maintenance.”
The results of this study will be important to plant biologists
working across a range of fields such as embryo and organ formation,
sexual reproduction and evolution. These findings, in addition to
the proposal of an equivalent scenario in groups such as fungi andanimals, will also be relevant to researchers in the wider biologicalsciences.

Reference

Tetsuya Hisanaga, Shota Fujimoto, Yihui Cui, Katsutoshi Sato, Ryosuke
Sano, Shohei Yamaoka, Takayuki Kohchi, Frédéric Berger, Keiji Nakajima.
2021. Deep evolutionary origin of gamete-directed zygote activation by
KNOX/BELL transcription factors in green plants. eLife, 10, e57090.

Egg cells of the liverwort Marchantia polymorpha.

20

NAIST Research Highlights

Plant Developmental Signaling

Root cap cell detachment in wild type (left) and autophagy-deficient mutant Arabidopsis thaliana (right).

Prof. Keiji Nakajima

A new role of autophagy in plant
cell differentiation revealed
Researchers at NAIST find that autophagy is required for root cap cell differentiation andseparation in Arabidopsis.

A

midlife career change is hard to pull off, because it caninvolve reinventing yourself and adopting a completely new
professional role. Now, scientists show that some plant cells geta helping hand from autophagy when they switch primary functions
partway through their lifespan.
In a study published in Development, a team of researchers led by
NAIST has revealed that autophagy—self-digesting machinery that
operates inside of eukaryotic cells—plays a crucial role in root growth.
That is, autophagy supports root cap cells transition from the centerof the cap, where they act as gravity sensors, to the outside of the cap,
where they function as secretory cells and eventually slough off.
Root cap cells are unique among plant cells in that they are subject
to constant turnover, while most of the other cells that make up theplant body are maintained throughout the plant's life. As the roots
grow and lengthen, the outermost cells on the root cap slough off, andinner cells take their place, changing function as well as location.
“The concerted cell turnover and functional specialization of root
cap cells require sophisticated subcellular reorganization,” explains
Tatsuaki Goh, lead author of the study. “However, the spatiotemporal
dynamics of this reorganization and the molecular and genetic
regulation of this process are not well characterized.”
To investigate this process of subcellular reorganization, theresearchers designed a microscopy technique that allowed them towatch Arabidopsis roots grow in real time while automatically tracking
the tip of the extending root. “What we saw is that, right before theoutermost layer of cells sloughed off of the root tip, there was a major
shift in the location of organelles within the cells,” says Keiji Nakajima,senior author of the study. “Importantly, the autophagy system needed
to be intact for this shift to take place.”
Having a functional autophagy system was also required for thecells to separate from the root caps in a highly ordered manner. “Ourfindings suggest that autophagy is an essential component of cell
clearance and organelle rearrangement. These are both associatedwith the functional specialization of the outermost root cap cells as
secretory cells, as well as with the cell separation process,” says Goh.
Autophagy is an evolutionarily conserved process in eukaryotic
cells, where it plays key roles in nutrient recycling and the stressresponse, although its role is less well understood in plants. Thus,
the novel role for autophagy in plants revealed by this study has
important implications for the conservation of core autophagy factors
in plant genomes.

Reference

Tatsuaki Goh, Kaoru Sakamoto, Pengfei Wang, Saki Kozono, Koki Ueno,
Shunsuke Miyashima, Koichi Toyokura, Hidehiro Fukaki, Byung-Ho
Kang, Keiji Nakajima. 2022. Autophagy promotes organelle clearance andorganized cell separation of living root cap cells in Arabidopsis thaliana.
Development, 149, dev200593.

NAIST Research Highlights

21

Plant Stem Cell Regulation and Floral Patterning
Prof. Toshiro Ito

How to beat the heat:
Memory mechanism allows plants
to adapt to heat stressResearchers at NAIST find that plants adapt to heat stress via an epigenetic memory mechanism,
where JUMONJI proteins control small heat shock genes.

“If you can’t stand the heat, get out of the kitchen,” as the oldsaying goes. But for organisms that can’t leave the proverbial
kitchen when things get too hot, there’s another way: scientists
have discovered that plants can gain heat tolerance to better adapt tofuture heat stress, thanks to a particular mechanism for heat stress‘memory’.
In a study published in Nature Communications, a team ofresearchers led by NAIST has revealed that a family of proteins that
control small heat shock genes enables plants to ‘remember’ how todeal with heat stress.
Climate change, especially global warming, is a growing threat toagriculture worldwide. Because plants can’t move to avoid adverse
conditions, such as potentially lethal high temperatures, they need to
be able to deal with factors such as heat stress effectively to survive.
Therefore, improving the heat tolerance of crop plants is an important
goal in agriculture.
“Heat stress is often repeating and changing,” says lead author of
the study Nobutoshi Yamaguchi. “Once plants have undergone mildheat stress, they become tolerant and can adapt to further heat stress.
This is referred to as heat stress ‘memory’ and has been reported to becorrelated to epigenetic modifications.” Epigenetic modifications are
inheritable changes in the way genes are expressed, and do not involve
changes in the underlying DNA sequences.
“We wanted to discover how plants retain a memory of environmental changes,” explains Toshiro Ito, senior author. “We examined
the role of JUMONJI (JMJ) proteins in acquired temperature toler-

Removal of H3K27me3 in response to heat.
22

NAIST Research Highlights

ance in response to recurring heat within a few days.”JUMONJI proteins are histone demethylases. Demethylases
are enzymes that remove methyl groups from molecules such asproteins, particularly histones, which provide structural support tochromosomes. The team revealed that plants are able to maintainheat memory because of lowered H3K27me3 (histone H3 lysine 27
trimethylation) on small heat shock genes.
“We found that these proteins are necessary for heat acclimationin Arabidopsis thaliana. These results, along with future studies, willfurther clarify the mechanisms of plant memory and adaptation,” says
Yamaguchi.
This research will be relevant to genetic research in a number offields, including biology, biochemistry, ecology, and environmental
and agricultural sciences, and is applicable to the study of animals
as well as plants. Understanding the epigenetic memory mechanism
revealed in this study will help in working with heat tolerance tomaintain the food supply in natural conditions.

Reference

Nobutoshi Yamaguchi, Satoshi Matsubara, Kaori Yoshimizu, Motohide
Seki, Kouta Hamada, Mari Kamitani, Yuko Kurita, Yasuyuki Nomura,
Kota Nagashima, Soichi Inagaki, Takamasa Suzuki, Eng-Seng Gan, Taiko
To, Tetsuji Kakutani, Atsushi J. Nagano, Akiko Satake, Toshiro Ito. 2021.
H3K27me3 demethylases alter HSP22 and HSP17.6C expression in
response to recurring heat in Arabidopsis. Nature Communications, 12, 3480.

Plant Stem Cell Regulation and Floral Patterning
Prof. Toshiro Ito

The flower clock:
How a small protein helps flowers
to develop right and on time
Researchers at NAIST find that KNUCKLES, a small multi-functional protein, supports thecorrect timing of floral development for the proper formation of flower reproductive organs.

How flowers form properly within a limited timeframe has been a mystery, at least until now.
scientists have discovered how a multi-tasking
protein helps flowers to develop as expected.
In a study published in Proceedings of the National
Academy of Sciences U.S.A., researchers have revealed
that a small protein plays multiple roles to ensure thatfloral reproductive organs are formed properly within ashort space of time.
Flowers develop from floral meristems, which
differentiate to produce the sepals, petals, stamens, andcarpels. The proper development of these floral organs
depends on meristem development being completed
within a certain time period. In the early stages offlower development, stem cells provide the cell source
for floral organ formation. In floral meristems, stem cell
activities are maintained via a feedback loop between
WUSCHEL (WUS), a gene that identifies floral stem
cells, and CLAVATA3 (CLV3), a stem cell marker genethat is activated and sustained by WUS.
“A small protein called KNUCKLES (KNU)
represses WUS directly, which leads to the completionof floral stem cell activity at the right time,” says leadauthor of the study Erlei Shang. “What isn’t fully
understood is how the robust floral stem cell activity
finishes within a limited time period to ensure carpel
development.”
KNUCKLES expression in floral meristem of Arabidopsis thaliana.
“The team’s research revealed that in Arabidopsis
thaliana, KNU can completely deactivate the robust
within a short time window, and ensures that flower reproductive
floral meristems at a particular floral stage, thanks to the multiple
organs are properly formed,” says corresponding author Bo Sun.
functions that KNU carries out via its position-specific roles,” says
The results of this research will be useful for genetic studies ofsenior author Toshiro Ito at NAIST.
food crop species such as rice, tomatoes, and maize. An understandKNU both represses and silences WUS, and directly represses
ing of the floral meristem termination mechanism discovered in this
CLV3 and CLV1 (a gene that encodes a receptor for the CLV3
study will benefit crop yields for food production globally.
peptide). Consequently, KNU eliminates the CLV3-WUS feedback
loop via transcriptional and epigenetic mechanisms (i.e., those that do
not involve changes in the underlying DNA sequences). Additionally,KNU interacts physically with the WUS protein, which inhibits WUS
Referencefrom sustaining CLV3, disrupting interactions that are required for
Erlei Shang, Xin Wang, Tinghan Li, Fengfei Guo, Toshiro Ito, Bo Sun.
the maintenance of floral meristems.
2021. Robust control of floral meristem determinacy by position-specific“Our results reveal a regulatory pathway where KNU plays a key
multifunctions of KNUCKLES. Proceedings of the National Academy ofrole in supporting the completion of floral meristem development
Sciences U.S.A., 118, e2102826118.
NAIST Research Highlights

23

Plant Physiology
Prof. Motomu Endo

The circadian clock
makes sure plant cells havethe time of their lives
Researchers at NAIST find that the circadian clock regulates cell cycle
progression and cell differentiation in Arabidopsis, using a novel algorithm
to reconstruct actual-time gene expression patterns from single-cell RNAseq
datasets.

T

hey say timing is everything, and that couldn’t be moretrue for cell cycle progression and differentiation. Now,
a team of researchers led by NAIST has found that thecircadian clock is crucial for proper plant development. In a study
published in Cell Reports, they showed that the circadian clock
plays a guiding role in plant cell differentiation.
The circadian clock is involved in both cell-cycle progression
and cell fate transitions. The involvement of circadian clocks inthe process of differentiation has been shown in many multicellular organisms; however, how plant circadian clocks regulate celldifferentiation remains unclear.
“Elucidating how the circadian clock is involved in celldifferentiation is important to understand the basis of cell fate determination,” explains Motomu Endo, senior author of the study.
“However, this has been difficult to investigate in plants because it
is challenging to isolate single plants’ cells, and existing analyticalmethods rely on “pseudo-time” analysis that does not accuratelyreflect normal circadian rhythms.”
To address these challenges, the researchers used tiny glass
tubes to isolate individual cells from developing plants and analyzed the expression of various genes related to circadian rhythms
and cell differentiation in each cell. They then developed a newalgorithm called PeakMatch to reconstruct actual-time gene
expression patterns from the single-cell datasets.
“Using this powerful approach, we were able to show thatthe expression profile of clock genes is changed prior to cell

24

NAIST Research Highlights

differentiation,” states Endo. “Specifically, in early differentiating
cells, the induction of the clock gene LUX ARRYTHMO directly
targets genes involved in cell-cycle progression to regulate celldifferentiation.”
Further investigation showed that large-scale changes in
the circadian clock profile in undifferentiated cells induce theexpression of the clock gene LUX, which directly triggers celldifferentiation. “Taken together, our results show that the plantcircadian clock plays a guiding role in cell differentiation,” says
Endo. “Importantly, our study also provides an approach fortime-series analysis at single-cell resolution.”
Because the development of circadian rhythms during celldifferentiation is observed in animals as well as in plants, thefinding that clock genes directly regulate cell fate determinationand cell division may help understand how cell differentiationis controlled in multicellular organisms. The newly developed
PeakMatch algorithm can also be applied to all kinds of single-celltranscriptomes in other organisms.
Reference

Kotaro Torii, Keisuke Inoue, Keita Bekki, Kazuya Haraguchi, Minoru
Kubo, Yuki Kondo, Takamasa Suzuki, Akane Kubota, Kyohei Uemoto,
Hanako Shimizu, Masato Saito, Hiroo Fukuda, Takashi Araki, Motomu
Endo. 2022. A guiding role of the Arabidopsis circadian clock in celldifferentiation revealed by time-series single-cell RNA sequencing. Cell
Reports, 40, 111059.

NAIST Research Highlights

25

Plant Symbiosis

Prof. Satoko Yoshida

When plants attack:Parasitic plants use ethylene
as a host invasion signal
Researchers at NAIST find that the plant hormone ethylene mediates theinvasion of hosts by parasitic plants, which could provide new ways tocontrol a range of parasitic weeds.

Mutants that reveal the secrets of
how plants attack? No, it’s not ascene from a science fiction movie,
but you could be forgiven for thinking that.
Instead, it’s a scene from real life. A team ofresearchers led by NAIST has reported in a
new study in Science Advances that parasitic
plants use the plant hormone ethylene as asignal to invade the roots of host plants.
To develop a successful parasitic relationship, parasitic plants form a specialized
structure, the haustorium which attaches toand invades the host plant. The formationof haustoria is regulated by signal molecules
derived from the host plant and allows the26

NAIST Research Highlights

parasitic plant to absorb water, nutrients, andsmall materials from the host plant.
“To understand the genetic programsfor haustorium development, we identified
mutants that displayed haustorial defects onhost invasion,” says lead author of the study
Songkui Cui. “Genome sequencing showedthat these mutants have defective ethylene
signaling, and it turned out that ethylene
signaling genes are crucial for the parasitic
plant to infect its host plant.”
Ethylene is a gaseous plant hormone thatis involved in fruit ripening, aging of leaves,and the formation of root nodules. Ethylene
is also widely involved in plant interactions

with viruses and numerous organisms, suchas insects and bacteria, lending either resistance or susceptibility to plants depending on
the types of pathogens.
“Our results indicate that ethylene mediates host recognition in parasitic plants forhost invasion,” explains project leader Satoko
Yoshida. “This is the first time that the mediation of host invasion by parasitic plant genes
has been identified via forward genetics. Ourfindings offer a new understanding of how
a parasitic plant uses the ethylene molecule
to tweak haustorium development and hostinvasion.”
Forward genetics is used to identify genes,

or sets of genes, that produce a particularcharacteristic in an organism. The model
species used in this study is from a familyof parasitic plants that includes destructive
weeds. But the molecular basis for theirparasitism has been largely unexplored until
now.
“Our results suggest that parasitic plants
have taken over ethylene signaling for parasitism at multiple stages of their life cycle,
such as germination, haustorium growth termination, and host invasion. This knowledge
could provide new ways to use ethylene andethylene inhibitors to control a broader rangeof parasitic weeds, including those that don’t

rely entirely on hosts to complete their life
cycle, by manipulating haustorial function,”says Cui.

Reference

Songkui Cui, Tomoya Kubota, Tomoaki Nishiyama,
Juliane K. Ishida, Shuji Shigenobu, Tomoko F. Shibata,Atsushi Toyoda, Mitsuyasu Hasebe, Ken Shirasu,
Satoko Yoshida. 2020. Ethylene signaling mediates
host invasion by parasitic plants. Science Advances, 6,
eabc2385.

The model parasitic plant, Phtheirospermum
japonicum, and its haustorium. The rightlower photo shows xy lem conne ction
between the parasitic (P) and host plants (H).
NAIST Research Highlights

27

Plant Growth Regulation
Prof. Masaaki Umeda

Damage control:
Plants juggle genome maintenanceand growth by being organized
Researchers at NAIST find that control of the plant hormones cytokinin and auxin help damaged
plants to minimize cell death while maintaining their genomes and organ development.

Humans pride themselves on being able tomultitask, especially under pressure. But
it turns out that we aren’t the only ones
who are organized: scientists have discovered thatplants balance genome maintenance with organ
growth by organizing different responses to DNA
damage.
In a study published in Science Advances, ateam of researchers led by NAIST has revealed
that plants use combined control of the plant
hormones cytokinin and auxin to organize DNA
damage responses while maintaining growth.
Plants are highly adaptable organisms thatnever stop growing, thanks largely to the functions
carried out by their roots. Because of the essential
role that roots play in plant growth, root development is highly responsive to fluctuating conditions
underground, including environmental stresses.
These stresses tend to limit root growth, and candamage DNA.
“Plants actively respond to DNA damage,
Arabidopsis root tips with increased cytokinin (left) and decreased auxin signals (right) inand these responses are controlled by the cell
response to DNA damage.
cycle checkpoint mechanism,” says project leader
Masaaki Umeda. “The cell cycle is stopped at a
The study explains previously unidentified mechanisms of keeping
certain stage so that DNA can be repaired, or in extreme cases, cell
genomesstable in a developmental setting, which allow continuous
death initiated. What we wanted to know was how the DNA damage
organ
growthin fluctuating environments. The results of this study
responses are organized to allow continuous root growth.”
will
have
wide-ranging
applications to research on agricultural techUnlike animal cells, plant cells aren’t mobile, so plants can’t replacenologies,
particularlyfor
plant growth and development in extreme
dead cells with surrounding living cells. To simultaneously maintainenvironments.
Futurestudies
will illuminate how plants develop
their genomes and allow organ development, plants initiate different
organsbeyondtheembryonic
stage
via the maintenance of genome
DNA damage responses that are dependent on the different types ofstability
inchanging
conditions.
cells in tissues.
“We found that the combined control of two plant hormones, cytokinin and auxin, organizes different DNA damage responses at thecost of minimized cell death,” explains first author Naoki Takahashi.
“This is a sophisticated way to retain stem cells while maintaining
Reference
their genomes in damaged plants.”
Naoki Takahashi, Soichi Inagaki, Kohei Nishimura, Hitoshi Sakakibara,
A decrease in auxin triggers stem cell death and stops the cell
Ioanna Antoniadi, Michal Karady, Karin Ljung, Masaaki Umeda. 2021.
cycle at the G2 phase, while increased cytokinin signals support early
Alterations in hormonal signals spatially coordinate distinct responses
endoreplication—a cell cycle variant where the genome is duplicatedto DNA double-strand breaks in Arabidopsis roots. Science Advances, 7,
without cell division.
eabg0993.

28

NAIST Research Highlights

Functional Genomics and Medicine
Associate Prof. Yasumasa Ishida

A single allele deletion in geneencoding Zbtb38 leads to early
embryonic deathResearchers at NAIST find that heterozygous loss in gene encoding methyl-CpG binding
protein Zbtb38 leads to early embryonic death via the suppression of transcription
factors Nanog and Sox2.

Dfurther research to find out how Zbtb38 works during embryogenesis.
NA methylation is a major epigenetic modification that isMore research needs to be done to elucidate the specific molecularcrucial for mammalian development. For instance, DNAmechanisms. Zbtb38 is found in all tissues, and it is linked to height,
methylation is central to inexhaustible biological processes,
cancers, neurodegenerative diseases and rheumatoid arthritis, etc.
such as gene regulation and cell fate decisions. In mammals, DNA
Thus, the creation and analysis of tissue-specific Cre-mediated knockmethyltransferases are key for blastocysts to re-establish global DNA
out mice will help us understand Zbtb38's physiological functions andmethylation patterns during implantation. This is critical for passing
Zbtb38-linked diseases,” concludes Matsuda.
on epigenetic information to the next generation. On the other hand,
The findings of this work will interest developmental biologists as
the role of methyl-CpG binding proteins (MBPs) that bind methylit emphasizes the epigenetic significance of DNA methylation during
ated CpG as part of the DNA methylation processes is still unclear.
the early stages of pregnancy.
However, a previous study conducted by researchers at NAIST
clarified that; Zbtb38, also known as CIBZ, is a zinc finger type of
MBP that is pivotal for the growth of mouse embryonic stem (ES)cells. They further demonstrated that Zbtb38 facilitates the expressionof Nanog, which is fundamental for the growth of ES cells. However,what Zbtb38 does in real life, is still a mystery.
Reference
In a further quest to solve this mystery, the same scientists at
Miki Nishio, Takuya Matsuura, Shunya Hibi, Shiomi Ohta, Chio Oka,NAIST, led by Eishou Matsuda, used Cre-loxP technology to make
Noriaki Sasai, Yasumasa Ishida, Eishou Matsuda. 2022. Heterozygous lossof Zbtb38 leads to early embryonic lethality via the suppression of Nanog
conditional Zbtb38 knockout mice. Their ground-breaking researchand Sox2 expression. Cell Proliferation, 55, e13215.
revealed that a single Zbtb38 allele deletion in the germline led to adecrease in epiblast cell growth and an increase
in apoptosis soon after implantation, which led toearly embryonic death. Nanog, Sox2 and genes that
control epiblast growth and differentiation became
dysfunctional when Zbtb38 was lost in heterozygous
embryos.
“Our findings indicate that germline lossof the Zbtb38 single allele reduces epiblast cellproliferation and increases apoptosis shortly
after implantation, resulting in early embryoniclethality. Heterozygous Zbtb38 deficiency reduced
the expression of Nanog, Sox2 and genes involvedin epiblast proliferation, differentiation and cell
viability. This finding shows that a methyl-CpG
binding protein has a role in controlling embryonicphenotype,” explains Matsuda.
“For the first time we demonstrated a link to anembryonic function for a protein that has long been
known to bind methyl-CpG,” says co-author YasuHeterozygous loss of Zbtb38 causes decreased epiblast cell proliferation and increased apoptosis
shortly after implantation, leading to early embryonic lethality.
masa Ishida. “This presents a huge opportunity for

NAIST Research Highlights

29

Molecular Immunobiology
Prof. Taro Kawai

Tropical ginger treatment for
blocking inflammation
Researchers at NAIST identify antioxidant properties of a ginger-derived compound that may
help fight inflammatory diseases.

M

The researchers grew immune cells in culture that were obtained
any natural compounds have various anti-inflammatory andfrommouse bone marrow, and also used a mouse model of colitis.
other beneficial properties that humans have been utilizing
ACA
was added to the growing cells and the live mice were given
for medicinal purposes for hundreds of years. However,the compound in their food. The researchers then examined thethe specific molecular mechanisms behind these health-promoting
effects on ROS production, secretion of IL-1β, and other markers ofeffects are not always clear. One such compound is 1’-acetoxychavicol
inflammation.
acetate, or ACA, which comes from the tropical ginger Alpinia plant.
“Cells treated with ACA had significantly reduced IL-1β
Now, a team of researchers led by NAIST has identified how ACA canproduction, as well as lower levels of ROS,” explains Taro Kawai,
help in the treatment of inflammatory diseases.
senior author. “ACA could also
In a report published ininhibit NLRP3 inflammasome
International Immunology, they
activation in the colitis mousefound that ACA attenuatesmodel.” These in vivo results
mitochondrial damage throughare promising, as they suggest
de c re asing m ito chond r i a l
ACA has the potential to treat
reactive oxygen species (ROS),
or prevent the development
blocking activation of a crucial
of inflammatory diseases. “Inprotein complex known as theterestingly, we did not observe
nucleotide-binding oligomerhigh levels of immune cellization domain-like receptor
death when using ACA, whichfamily pyrin domain-containmeans that it may be relatively
ing protein 3 (NLRP3) inflamsafe,” continues Ori.
masome. Many inflammatory
This work provides novel
diseases, like inflammatory
evidence for a specific molecbowel disease, display improperular mechanism governingand chronic activation of thisthe previously obser ved
complex.
anti-inflammatory properties
Previous work has suggestedo f AC A . F u r t h e r m o r e , i tthat the NLRP3 inflammasome
highlights the potential of ACA
pl ays a signif icant role infor therapeutic use in diseases
promoting inflammation by
mediated by IL-1β molecules,
secreting a molecule called ILor associated with cytokine
1β. This acts as a messenger that
ACA ameliorates mitochondrial damage, leading to the suppression ofstorm occurrence, as seen in
recruits various immune cells
NLRP3-inflammasome activity and subsequent IL-1b release.
patients suffering from severe
to the site of injury or infection.
COVID-19.
Additional studies described
how production of ROS can help trigger activation of the NLRP3
inflammasome. Because other groups showed that ACA can reduce
ROS production in certain immune cells, the researchers became
curious how this compound would impact the NLRP3 inflammasome
and its functions.
“Many disease pathogeneses involve dysregulation of the inflamReference
masome,” says Daisuke Ori, co-lead author of the study. “Blood cells
Sophia P. M. Sok, Daisuke Ori, Ayana Wada, Haruna Okude, Takumi
from people suffering from rheumatoid arthritis or other autoimmune
Kawasaki, Masatoshi Momota, Noor Hasima Nagoor, Taro Kawai. 2021.
disorders frequently have increased levels of inflammasome-derived
1’-acetoxychavicol acetate inhibits NLRP3-dependent inflammasome
IL-1β. Therefore, targeting the NLRP3 inflammasome with a comactivation via mitochondrial ROS suppression. International Immunology,
33, 373‒386.
pound like ACA may be a promising therapeutic strategy.”30

NAIST Research Highlights

Molecular Immunobiology
Prof. Taro Kawai

Alveolar macrophages help
CD8+ T cells go (anti-)viral
Researchers at NAIST find that lung-resident alveolar macrophages act as antigen-presenting cells
to prime CD8+ T cell expansion in the lungs in response to viral infection.

The human immune system is a highly complex
network of cells, signals, and responses thatis tightly regulated to ensure that the bodycan fight off infection without damaging its own
tissues. Now, scientists report a new way in which
the immune system protects lung tissue from viral
infections.
In a study published in Cell Reports, a teamof researchers led by NAIST has revealed that
antigen-specific killer T cells (CD8 + T cells)
rapidly expand in the lungs when they encounter
antigen-presenting alveolar macrophages (AMs) toprotect against viral infection.
CD8+ T cells confer protective immunity against
infection with respiratory viruses, such as influenza
A virus (IAV) and severe acute respiratory syndrome
coronavirus 2 (SARS-CoV-2), by killing infected
cells. In order to target the correct cells for killing,
naive CD8+ T cells must be primed by contact with
antigen-presenting cells (APCs), which mediate
the uptake of virus-infected cells and present theirantigens, in a process known as cross-presentation.
The primed CD8+ T cells then clonally expand anddifferentiate into effector or long-lived antigen-speInfluenza virus-specific CD8+ T cells induced in lymph nodes as a result of influenza virus
cific memory T cells.
+
infection or vaccination circulate throughout the body.
“Multiple cell types can present antigen to CD8 T
cells in the lungs, although the role of tissue-resident
macrophages in this process is unclear,” explains
may improve the efficacy of CD8+ T cell-dependent cellular immuniTakumi Kawasaki, lead author of the study. “AMs are the first cells inty,” says Kawai.
the lungs that encounter infectious materials, environmental particles,
Given that the lung is a major tissue for IAV and SARS-CoV-2
surfactants, and dying cells, and they are important for the host deinfection, the findings from this study regarding the mechanism
fense against bacterial and fungal infection, so we suspected that theyof lung-resident memory CD8+ cell expansion are expected to lead
were also important in protecting against respiratory virus infection.”to the development of new vaccines that induce cellular immunity.
To test this, the researchers explored the mechanisms by which
Virus-specific antigen-presenting AMs could be delivered as a type of
APCs instruct antigen-specific CD8+ T cells in the lungs. First, mice“cell transplant vaccine” in the future.
were primed by vaccination with a specific antigen or infection with
IAV, and then they were subjected to secondary immunization or
re-infection. “We determined that antigen-presenting AMs present
inhaled antigen to memory CD8+ T cells,” says senior author of thestudy, Taro Kawai, “and that this resulted in a rapid expansion ofReference
antigen-specific CD8+ T cells in the lungs.”
Takumi Kawasaki, Moe Ikegawa, Kosuke Yunoki, Hifumi Otani, Daisuke
Furthermore, the researchers found that AMs help to develop
Ori, Ken J Ishii, Etsushi Kuroda, Shiki Takamura, Masahiro Kitabatake,resident memory-type cell population by producing interleukin 18.
Toshihiro Ito, Ayako Isotani, Taro Kawai. 2022. Alveolar macrophages
Importantly, administration of antigen-loaded AMs to mice induced
instruct CD8+ T cell expansion by antigen cross-presentation in lung. Cell
Reports, 41, 111828.
the proliferation of resident memory-type CD8+ T cells. “This strategy
NAIST Research Highlights

31

Molecular Medicine and Cell Biology
Prof. Shiro Suetsugu

How cellular fingertips may help
cells "speak" to each otherResearchers at NAIST broaden the known functions of an under-appreciated cell structure, withpossible applications in wound closure and cancer therapy.

W

hat if you found out that you could heal using only
a finger? It sounds like science fiction, reminiscentof the 1982 movie E.T. Well, it turns out that yourbody's own cells can do something similarly unexpected. A
team of researchers led by NAIST has reported in Developmental Cell a means by which cells may use "fingers" to communicate instructions for wound closure.
NAIST project leader Shiro Suetsugu has devoted his career
to studying how cells shape themselves, initiate and accept
communication among one other. An under-appreciatedmeans of doing so is through filopodia, small finger-likecellular projections that are more commonly known to help
certain cells crawl in the body. "Filopodia are well-recognized
as cellular locomotion machinery. Less understood is how
filopodia help cells communicate, and the molecular details of
how this is done," says Suetsugu.
A focus of this line of research should be the proteins
known by the acronym I-BAR. I-BAR proteins are well-known
to help bend the plasma membrane, the "skin" of many cells,
for filopodia formation and thus facilitate movement. "We
identified an I-BAR protein that severs filopodia," says Suetsugu. An important element of this scission may be mechanicalforce, a stimulus that your body commonly applies to cells.
"Laser experiments showed that the force required for scission
is approximately 8‒20 kilopascals. These forces are similar to
the 4‒13 kilopascals, experienced by cells in blood capillaries,"
Suetsugu says.
Severed filopodia go on to form structures called extracelThe lattice light sheet microscopic images of the filopodia by expressing the I-BAR
lular vesicles, a popular research topic in biology. Extracellulardomain protein MIM.
vesicles were used to basically be considered the trash bags ofcells, used for disposing cellular waste. However, the vesicles
BAR proteins are also pertinent to cell locomotion. By learning moreare now considered to be communication packets rather than waste
about how these proteins aid cell-cell communication, we may findbags. "The pertinence of these vesicles to cancer metastasis has piqued
better ways to stop cancer cells from spreading," he says.
researchers' and clinicians' interest," notes Suetsugu.
What does this have to do with cell-cell communication? A
simulated cell-scale wound healed faster when it was treated withfilopodia-derived extracellular vesicles than if untreated. In otherwords, an I-BAR protein first induced filopodia scission and vesicle
production. These vesicles then sent cellular signals that promoted
cell migration toward one another, in a way that may promote woundclosure.
By understanding how cells fully use their molecular machinery tosend instructions to other cells, Suetsugu is optimistic that medical
practitioners will develop new means to safely treat cancer and otherdiseases. "Certain BAR proteins are pertinent to cancer cell biology.

32

NAIST Research Highlights

Reference

Tamako Nishimura, Takuya Oyama, Hooi Ting Hu, Toshifumi Fujioka,
Kyoko Hanawa-Suetsugu, Kazutaka Ikeda, Sohei Yamada, Hiroki Kawana,
Daisuke Saigusa, Hiroki Ikeda, Rie Kurata, Kayoko Oono-Yakura, Manabu
Kitamata, Kazuki Kida, Tomoya Hikita, Kiyohito Mizutani, Kazuma
Yasuhara, Yuko Mimori-Kiyosue, Chitose Oneyama, Kazuki Kurimoto,
Yoichiroh Hosokawa, Junken Aoki, Yoshimi Takai, Makoto Arita, Shiro
Suetsugu. 2021. Filopodium-derived vesicles produced by MIM enhance
the migration of recipient cells. Developmental Cell, 56, 842‒859.

Molecular Medicine and Cell Biology
Prof. Shiro Suetsugu

AI knows where your proteins go
Researchers at NAIST find that machine learning can predict the subcellular locations offunctionally related proteins.

F

acial recognition software can be used to spot a face in a crowd;
but what if it could also predict where someone else was in
the same crowd? While this may sound like science fiction,scientists have now shown that artificial intelligence can accomplish
something very similar on a cellular level.
In a study published in Frontiers in Cell and Developmental Biology,
researchers from NAIST have revealed that a machine learning
program can accurately predict the location of proteins related toactin, an important part of the cellular skeleton, based on the locationof actin itself.
Actin plays a key role in providing shape and structure to cells,
and during cell movement helps form lamellipodia, which are fanshaped structures that cells use to “walk” forwards. Lamellipodia also
contain a host of other proteins that bind to actin to help maintain the
fan-like structure and keep the cells moving.
“While artificial intelligence has been used previously to predictthe direction of cell migration based on a sequence of images, so
far it has not been used to predict protein localization,” says leadauthor of the study, Shiro Suetsugu. This idea came in during thediscussion with Yoshinobu Sato at the Data Science Center in NAIST.
“We therefore sought to design a machine learning algorithm thatcan determine where proteins will appear in the cell based on theirrelationship with other proteins.”
To do this, the researchers trained an artificial intelligence system
to predict where actin-associated proteins would be in the cell byshowing it pictures of cells in which the proteins were labeled withfluorescent markers to show where they were located. Then, they gave

the program pictures in which only actin was labeled and asked it totell them where the associated proteins were.
“When we compared the predicted images to the actual images,
there was a considerable degree of similarity,” states Suetsugu. “Ourprogram accurately predicted the localization of three actin-associated proteins within lamellipodia; and, in the case of one of these
proteins, in other structures within the cell.”On the other hand, when the researchers asked the program topredict where tubulin, which is not directly related to actin, would bein the cell, the program did not perform nearly as well. “Our findings
suggest that machine learning can be used to accurately predict thelocation of functionally related proteins and describe the physicalrelationships between them,” says Suetsugu.
Given that lamellipodia are not always easy for non-experts tospot, the program developed in this study could be used to quickly
and accurately identify these structures from cell images in the
future. In addition, this approach could potentially be used as a sortof artificial cell staining method to avoid the limitations of current
cell-staining methods.

Reference

Kei Shigene, Yuta Hiasa, Yoshito Otake, Mazen Soufi, Suphamon
Janewanthanakul, Tamako Nishimura, Yoshinobu Sato, Shiro Suetsugu.
2021. Translation of cellular protein localization using convolutional
networks. Frontiers in Cell and Developmental Biology, 9, 635231.

An example of a generated image of focal adhesion protein (center), which anchors actin
filaments, from an image of actin filaments (left).

NAIST Research Highlights

33

Applied Stress Microbiology

Prof. Hiroshi Takagi

Japanese sake:
The new pick-me-up?
Yeast strain makes
fatigue-fighting ornithine
Researchers at NAIST find that a mutant strain of sake yeast produces highlevels of the amino acid ornithine, which could be applied to brewing sake, a
traditional Japanese alcoholic beverage, as well as wine and beer.

34

NAIST Research Highlights

F

ans of sake, the traditional Japanese
alcoholic beverage, may have evenmore reason to enjoy it now: Japanesescientists have discovered that a mutant
strain of sake yeast produces high levels of
the amino acid ornithine.
In a study published this month in Metabolic Engineering, a team of researchers led by
NAIST has revealed that a mutant strain of
sake yeast produces 10 times the amount of
the amino acid ornithine compared with theparent yeast strain.
Ornithine is a non-protein-making aminoacid and a precursor to two amino acids—
arginine and proline. It has been found to
perform several physiological functions,such as reducing fatigue and improving sleep
quality.
“We wanted to obtain sake yeast strains
with improved ethanol tolerance,” says
first author of this study, Masataka Ohashi.
“During sake fermentation, the yeast is
exposed to high concentrations of ethanol,
which impedes yeast cell growth, viability andfermentation. Increased ethanol tolerancein sake yeast strains could improve ethanol
production and reduce fermentation time.”
To find ethanol-tolerant yeast strains,
the researchers isolated mutants that
accumulated proline, which can alleviate
ethanol toxicity, using a conventional mutagenesis (i.e., one that doesn’t involve genetic
modification). They also conducted wholegenome sequencing analysis, and performed
brewing tests with sake yeast strains. Thenthey identified and analyzed a new mutation
in a gene that encodes a variant of N-acetyl
glutamate kinase that increases intracellularornithine level.
“We previously constructed self-cloning
industrial yeast strains that accumulate
proline to increase ethanol tolerance andproductivity of yeast,” explains Hiroshi Takagi, corresponding author. “But those yeasts
have not been yet acceptable to consumers
because they’re considered to be geneticallymodified, even though a self-cloning yeast
has no foreign genes or DNA sequences—
they only have yeast DNA.”The researchers successfully isolated
non-genetically mo dif ied ye asts t hat
produced 10 times the amount of ornithinecompared with the parent strain, which is
widely used in Japanese sake breweries, andthe sake brewed with them contained 4–5
times more ornithine.
Takagi notes “ There are two major
purposes for breeding of industrial yeast:
improvement of fermentation ability withenhanced tolerance to environmental stresses
during fermentation processes and diversityof product taste and flavor with modified
metabolic pathways. In yeast, amino acidmetabolisms vary under different growthenvironments and the metabolic styles form
a complicated but robust network. The
elucidation of metabolic regulatory mechanisms and physiological roles for aminoacids is important fundamental research forunderstanding life phenomenon. The yeast
is reliable and safe in food production, andthus the development of novel strains that
overproduce ‘functional amino acids’ such as
ornithine, proline and branched-amino acids,
would greatly contribute to food-relatedindustries.”
The results of this study will contributeto the development of improved yeast strains
for production of high levels of ornithine,
and the strain obtained in this study could
be readily applied to sake, wine, and beer
brewing. Ornithine-accumulating yeast
strains could also be used in the productionof ornithine-rich dietary supplements madefrom these yeasts and their products.
Reference

Masataka Ohashi, Ryo Nasuno, Shota Isogai,
Hiroshi Takagi. 2020. High-level production ofornithine by expression of the feedback inhibitioninsensitive N-acetyl glutamate kinase in the sake yeast
Saccharomyces cerevisiae. Metabolic Engineering, 62,
1–9.

Overview of amino acid metabolism in yeast.
NAIST Research Highlights

35

Environmental Microbiology
Prof. Shosuke Yoshida

How a bacterium
may help solve the plastic
pollution crisis

Researchers at NAIST find that the bacterium Ideonella sakaiensis can
not only degrade environmentally problematic petroleum-based plastics but
can also sustainably produce biodegradable plastics, which has promising
environmental implications.

P

lastic pollution is one of the mostpressing environmental issues of ourtime. The accumulation of petroleum-based plastics is having devastating
effects on our environment, wildlife, andhuman health. In a study published in
Scientific Reports, a team of researchers led
by NAIST has revealed a bacterium that is
not only able to degrade difficult-to-recycle
petroleum-based plastics but can alsosustainably produce more environmentallyfriendly biodegradable plastics.
Petroleum-based plastics, includingpoly(ethylene terephthalate) (PET), areextensively used in everyday products, suchas single-use plastic bottles, textiles, and foodwrappers. While such products are disposedof rapidly after use, they persist in the envi36

NAIST Research Highlights

ronment for hundreds of years. The plastic
pollution resulting from our throw-away
culture has now exceeded manageable levels
and is overwhelming the planet’s ability todeal with it. The environmental impacts are
becoming increasingly obvious, with wildlife
and human health increasingly threatened.
Although reducing the manufacture of
unnecessary single-use plastics and improving waste management systems will help
ease the pollution crisis, our reliance on theconvenience of plastic products is unlikely
to be abated any time soon. Researchers are
therefore looking at alternative approaches to“clean up” the more persistent plastics fromour environment and it appears that microbes
may offer some promising solutions.
“Certain bacteria harbor the necessary

enzymes to degrade PET, the most problematic plastic environmentally,” explains seniorauthor Shosuke Yoshida. “Our research has
shown that the bacterium Ideonella sakaiensis
converts PET into poly(3-hydroxybutyrate)(PHB), a type of poly(hydroxyalkanoate)(PHA) plastic that is biodegradable,” hecontinues.
This finding is particularly promisingbecause it addresses two current problems
for the sustainability of plastics: degrading
the most persistent form of petroleum-based
plastic while sustainably producing biodegradable plastics.
“We believe that this discovery could
be significant in tackling plastic pollution,”
Yoshida states, “as we show that the PET-degradation and PHB-synthesis pathways arefunctionally linked in I. sakaiensis. Thismight provide a novel pathway where a single
bacterial species breaks down difficult-to-recycle PET plastics and uses the products tomake biodegradable PHA plastics.”
Given the overwhelming challenge of
dealing with worldwide plastic pollution, thisnovel bacterial approach may be a significant
part of the solution.

Reference

Ry o g a F u j i w a r a , R i k a k o S a n u k i , H i r o h a r u
Ajiro, Toshiaki Fukui, Shosuke Yoshida. 2021.
Direct fermentative conversion of poly(ethylene
terephthalate) into poly(hydroxyalkanoate) by
Ideonella sakaiensis. Scientific Reports, 11, 19991.

Ideonella sakaiensis cultured on poly(ethylene terephthalate) (PET) accumulatespoly(3-hydroxybutyrate) (PHB).
NAIST Research Highlights

37

Structural Life Science
Prof. Tomoya Tsukazaki

A protein with an unprecedented
fold helps bacteria uptake
thiosulfate as a sulfur source
Researchers at NAIST discover that the YeeE protein allows bacteria to uptake thiosulfate from theenvironment for cysteine synthesis, showing a unique hourglass shape that allows YeeE to undergo
minimal conformational changes for the uptake compared with other transporter proteins.

A

new study led by researchers at NAIST, in Science Advances hasreported the crystal structure of YeeE, a membrane protein thatallows bacteria to uptake thiosulfate from the environment
in order to synthesize L-cysteine. The structure reveals that YeeE
has a characteristic hourglass shape that results in a sophisticatedmechanism for the uptake, providing fundamental information that
could greatly lower cysteine
production costs in industry.
Most people know thatwater is made of hydrogenand oxygen atoms, andcarbon compounds are foundin all life on earth. However,other elements like sulfur are
also indispensable for life, andsulfur-based molecules likeL-cysteine are essential formany of our proteins. L-cysteine is also commercially
important, as it is heavily usedby the food, cosmetic andpharmaceutical industries.
“In nature, L-cysteine isproduced by microorganismsthat collect inorganic sulfur
YeeE structure.
in the soil. If the secretionand production efficiencyof L-cysteine by microorganisms can be dramatically improved, thisprocedure will be superior to existing methods, such as in the production of glutamic acid by Corynebacterium glutamicum,” says Tomoya
Tsukazaki, an expert in structural biology.
Bacteria can take up both sulfate and thiosulfate ions from theenvironment in order to synthesize L-cysteine. The efficiency ofthe synthesis from thiosulfate is higher because of fewer chemical
reaction steps. To examine which proteins are crucial for thiosulfate
uptake, the researchers conducted a series of genetic studies, finding
YeeE.
To understand how YeeE physically conforms for the transport,
the researchers uncovered crystal structures of YeeE that revealed an
unprecedented fold forming an hourglass shape. “Both the inner andouter surfaces of YeeE are indented toward the center. We think this38

NAIST Research Highlights

shape is crucial for initiating the uptake and conducts thiosulfate,”
explains Yoshiki Tanaka, the first author of the study.
Molecular dynamics simulations implied that the uptake occurs
by passing the thiosulfate ion through three key sites in the YeeE
structure. In the model, the first site attracts a thiosulfate ion to
the positively charged surface. Then using S-H-S type hydrogenbonds, YeeE passes the ion
to the other two sites andon to the cytoplasm without
itself undergoing any major
conformational changes.
This mechanism for uptake is quite unusual among
membrane transporters
according to Tanaka. “There isa lot less movement comparedwith transporters that haveinward- and outward-facing
structures or use rocking
bundle motions. YeeE is not
structurally designed to use
these other mechanisms,” hesays.
Tsukazaki adds that the
new mechanism expands ourknowledge of nutrient transport into a cell, knowledge
that can be exploited for industrial purposes. “Comparatively little is
known about the YeeE family of transport proteins. Through morestudy of the structure for the uptake and genetic modifications, wemight be able to artificially design ‘super’ YeeE that enhance L-cysteine productivity via high thiosulfate uptake” he says.

Reference

Yoshiki Tanaka, Kunihito Yoshikaie, Azusa Takeuchi, Muneyoshi Ichikawa,
Tomoyuki Mori, Sayaka Uchino, Yasunori Sugano, Toshio Hakoshima,
Hiroshi Takagi, Gen Nonaka, Tomoya Tsukazaki. 2020. Crystal structureof a YeeE/YedE family protein engaged in thiosulfate uptake. Science
Advances, 6, eaba7637.

Systems Neurobiology and Medicine
Prof. Naoyuki Inagaki

Shootin1a – The missing link
underlying learning and memory
Researchers at NAIST find that the shootin1a protein is crucial for allowing dendritic spines tochange in size, which is an important process underlying learning and memory.

In neurons, changes in the size of dendritic spines—small cellular
protrusions involved in synaptic transmission—are thought tobe a key mechanism underlying learning and memory. However,the specific way in which these structural changes occur remains
unknown. In a study published in Cell Reports, a team of researchers
led by NAIST has revealed that the binding of cell adhesion moleculeswith actin, via an important linker protein in the structural backbone
of synapses, is vital for this process of structural plasticity.
Actin proteins make up an important part of a cell’s structure, or cytoskeleton, and allow for dynamic changes in this structure by forming
microfilaments when growth or movement is required. It was originally
thought that the polymerization of actin was all that was needed for
dendritic spines to change size in response to synaptic activation, but
researchers at NAIST found that this process alone was not enough tocause structural plasticity, and decided to address this problem.
“Current models of structural plasticity in dendritic spines do nottake mechanical force into account,” says Naoyuki Inagaki, corresponding author. “We had already identified the role of shootin1a, a
protein involved in neuronal development, in axon growth and so we
wanted to investigate whether this protein might also have a role inthe structural plasticity of dendritic spines.”
To explore this question, the researchers used neurons of control
and shootin1a knockout rodents to examine whether shootin1a
was involved in the formation of dendritic spines. The researchers
wanted to determine if mechanical force was generated in dendritic
spines by the shootin1a-mediated coupling of actin and cell adhesion

molecules—cell-surface proteins that bind cells together at synapses—
similar to what they had observed in axons.
“The results were clear,” explains Inagaki. “We found that shootin1a mechanically linked polymerizing actin with cell adhesion molecules in dendritic spines, and revealed that synaptic activity enhanced
this coupling, thus allowing the actin filaments to push against themembranes and enlarge spines.” The results of this study are the firstto link mechanical force with synaptic activity-dependent dendritic
spine plasticity and provide new insights into the mechanisms ofstructural plasticity in these spines.
Given that changes in activity-dependent dendritic spine plasticity
have been implicated in multiple neuropsychiatric and neurodegenerative disorders, including autism spectrum disorder and Alzheimer’s
disease, these findings are important because they suggest thatshootin1a disruption may lead to the development of neurologicaldisorders. Future studies into this mechanism of structural plasticity
in dendritic spines might provide new drug targets for these disorders.

Reference

Ria Fajarwati Kastian, Takunori Minegishi, Kentarou Baba, Takeo
Saneyoshi, Hiroko Katsuno-Kambe, Singh Saranpal, Yasunori Hayashi,
Naoyuki Inagaki. 2021. Shootin1a-mediated actin-adhesion coupling
generates force to trigger structural plasticity of dendritic spines. Cell
Reports, 35, 109130.

Fluorescence time-lapse images of dendritic spines (A) and time course of their volume changes (B) of hippocampal neurons inslice culture.
NAIST Research Highlights

39

Plant Cell Function
Prof. Takashi Hashimoto

Hired blade:Anchoring complex in plant cells
recruits its own katana sword
Researchers at NAIST find that an anchoring complex stabilizes microtubule creation sites within
plant cells, then recruits katanin—named after the katana sword—to cut new microtubules.

T

recruits katanin to the same location to enable the efficient release of
he katana, a Japanese sword, may be thought of solely as adaughter microtubules.
weapon used by the samurai. But scientists from Japan have
“These ‘glue-and-cut’ functions performed by Msd1-Wdr8 anddiscovered that not only do plants wield their own katanas
their effects on microtubule stability may seem confusing at first,
within their cells, they recruit them to specific locations within those
but they probably enable strict control of microtubule release by thecells to do their work.
katanin activity,” explains Hashimoto.
In a study published in Nature Communications, a team of reThis study will inform future research on whether the Msd1-Wdr8
searchers led by NAIST has revealed that the enzyme katanin, which
complex in animal cells also recruits katanin, and whether other sites
is named after the katana, is used by an anchoring complex to cut
use similar mechanisms for the stabilization and release of daughter
microtubules at specific locations of the framework within individual
microtubules. The results of this study will be of interest to cell biolplant cells.
ogists, especially those working on cytoskeletons, in plants and other
Katanin severs microtubules in cells, which is an important steporganisms.
in cell division and central to the development of many organisms,
including plants and animals. Microtubules form part of the cytoskeleton, a complex network of protein filaments found in all cells. The
severing performed by katanin enables mobility, which is importantduring development, and treadmilling—a phenomenon where one
end of a filament lengthens as the other shrinks, which results in asection of filament that seems to ‘move’ like a treadmill.
“Katanin severs microtubules at specific locations in plant andReference
animal cells, and this leads to active reorganization of the microtubule
Noriyoshi Yagi, Takehide Kato, Sachihiro Matsunaga, David W. Ehrhardt,
cytoskeleton,” says senior author of the study Takashi Hashimoto. “But
Masayoshi Nakamura, Takashi Hashimoto. 2021. An anchoring complex
the mechanisms for targeting this extraordinary enzyme at specificrecruits katanin for microtubule severing at the plant cortical nucleationsites. Nature Communications, 12, 3687.
sites within the cell are not well understood—these are what we
wanted to investigate.”
The team’s genetic
and cell biology research
results showed that the
microtubule anchoring
complex Msd1-Wdr8 isused to stabilize microtubule nucleation sites(where microtubules areformed) in plant cells toprevent early release of
the new microtubules( c a l l e d ‘d a u g h t e r
microtubules’). But in aseemingly counterintuitive twist, Msd1-Wdr8
The anchoring complex Msd1-Wdr8 stabilizes the base of the Y-shaped nucleation structure and recruit katanin to sever thethen turns around andbasal end of the daughter microtubule.

40

NAIST Research Highlights

MATERIALSSCIENCEBio-Process Engineering
Prof. Yoichiroh Hosokawa

Antibiotic resistance testing
no longer impeded by time
Researchers at NAIST develop a novel method for antibiotic resistance testing that mighttransform microbial screening in clinical and research labs.

S

datasets, the machine learning tool could automatically label theignificant time is needed to determine the drug susceptibility
reference dataset as the “learning” dataset and use it to learn the
profile of a bacterial infection. Now, a team of researchers led
characteristics of an untreated bacterium. By real-time comparison
by NAIST has published reports on a technology that willwith antibiotic-treated cells, the tool can identify whether the bacteria
dramatically speed up this otherwise slow process and possibly help
are susceptible to the drug and can even identify what proportion
save lives.
of bacterial cells are resistant in a mixed-resistance population.
The U.S. CDC states that antibiotic-resistant infections are respon"Although there was a misidentification error of less than 10% in oursible for killing over a million people worldwide every year. Central
work, there was a clear discrimto managing resistant infections
ination between susceptible andis quickly identifying an approresistant cells within 2 hours ofpriate treatment to which theantibiotic treatment," explains
infective bacteria are susceptible.
Yoichiroh Hosokawa, another"Oftentimes susceptibility results
senior author in the group.
are needed much faster thanThis work is not limited toconventional tests can deliver
rapid evaluations of infections
them," says Yaxiaer Yalikun,
in clinical settings. For example,
senior author. "To address this,drug discover y researchers
we developed a technology that
could use it to conduct quick
can meet this need."initial investigations of drug
The group’s work is based
efficacy against any cell, as longon impedance cytometry which
as the cellular response results inmeasures the dielectric propera change in dielectric properties.
ties of individual cells with high
Impedance cytometry mightthroughput—over a thousand
become a staple of clinical andcells per minute. Because theresearch labs in the coming
electrical readout of a bacterium
years.
corresp onds to its physicalresponse to an antibiotic, one has
Schematic of the intelligent impedance system, consisting of a parallela straightforward means of deimpedance cytometry and a machine learning-based detection system.
termining whether the antibiotic
works against the bacteria. Conventional impedance cytometry involves analyzing the test (antibiotic
treated) and reference (untreated) particles in one sample followedby calibrating the impedance of the two particles—both steps require
References
technical specialists to carry out extensive post-processing, which was
TaoTang, Xun Liu, Yapeng Yuan, Tianlong Zhang, Ryota Kiya, Yang Yang,a major limitation the group was determined to overcome.
Yoichi Yamazaki, Hironari Kamikubo, Yo Tanaka, Ming Li, Yoichiroh
In a study published in ACS Sensors, the group develops a novel
Hosokawa, Yaxiaer Yalikun. 2022. Parallel impedance cytometry for
impedance cytometry method that simultaneously analyses the test
real-time screening of bacterial single cells from nano- to microscale.
ACS Sensors, 7, 3700–3709.
and reference particles in separate channels, creating easily analyzable
TaoTang, Xun Liu, Yapeng Yuan, Ryota Kiya, Tianlong Zhang, Yang Yang,
separate datasets. This cytometry had nanoscale sensitivity, allowing
Shiro Suetsugu, Yoichi Yamazaki, Nobutoshi Ota, Koki Yamamoto,for detection of even minute physical changes in bacterial cells. In
Hironari Kamikubo, Yo Tanaka, Ming Li, Yoichiroh Hosokawa, Yaxiaer
a concurrent study published in Sensors and Actuators B, the group
Yalikun. 2022. Machine learning-based impedance system for real-time
designed a machine learning tool to analyze the impedance cytometry
recognition of antibiotic-susceptible bacteria with parallel cytometry.
Sensors and Actuators: B. Chemical, 374, 132698.
data. Because the new cytometry method splits the test and reference

42

NAIST Research Highlights

Bio-Process Engineering
Prof. Yoichiroh Hosokawa

Feeling the pressure
Researchers at NAIST refine the equations for mechanical stiffness to apply to plant cell walls
based on elastic shell theory and verified using finite element simulations, which may lead to abetter understanding of how plants resist stress.

A

The work helps generalize our understanding of stiffness forteam of researchers led by NAIST has shown how the stiffness
living systems. This knowledge can be applied to help ensure thatof plant cell walls depends on their elasticity and internal
plants maintain their structure even under stressful situations, such as
turgor pressure using elastic shell theory. By utilizing atomic
during periods of water deprivation.
force microscopy (AFM) combined with finite element computer
simulations, they were able to show that cell stiffness is very sensitive
to internal turgor pressure.
Many people will have fond memories from their school days
looking at onion peels under a microscope. While the individual
Referencecells might have seemed then like simple rectangles, the stability ofSatoru Tsugawa, Yuki Yamasaki, Shota Horiguchi, Tianhao Zhang, Takara
plant cells reflects complex combinations of forces. In addition to
Muto, Yosuke Nakaso, Kenshiro Ito, Ryu Takebayashi, Kazunori Okano,the cell membrane which is similar in animals, plant cells also have a
Eri Akita, Ryohei Yasukuni, Taku Demura, Tetsuro Mimura, Ken’ichi
rigid cell wall that provides structural integrity. Turgor, meaning the
Kawaguchi, Yoichiroh Hosokawa. 2022. Elastic shell theory for plant cellnormal rigidity of cells due to the pressure from its contents, is also awall stiffness reveals contributions of cell wall elasticity and turgor pressurein AFM measurement. Scientific Reports, 12, 13044.
critical factor in maintaining balance with the environment. Too little
pressure can cause the cell to shrink. Cells can regulate theirturgor pressure osmotic flows that tend to balance the salt
concentrations between the interior and the outside of thewall. However, the resulting mechanical properties of plant
cells remain nebulous. For example, using AFM alone todetermine the stiffness from cell wall deformation makes it
difficult to separate the contributions from the tension of thecell wall, cell geometry and turgor pressure.
Now, the research team led by NAIST has used finite
element method (FEM) simulations to verify a new formulabased on elastic shell theory. This allowed them to interpretthe apparent stiffness observed using AFM. The teamstudied onion epidermal cells, which are a model system
for understanding the physical properties of plant cells.
“Looking at the force versus indentation data suggested thatthe standard equations were not sufficient for interpretingthe apparent stiffness of plant cells,” senior author Yoichiroh
Hosokawa says.
Based on the FEM simulations, the elastic shell theory
equation was shown to be better at describing the AFM
response of the onion cells, compared with the conventionalmodel used for objects without internal turgor pressure.
Moreover, their findings suggest that tension caused by turgor pressure regulates cell stiffness, which can be modified
by slight changes, on the order of 0.1 megapascals. “Ourtheoretical analysis paves the way for a more complete understanding of the forces inherent in a plant cell,” Hosokawa
Overview of the study system that utilizes an atomic force microscopy (AFM) combinedsays.
with finite element computer simulations.

NAIST Research Highlights

43

Bio-Process Engineering
Prof. Yoichiroh Hosokawa

Separating tiny bacteria
by shape: Simple techs
for E. coli sorting
Researchers at NAIST create a lab-on-a-chip with a viscoelastic fluid that can separate spherical from
elongated bacteria, which can help standardize biological research and improve medical testing.

A

team of researchers led by NAIST has
achieved shape-based separation ofEscherichia coli (E. coli) bacteria using aviscoelastic fluid inside microfluidic channels.
This “lab-on-a-chip” system has the capability
to assist in making scientific experiments more
reproducible, as well as provide more accurate
assessments of the severity of bacterial infections
based on patient samples.
The task of separating tiny bacteria based on
their shape is a vital step in scientific and clinicalapplications, but also a difficult one. Whether
an E. Coli bacterium is in a round or elongatedconfiguration can indicate its state of biological
function, and the ability to prepare homogeneous
populations with uniform shapes may help
improve experimental reproducibility. Moreover,being able to sort samples into sub-populations
based on their shape may help diagnose patient
health or assess environmental contamination.
However, this ability has been difficult to achieve,
especially at the scales needed to be practical.
The design of the viscoelastic microfluidic device for shape-based separation of drug-treated E.
Now, the research team led by NAIST hascoli.
been able to separate drug-treated E. coli bacteriausing microfluidic channel containing poly-eththe separated E. coli populations to determine linkage between cell
ylene-oxide (PEO), a viscoelastic fluid. The elongation of the bacteria,
shape, antibiotic susceptibility and gene function,” author Yoichiroh
as measured by the aspect ratio, ranged from 1.0 (spherical) to 5.5
Hosokawa says.
(rod-shaped). Microchannels made of poly(dimethylsiloxane) were
The team hopes the microfluidic platform they developed will becreated with photolithography. PEO, which has both high viscosity
adopted by the broader scientific community for different applications
and elasticity, was made to flow along the edges of an expanding
in the fields of microbiology, molecular biology and biomedicine.
V-shaped chamber. The elongated bacterium followed this flow to becollected at the edges of the device.
“The shape of a E. coli bacterium changes based on its attachment,
motility, dispersal, predation and differentiation behaviors,” author
Reference
Yalikun Yaxiaer explains. A rod-like configuration is more efficient
Tianlong Zhang, Hangrui Liu, Kazunori Okano, Tao Tang, Kazuki Inoue,
for swimming, while a spherical shape may help it form communities.
Yoichi Yamazaki, Hironari Kamikubo, Amy K Cain, Yo Tanaka, David
These changes are important milestones for the progression of disease,
Inglis, Yoichiroh Hosokawa, Yalikun Yaxiaer, Ming Li. 2022. Shape-based
and whether the bacteria are swimming free or forming communities.
separation of drug-treated Escherichia coli using viscoelastic microfluidics.
Lab on a Chip, 22, 2801–2809.
“This technology will allow downstream genomics experiments on

44

NAIST Research Highlights

Bio-Process Engineering

Precise total assessment of the cells with impedance signals.

Prof. Yoichiroh Hosokawa

From outside to inside:
A rapid and precise total
assessment method for cells
Researchers at NAIST show that using four frequencies of applied voltage can improve themeasurement of cell size and shape during impedance cytometry, enabling to enhance the speed
and accuracy of biological experiments.

H

aving a good eye for detail is an essential skill for many professions. In particular, biologists use special techniques andadvanced technology to analyze individual cells with unprecedented precision. Impedance cytometry is one experimental methodthat can reveal specific characteristics of living single cells. This technique requires electrical penetration, in which high-frequency current
can freely pass through the cell membrane, without damaging thecell. Now, a team of researchers led by NAIST has determined optimal
conditions to perform impedance cytometry. Their work may lead to
rapid assessment of cells during culture in biological experiments.
An improved method for measuring the morphology and biomassof single cells using impedance cytometry has been introduced
in a recently published study in Microsystems & Nanoengineering.
Impedance cytometry involves applying high-frequency voltages
to electrodes to measure complex impedance, which can provide
information about the shape and effective volume of the cell. In the
study, the researchers used different phases of voltage signals at four
frequencies. They showed that applied voltages with frequencies ofaround 7 MHz are able to pass through the membrane of Euglena
gracilis cells. Higher frequencies can monitor changes in biomass,
while lower frequencies can track volume changes.
When a high-frequency electrical field penetrates the cell membrane, the uneven intracellular distribution tilts the impedance pulses

to the left or right, which has been verified in simulation and experiments. “Ultimately, our method for determining the conductivity of
the cell membrane relies on the degree of tilt caused by the electricalpulses,” says author Yoichiroh Hosokawa. The team also performed
calibration studies using beads to better understand the underlying
physical mechanisms of this effect.
“This research enables the easy determination of the electricalpenetration of a cell membrane, and the proposed platform is applicable to multiparameter assessment of the organism’s state during
cultivation,” says senior author Yaxiaer Yalikun. This platform may beeasily integrated into microfluidic systems for the scalable monitoringof biological experiments.
The need for efficient and highly accurate analysis of living single
cells may be met by this new impedance cytometry method developedby the research team. Future applications could be extended to cells
in mammals to monitor specific membrane changes in fields such as
oncogenesis and cell aging.
Reference

Tao Tang, Xun Liu, Yapeng Yuan, Tianlong Zhang, Ryota Kiya, Yang Yang,
Kengo Suzuki, Yo Tanaka, Ming Li, Yoichiroh Hosokawa, Yaxiaer Yalikun.
2022. Assessment of the electrical penetration of cell membranes using fourfrequency impedance cytometry. Microsystems & Nanoengineering, 8, 68.
NAIST Research Highlights

45

Solid-state Information Physics

Prof. Tomohiro Matsushita

Old silicon learns
new tricks
Researchers at NAIST fabricate arrays of atomically smooth iron-coatedsilicon crystals pyramids with unusual magnetic properties, which could
enhance the functionality of 3D spintronics and other technologies.

46

NAIST Research Highlights

Schematics representing Ewald sphere and reciprocal lattice rods from a pyramid surface,reflecting diffraction patterns.

Ultra-small integrated circuits have
revolutionized mobile phones, home
appliances, cars, and other everyday
technologies. To f ur t her mini aturize
electronics and enable advanced functions,circuits must be reliably fabricated in three
dimensions. Achieving ultrafine 3D shapecontrol by etching into silicon is difficult
because even atomic-scale damage reduces
device performance.
A team of researchers led by NAIST
has reported, in a new study seen in Crystal
Growth and Design, silicon etched to adopt
the shape of atomically smooth pyramids.
Coating these silicon pyramids with a thinlayer of iron imparts magnetic properties that
until now were only theoretical. Ken Hattoriat NAIST is widely published in the field of
atomically controlled nanotechnology. One
focus of Hattori's research is in improving thefunctionality of silicon-based technology.
“Silicon is the workhorse of modern electronics because it can act as a semiconductor
or an insulator, and it’s an abundant element.
However, future technological advances
require atomically smooth device fabricationin three dimensions,” says Hattori.
A combination of standard dry etching
and chemical etching is necessary to fabricate
arrays of pyramid-shaped silicon nanostructures. Until now, atomically smooth surfaces
have been extremely challenging to prepare.
“Our ordered array of isosceles silicon
pyramids were all the same size and had flat
facet planes. We confirmed these findings by
low-energy electron diffraction patterns andelectron microscopy,” explains lead author of
the study Aydar Irmikimov.
An ultrathin—30 nanometer—layerof iron was deposited onto the silicon to
impart unusual magnetic properties. The
pyramids' atomic-level orientation defined
the orientation—and thus the properties—of
the overlaying iron. “Epitaxial growth of ironenabled shape anisotropy of the nanofilm.
The curve for the magnetization as a functionof the magnetic field was rectangular-likeshaped but with breaking points which werecaused by asymmetric motion of magnetic
vortex bound in pyramid apex,” explains
Hattori.
The researchers found that the curve hadno breaking points in analogous experiments
performed on planar iron-coated silicon.
Other researchers have theoretically predicted the anomalous curve for pyramid shapes,
but the NAIST researchers are the first to
have shown it in a real nanostructure.
“Our technology will enable fabricationof a circular magnetic array simply by tuning
the shape of the substrate,” says Irmikimov.
Integration into advanced technologies suchas spintronics—which encode informationby the spin, rather than electrical charge, ofan electron—will considerably accelerate thefunctionality of 3D electronics.

Reference

Aydar Irmikimov, Liliany N. Pamasi, Azusa N.
Hattori, Takaaki Higashi, Shunta Takahashi, Emilia E.
Hashamova, Xiaoqian Shi, Fangzhun Guo, Nobuyoshi
Hosoito, Ai I. Osaka, Hidekazu Tanaka, Ken Hattori.
2021. Atomically architected silicon pyramid singlecrystalline structure supporting epitaxial material
growth and characteristic magnetism. Crystal Growthand Design, 21, 946–953.
NAIST Research Highlights

47

Organic Electronics
Prof. Masakazu Nakamura

Easy fabrication of
next-generation,
super-flexible
electronic circuits
Researchers at NAIST fabricate one-dimensional soft n-type semiconductors by a method that simplifies layer-by-layer assembly, which could befoundational to developing wearable electronics technologies with CMOS
circuits.

F

lexible semiconductors are essential
f or f utu re w e ar a b l e e l e c t ron i c stechnologies, but have been difficult
to integrate into complex architectures.
Now, in a study published in Advanced
Electronic Materials, researchers from NAIST
have developed a straightforward means offabricating high-quality soft semiconductors
for advanced electrical circuits.
Modern integrated circuit technology
dep ends on basic elements known ascomplementary metal oxide semiconductor(CMOS) circuits. Silicon is the semiconductor component of most modern CMOS
technology. However, because future CMOS
circuits must (for example) mold to the shapeof a body or integrate into clothing, muchwork has focused on developing soft, flexible,
polymer-based semiconductors.
48

NAIST Research Highlights

Several technical challenges must be
overcome to integrate such semiconductors,
especially n-type ones which flows electrons,
into CMOS circuits. For example, preparing
high-quality, layer-by-layer structures—
important for CMOS device functionality—
tends to be rather slow and challenging.
Solving these challenges is the problem thatthe researchers at NAIST sought to address.
"Ideally, one would be able to deposit
polymer films onto liquid substrates for easeof transfer to any other substrate," explains
Manish Pandey, lead author. "Our strategy
offers superior control over the resulting
semiconductor film morphology, which is
critical to the electrical properties, comparedwith conventional solution processing."
This work is based on unidirectional
floating film transfer. By using a liquid

substrate that does not dissolve the polymer,
a solvent-dissolved polymer can be added
dropwise onto the substrate, in a manner that
forms a one-dimensional floating polymer
film. Upon evaporation of the solvent, thepolymer molecules orient perpendicular tothe length direction of the film. This molecular morphology optimizes the electricalproperties of the polymer film. Once thefilm solidifies, one can easily transfer it onto
another substrate—e.g., for layer-by-layer
deposition.
"We prepared an n-channel transistor thatexhibited nearly no threshold voltage, which
is important for maintaining power efficiency," says Masakazu Nakamura, senior author.
"By using our approach, preparing andintegrating n-channel as well as p-channeltransistors into one device—based on flexible

semiconductors—should be straightforward."
This work succeeded in preparing
one-dimensional, polymer-based semiconductor films in an inexpensive mannerthat is straightforward to replicate. The
NAIST researchers' polymer film assembly
methodology will be useful for advancing theprospects of flexible electronics, and helping
to find replacements for silicon in upcoming
wearable CMOS technology.

Reference

Manish Pandey, Yuya Sugita, Jumpei Toyoda, Shohei
Katao, Ryo Abe, Yongyoon Cho, Hiroaki Benten,
Masakazu Nakamura. 2023. Unidirectionally aligned
donor–acceptor semiconducting polymers in floating
films for high-performance unipolar n-channelorganic transistors. Advanced Electronic Materials, 9,2201043.

Overview and characteristics of the unidirectional floating film transfer method
developed in this study.
NAIST Research Highlights

49

Organic Electronics
Prof. Masakazu Nakamura

Shedding light on polymer
solar cells: Illuminating how solvent
additives improve efficiency
Researchers at NAIST use photoconductive atomic force microscopy to investigate the role of solvent
additives in the performance of all-polymer blend solar cells.

A

ll-polymer blend solar cells are expected to play an importantrole in the transition to clean energy technologies because they
can be easily produced in large-scale flexible sheets. However,their performance has lagged behind that of more traditional siliconalternatives, as well as other organic solar cells.
All-polymer blend solar cells are formed by combining twopolymer solutions that solidify into a film on an electrode with in
the form of interpenetrating networks, a kind of “phase-separation”.
The introduction of solvent additives to the polymer solution has
been shown to increase the efficiency of all-polymer blend solar cells.
However, the exact process underlying this improvement has notbeen fully understood. Now, in a study recently published in ACS
Applied Polymer Materials, researchers from NAIST have investigatedthe performance enhancement mechanism using photoconductive
atomic force microscopy (PC-AFM). Their findings are expected tohelp accelerate the widespread application of polymer-based solar
cells.
"The empirical nature of solvent additive-mediated efficiency
enhancement has hindered the optimization of all-polymer blend
solar cell performance, so there has been an urgent need for a greater
understanding of the process," explains senior author Hiroaki Benten.
"To that end, we used PC-AFM to interrogate the nanoarchitecturethat underpins the performance enhancement."
PC-AFM is an advanced microscopy technique that allows
photocurrents to be visualized with nanometer-scale resolution. The
researchers found that trace solvent additives improved the power
conversion and photocurrent density of an all-polymer blend solar

cell by a factor of up to ~3 by enhancing the ordering and crystallization of the polymer microstructure in the solar cell without damaging
the phase-separated structure.
Absorption spectroscopy measurements further confirmed thatthe trace additives improved the ordering in the polymer microstructures. By forming a network that efficiently transports the photogenerated charges to the external electrode, the flow of photocurrent isincreased.
“We found that local photocurrents were enhanced, somewhat
like forming a new charge current highway, while the scale of phaseseparation that is critical to device functionality was retained," says
coauthor Masakazu Nakamura. "We believe that this insight will bebroadly applicable to all-polymer blend solar cells, not just those
based on our choice of polymers."
The results of the study are expected to be important for optimizing the performance of all-polymer blend solar cells. By using
the findings to minimize laboratory trial-and-error, it is hoped that
researchers can speed up ongoing bench-to-market efforts, taking us
a step closer to high-performance solar cells that are environmentally
sustainable and easy to produce on a large scale.

Reference

Yuji Yamagata, Hiroaki Benten, Toshiki Kawanishi, Masakazu Nakamura.
2022. Nanoscale observation of the influence of solvent additives on allpolymer blend solar cells by photoconductive atomic force microscopy.
ACS Applied Polymer Materials, 4, 169–178.

Improvements of the device performance by the introduction of solvent additives to the polymer solution.
50

NAIST Research Highlights

Organic Electronics
Prof. Masakazu Nakamura

Fill me up: Improved efficiency of
all-polymer solar cells
Researchers at NAIST reveal the science that underpins the efficiency of a versatile class of
solar cells, which might help solve the environmental problems caused by relying solely on
silicon-based solar cells.

S

was substantial charge delocalization in the donor and acceptor
olar cells are an important contributor to a renewable energy
domains," explains Masakazu Nakamura, senior author. "Appropriate
supply, but solar panel waste will quickly become an enormous
aggregation of the polymer donors and acceptors led to a substantiallyproblem. Now, in a study recently published in Journal ofordered local structure of the polymer, which helped keep the separaMaterials Chemistry A, researchers from NAIST have investigated thetion of the electrons from the holes."science that might help improve the utility of easily mass-producible,
Even if researchers completely solve the efficiency problem of
all-polymer-based solar cells.
all-polymer solar cells, they'll still need to improve on the 10-year
Globally, approximately one-third of electricity currently comes
service life of the most advanced research prototypes. Additionalfrom renewable sources. Silicon-based solar cells are the major
research efforts include optimizing the film morphology, and evencontributor, but there's an increasing problem: what to do with thedeveloping hybrid polymer/silicon solar cells, to optimize energy
panels after their 30-year lifetime. A May 2022 article in Chemical &collection and efficiency. In the coming years, solar cells might look
Engineering News lays out the problem: even when facilities recycleand function completely different from—and better than—modern
the frames and covers of the panels, the most valuable or even toxic
technology.
elements are simply disposed. With a forecasted 80 million metrictons of solar panel waste to have been produced by 2050, this is amassive waste problem.
Polymer-based solar cells are a possible, less-wasteful solution.
Reference
Such panels are thin and flexible, and thus are in principle quite
Ren Hagio, Hiroaki Benten, Yunju Na, Zhiyuan Liang, Naoki Oyaizu,
versatile. Nevertheless, they have certain problems; e.g., a lower power
Junya Suzuki, Shota Kubota, Min-Cherl Jung, Hirotaka Kojima, Masakazu
conversion efficiency than silicon. "This efficiency is substantially
Nakamura. 2022. Chain aggregation dictates bimolecular charge
limited by the fill factors: commonly less than 60%, even in advanced
recombination and fill factor of all-polymer blend solar cells. Journal of
Materials Chemistry A, 10, 21727–21737.
devices," says corresponding author Hiroaki Benten. "The sciencet h at u nd e r pi ns t he l i m ite defficiency of all-polymer blend
solar cells remains insufficiently
unexplored."A ground-breaking result of
this research is the high fill factor:
ca. 70%, which remained ca. 60%
even for polymer films several
hundred nanometers t hick.
Competing polymer technology
exhibits a ca. 40% fill factor at
this thickness. This is because
bimolecular recombination offree electrons with free holes substantially inhibited the fill factor
prior work, but was suppressed in
the current study.
W h at s up p re s s e d b i m o lecular recombination withinHigh fill factors of the all-polymer blend solar cells are achieved by long charge-carrier lifetimes owing to low
the polymer blends? "Therebimolecular charge recombination coefficients.

NAIST Research Highlights

51

Biomimetic and Technomimetic Molecular Science

Prof. Gwénaël Rapenne

Gearing up
nanoscale machines
Researchers at NAIST in partnership with University of Paul Sabatier advance the science of molecular-scale gear trains for transmitting rotational
force with the visualization of rotational snapshots by scanning tunnelingmicroscopy.

G

ear trains have been used for centuries to translate changes ingear rotational speed into changes in rotational force. Cars,
drills, and basically anything that has spinning parts use them.
Molecular-scale gears are a much more recent invention that could
use light or a chemical stimulus to initiate gear rotation. Researchersat NAIST in partnership with University of Paul Sabatier, France,
have reported in a new study published in Chemical Science a means
to visualize snapshots of an ultrasmall gear train—an interconnected
chain of gears—at work.
NAIST project leader Gwénaël Rapenne has devoted his career tofabricating molecular-scale mechanical devices, such as wheels andmotors. Researchers recently designed a cogwheel for a molecular
gear train but currently have no means to visualize the gears in action.
"The most straightforward way to monitor the motion of molecular
gears is through static scanning tunneling microscopy images. For
these purposes, one of the teeth of the cogwheels must be either
sterically or electrochemically distinct from the other teeth," explains
Rapenne.

52

NAIST Research Highlights

The researchers first created a molecular cogwheel comprising
five paddles, where one paddle is a few carbon atoms longer than theother four paddles. However, as they showed last year, differences in
paddle length disrupt the coordinated motion along the gear train.
Thus, differences in paddle electrochemistry are a more promising
design approach but synthetically more challenging.
"We used computational studies to predict whether electron-withdrawing units or metal chemistry could tailor the electronic properties of a paddle, without changing paddle size," says Rapenne. Suchtailored properties are important because one can observe them as
differences in contrast by using scanning tunneling microscopy, andthereby facilitate static imaging.
"Our pentaporphyrinic cogwheel prototypes contained one paddle
with either a cyanophenyl substituent or a zinc—rather than nickel—
metal center,” explains Rapenne. “Various spectroscopy techniques
confirmed the architectures of our syntheses."
How can researchers use these cogwheels? Imagine shining ahighly focused beam of light, or applying a chemical stimulus, to one

of the gears to initiate a rotation. By so doing, one could rotate a seriesof cogwheels in a coordinated manner as in a conventional gear train,
but on a molecular scale which consists in the ultimate miniaturizatio
of devices. "We now have the means to visualize such rotations," notes
Rapenne.
By using this development to carry out single-molecule mechanics
studies, Rapenne is optimistic that the broad research community will
have a powerful new design for integrated nanoscale machines. "We're
not there yet, but are working collaboratively to make it happen assoon as possible," he says.

Reference

Seifallah Abid, Yohan Gisbert, Mitsuru Kojima,
Nathalie Saffon-Merceron, Jérôme Cuny, Claire
Kammerer, Gwénaël Rapenne. 2021. Desymmetrised
pentaporphyrinic gears mounted on metallo-organic
anchors. Chemical Science, 12, 4709–4721.

A train of molecular gears composed of star-shape molecules.
NAIST Research Highlights

53

Photonic and Reactive Molecular Science
Prof. Tsuyoshi Kawai

Invisible X-rays turn blue
Researchers at NAIST develop a new reaction system that can detect X-rays at the highest
sensitivity ever recorded by using organic molecules.

A

new reaction system can detect X-rays
at the highest sensitivity ever recordedby using organic molecules. The system,
developed by researchers at NAIST, involves
the cycloreversion of terarylene, causing themolecule to switch reversibly between colorlessand blue isoforms in the presence or absenceof X-rays. With detection at safe doses, thisreaction system is expected to detect even the
faintest X-rays levels considered dangerous.
Photoreactive materials convert light input
into chemical output and are standard insemiconductor and 3D printing technologies.
Some of these materials are also used ineye-protection, such as how sunglasses can
reduce UV exposure by changing the lens color.
Similarly, workers at risk to X-ray radiation
are required to wear monitoring badges thatindicate dangerous levels through changes inphotoreactive materials.
However, Tsuyoshi Kawai, project leader,stresses that these badges do not completely
The hazardous radiations such as Ultra-Violet and X-ray oxidative triggers the color-changing
eliminate the risk. “Current materials for
cascade of new dyes for 1000 times.
wearable detectors are sensitive to about 1 Gy.
Ideally, safety management systems want about
version between two isoforms. The result was a sensitivity of up to
one hundred times more sensitivity,” he says. Kawai is an expert at
0.3 Gy, making it more than 1000 times more sensitive than current
increasing the photoconversion efficiency of photoreactive molecules,
commercial systems. Notably, 0.3 Gy is considered a safe exposure
having focused his attention primarily on terarylenes, organic
level, suggesting that no dangerous level will go undetected.
molecules with which his research team has consistently achieved exPhotoconversion reactions like photosynthesis or neural
ceptionally high reaction efficiencies. “We have steadily improved thestimulation in response to light in our eyes occurs at less than 100%number of molecules that can undergo photoconversion in response
efficiency (less than one molecule reacts to one photon). The system
to one photon. It was one to one in 2011 and today it becomes 33
designed by the researchers, however, could achieve 3300% (33molecules per one photon,” he says.
molecules per photon), showing the potential of organic molecules in
To increase the quantum yield of terarylenes, is to maximize theartificial systems. “I think this is the highest efficiency ever reported
number of changes that can be induced by a single photon. They havefor photoconversion with an organic molecule,” notes Kawai.
selected terarylenes because of their reversibility, meaning that themolecule can be converted back to the starting blue isoform upon
exposure to ultraviolet light allowing for the system to be reset for
repeated use. Indeed, the color change is one of several reasons hebelieves organic molecules are preferable when considering X-ray
detectors.
“Photochromic organic detectors can report X-rays througheasily observed color changes and are recyclable and easy to process,”
Kawai says. The key modification to the terarylene molecules was
the addition of a phenyl group to only one of the molecules two
phenylthiophene groups, which allowed for reversible photocon-

54

NAIST Research Highlights

Reference

Ryosuke Asato, Colin J. Martin, Jan Patrick Calupitan, Ryo Mizutsu,
Takuya Nakashima, Go Okada, Noriaki Kawaguchi, Takayuki Yanagida,
Tsuyoshi Kawai. 2020. Photosynergetic amplification of radiation input:
from efficient UV induced cycloreversion to sensitive X-ray detection.
Chemical Science, 11, 2504–2510.

Innovative Research and Education Programs
NAIST constantly strives to renew its research and education programs toward producing science andtechnology researchers prepared to meet the demands facing tomorrow’s global scientific community.
These programs are regularly awarded external funding for their wide-ranging benefits.

Program for Promoting the Enhancement of Research Universities (2013-2023)
Japanese Ministry of Education, Culture, Sports, Science andTechnology (MEXT) launched the Program for Promoting theEnhancement of Research Universities in October 2013, which is a
new type of research funding in Japan that aims to enhance researchcapabilities by utilizing University Research Administrators (URA)
who implement intensive reforms to strengthen the research
environment at their respective universities. NAIST is one of 22
universities and research institutions selected to receive support
through this program.
NAIST continues to conduct frontier-opening research while
expanding into new interdisciplinary fields in science and technology. With the establishment of a university-wide strategic research
infrastructure, NAIST endeavors to leverage its abundant resources

Formulating strategies
and plans based on
objective analysis data

to attain the new research materials and facilities necessary fornext-generation research, to disseminate its achievements andhuman resources around the globe, and to further expand its global
research and education network in order to contribute to the overall
advancement of science and technology.
Projects being supported through this program include (1) the
Creating New Research Streams Program which creates new
research domains promising a high global profile, (2) the Sustainable Development of Research Capabilities Program which enhances NAIST’s world-class research capabilities, and (3) the Joint
International Research Program which raises the global visibility
and standing of NAIST’s research capabilities.

Supporting the strategicacquisition of external
competitive research
funds

Enhancing theinternational
collaborative research
network

Reforming the research system to enhance
NAIST’s research capabilities

Top Global University Project (2014-2024)
In October 2014, NAIST was one of 37 universities selected to anotherprestigious MEXT initiative, the Top Global University Project. For aperiod of ten years, MEXT will support outstanding universities intheir efforts to reform institutional governance and collaborate with
top universities worldwide in order to strengthen international
competitiveness.
Through the Top Global University Project, NAIST has committed

to enhancing its international graduate courses by (1) including a
joint degree scheme, (2) developing a new model for graduate education based on top-notch research, (3) reforming institutional
governance and strategic agility, (4) creating a campus environmentthat supports trans-disciplinary education and cultural diversity, and(5) reorganizing its three graduate schools into a single entity toward
establishing new, flexible research groups.

NAIST Research Highlights

55

Outgrow your limits