bluuebunny/crossref_metadata_2025 huggingface dataset:
CellHeap: A scRNA-seq workflow for large-scale bioinformatics data analysis
Authors: Castro Maria Clicia S. , Silva Vanessa S. , Costa Maiana O. C. , Silva Helena S. I. L. , Walter Maria Emilia M. T. , Melo Alba C. M. A. , Ocaña Kary , dos Santos Marcelo T. , Nicolas Marisa F. , Carvalho Anna Cristina C. , Henriques-Pons Andrea , Silva Fabrício A. B.
Published: April 2023

Abstract

Background

Several hundred terabytes of single-cell RNA-seq (scRNA-seq) data are available in public repositories. These data refer to various research projects, from microbial population cells to multiple tissues, involving patients with a myriad of diseases and comorbidities. An increase to several Petabytes of scRNA-seq data available in public repositories is a realistic prediction for coming years. Therefore, thoughtful analysis of these data requires large-scale computing infrastructures and software systems optimized for such platforms to generate correct and reliable biological knowledge.

Results

This paper presents CellHeap, a flexible, portable, and robust platform for analyzing large scRNA-seq datasets, with quality control throughout the execution steps, and deployable on platforms that support large-scale data, such as supercomputers or clouds. As a case study, we designed a workflow to study particular modulations of Fc receptors, considering mild and severe cases of COVID-19. This workflow, deployed in the Brazilian Santos Dumont supercomputer, processed dozens of Terabytes of COVID-19 scRNA-seq raw data. Our results show that most of the workflow total execution time is spent in its initial phases and that there is great potential for a parallel solution to speed up scRNA-seq data analysis significantly. Thus, this workflow includes an efficient solution to use parallel computational resources, improving total execution time. Our case study showed increased Fc receptors transcription in macrophages of patients with severe COVID-19 symptoms, especially FCGR1A, FCGR2A, and FCGR3A. Furthermore, diverse molecules associated with their signaling pathways were upregulated in severe cases, possibly associated with the prominent inflammatory response observed.

Conclusion

From the CellHeap platform, different workflows capable of analyzing large scRNA-seq datasets can be generated. Our case study, a workflow designed to study particular modulations of Fc receptors, considering mild and severe cases of COVID-19, deployed on the Brazilian supercomputer Santos Dumont, had a substantial reduction in total execution time when jobs are triggered simultaneously using the parallelization strategy described in this manuscript. Regarding biological results, our case study identified specific modulations comparing healthy individuals with COVID-19 patients with mild or severe symptoms, revealing an upregulation of several inflammatory pathways and an increase in the transcription of Fc receptors in severe cases.
Global alignment and local curvature of microtubules in mouse fibroblasts are robust against perturbations of vimentin and actin
Authors: Blob Anna , Ventzke David , Rölleke Ulrike , Nies Giacomo , Munk Axel , Schaedel Laura , Köster Sarah
Published: April 2023

The eukaryotic cytoskeleton is an intricate network of three types of mechanically distinct biopolymers – actin filaments, microtubules and intermediate filaments (IFs). These filamentous networks determine essential cellular functions and properties. Among them, microtubules are important for intracellular transport and establishing cell polarity during migration. Despite their intrinsic stiffness, they exhibit characteristic bending and buckling in cells due to non thermal forces acting on them. Interactions between cytoskeletal filaments have been found but are complex and diverse with respect to their effect on the mechanical behavior of the filaments and the architecture of networks. We systematically study how actin and vimentin IFs influence the network structure and local bending of microtubules by analyzing fluorescence microscopy images of mouse fibroblasts on protein micropatterns. Our automated analysis averages over large amounts of data to mitigate the effect of the considerable natural variance in biological cell data. We find that the radial orientation of microtubules in circular cells is robust and is established independently of vimentin and actin networks. Observing the local curvature of microtubules, we find highly similar average bending of microtubules in the entire cell regardless of the cytoskeletal surrounding. Small systematic differences cannot be attributed directly to vimentin and actin densities. Our results suggest that, on average, microtubules in unpolarized mouse fibroblasts are unexpectedly independent of the rest of the cytoskeleton in their global network structure and their local response to forces.
Myosin Light Chain Dephosphorylation by PPP1R12C Promotes Atrial Hypocontractility in Atrial Fibrillation
Authors: Perike Srikanth , Gonzalez-Gonzalez Francisco J. , Abu-Taha Issam , Damen Frederick W. , Lizama Ken S. , Aboonabi Anahita , Capote Andrielle E. , Aguilar-Sanchez Yuriana , Levin Benjamin , Han Zhenbo , Sridhar Arvind , Grand Jacob , Martin Jody , Akar Joseph G. , Warren Chad M. , Solaro R. John , Ong Sang-Ging , Darbar Dawood , Goergen Craig J. , Wolska Beata M. , Dobrev Dobromir , Wehrens Xander H.T. , McCauley Mark D.
Published: April 2023

Background

Atrial fibrillation (AF), the most common sustained cardiac arrhythmia, increases thromboembolic stroke risk five-fold. Although atrial hypocontractility contributes to stroke risk in AF, the molecular mechanisms reducing myofilament contractile function remain unknown. We tested the hypothesis that increased expression of PPP1R12C, the PP1 regulatory subunit targeting atrial myosin light chain 2 (MLC2a), causes hypophosphorylation of MLC2a and results in atrial hypocontractility.

Methods

Right atrial appendage tissues were isolated from human AF patients versus sinus rhythm (SR) controls. Western blots, co-immunoprecipitation, and phosphorylation studies were performed to examine how the PP1c-PPP1R12C interaction causes MLC2a de-phosphorylation.In vitrostudies of pharmacologic MRCK inhibitor (BDP5290) in atrial HL-1 cells were performed to evaluate PP1 holoenzyme activity on MLC2a. Cardiac-specific lentiviral PPP1R12C overexpression was performed in mice to evaluate atrial remodeling with atrial cell shortening assays, echocardiography, and AF inducibility with EP studies.

Results

In human patients with AF, PPP1R12C expression was increased two-fold versus SR controls (P=2.0×10−2, n=12,12 in each group) with > 40% reduction in MLC2a phosphorylation (P=1.4×10−6, n=12,12 in each group). PPP1R12C-PP1c binding and PPP1R12C-MLC2a binding were significantly increased in AF (P=2.9×10−2and 6.7×10−3respectively, n=8,8 in each group).In vitrostudies utilizing drug BDP5290, which inhibits T560-PPP1R12C phosphorylation, demonstrated increased PPP1R12C binding with both PP1c and MLC2a, and dephosphorylation of MLC2a. Lenti-12C mice demonstrated a 150% increase in LA size versus controls (P=5.0×10−6, n=12,8,12), with reduced atrial strain and atrial ejection fraction. Pacing-induced AF in Lenti-12C mice was significantly higher than controls (P=1.8×10−2and 4.1×10−2respectively, n= 6,6,5).

Conclusions

AF patients exhibit increased levels of PPP1R12C protein compared to controls. PPP1R12C overexpression in mice increases PP1c targeting to MLC2a and causes MLC2a dephosphorylation, which reduces atrial contractility and increases AF inducibility. These findings suggest that PP1 regulation of sarcomere function at MLC2a is a key determinant of atrial contractility in AF.
Apolipoprotein B and Interleukin 1 Receptor Antagonist: Reversing the Risk of Coronary Heart Disease
Authors: Yang Fangkun , Huangfu Ning , Shen Jiaxi , Su Pengpeng , Zhu Lujie , Cui Hanbin , Yuan Shuai
Published: April 2023

Background

Epidemiological evidence for the link of interleukin 1 (IL-1) and its inhibition with coronary heart disease (CHD) remains controversial. We aim to investigate the cardiovascular effects of IL-1 receptor antagonist (IL-1Ra) and underlying mechanisms, as well as the potential interaction with lifestyle factors.

Methods

A comprehensive multivariable Mendelian randomization study was performed. Genetic variants identified from a genome-wide association study involving 30,931 individuals were used as instrumental variables for the serum IL-1Ra concentrations. Genetic associations with CHD (60,801 cases and 123,504 controls) were extracted from the CARDIoGRAMplusC4D consortium. Inverse-variance weighted method was utilized to derive effect estimates, while supplementary analyses employing various statistical approaches.

Results

Genetically determined IL-1Ra level was associated with increased risk of CHD (odds ratio (OR), 1.07; 95% CI: 1.03-1.17) and myocardial infarction (OR, 1.13; 95% CI: 1.04-1.21). The main results remained consistent in supplementary analyses. Besides, IL-1Ra was associated with circulating levels of various lipoprotein lipids, apolipoproteins and fasting glucose. Interestingly, observed association pattern with CHD was reversed when adjusting for apolipoprotein B (OR, 0.84; 95%CI: 0.71-0.99) and slightly attenuated on accounting for other cardiometabolic risk factors. Appropriate lifestyle intervention was found to lower IL-1Ra concentration and mitigate the heightened CHD risk it posed.

Conclusions

Apolipoprotein B represents the key driver, and a potential target for reversal of the causal link between serum IL-1Ra and increased risk of CHD/MI. The combined therapy involving IL-1 inhibition and lipid-modifying treatment aimed at apolipoprotein B merit further exploration.
Structural remodeling of the endoplasmic reticulum in response to extracellular signals requires αTAT1-induced microtubule acetylation
Authors: Ortiz Hannah R. , Flores Paola Cruz , Ramonett Aaron , Ahmed Tasmia , Ellis Nathan A. , Langlais Paul R. , Mythreye Karthikeyan , Lee Nam Y.
Published: April 2023

Dynamic changes in the endoplasmic reticulum (ER) morphology are central to maintaining cellular homeostasis. Microtubules (MT) facilitate the continuous remodeling of the ER network into sheets and tubules by coordinating with many ER-shaping protein complexes, although how this process is controlled by extracellular signals remains unknown. Here we report that TAK1, a kinase responsive to numerous growth factors and cytokines including TGF-β and TNF-α, triggers ER tubulation by activating αTAT1, an MT-acetylating enzyme that enhances ER-sliding. We show that this TAK1/αTAT-dependent ER remodeling promotes cell survival by actively downregulating BOK, an ER membrane-associated proapoptotic effector. While BOK is normally protected from degradation when complexed with IP3R, it is rapidly degraded upon their dissociation during the ER sheets-to-tubules conversion. These findings demonstrate a distinct mechanism of ligand-induced ER remodeling and suggest that the TAK1/αTAT pathway may be a key target in ER stress and dysfunction.
Changes to Astrocyte-associated Protein Expression at Different Timepoints of Cuprizone Treatment
Authors: Frankle Lana , Riley Amanda , Tomor Riely , Lee Hannah , Jarzembak Kole , Benedict Olesia , Sternbach Sarah , Shelestak John , McDonough Jennifer , Clements Robert
Published: April 2023

Glial cells, including astrocytes, microglia, and oligodendrocytes, are brain cells that support and dynamically interact with neurons and each other. These intercellular dynamics undergo changes during stress and disease states. In response to most forms of stress, astrocytes will undergo some variation of activation, meaning upregulation in certain proteins expressed and secreted and either upregulations or downregulations to various constitutive and normal functions. While types of activation are many and contingent on the particular disturbance that triggers these changes, there are two main overarching categories that have been delineated thus far: A1 and A2. Named in the convention of microglial activation subtypes, and with the acknowledgement that the types are not completely distinct or completely comprehensive, the A1 subtype is generically associated with toxic and pro-inflammatory factors, and the A2 phenotype is broadly associated with anti-inflammatory and neurogenic factors. The present study served to measure and document dynamic changes in these subtypes at multiple timepoints using an established experimental model of cuprizone toxic demyelination. The authors found increases in proteins associated with both cell types at different timepoints, with protein increases in the A1 marker C3d and the A2 marker Emp1 in the cortex at one week and protein increases in Emp1 in the corpus callosum at three days and four weeks. There were also increases in Emp1 staining specifically colocalized with astrocyte staining in the corpus callosum at the same timepoints as the protein increases, and in the cortex weeks later at four weeks. C3d colocalization with astrocytes also increased most at four weeks. This indicates simultaneous increases of both types of activation as well as the likely existence of astrocytes expressing both markers. The authors also found the increase in two A1 associated proteins (TNF alpha and C3d) did not show a linear relationship in line with findings from other research and indicating a more complex relationship between cuprizone toxicity and astrocyte activation. The increases in TNF alpha and IFN gamma did not occur at timepoints preceding increases in C3d and Emp1, showing that other factors also precipitate the subtypes associated (A1 for C3d and A2 for Emp1). These findings add to the body of research showing the specific early timepoints at which A1 and A2 markers are most increased during the course of cuprizone treatment, including the fact that these increases can be non-linear in the case of Emp1. This provides additional information on optimal times for targeted interventions during the cuprizone model.
Decoding humanin vitroterminal erythropoiesis originating from umbilical cord blood mononuclear cells and pluripotent stem cells
Authors: Wang Xiaoling , Zhang Wei , Zhao Siqi , Yan Hao , Xin Zijuan , Cui Tiantian , Zang Ruge , Zhao Lingping , Wang Haiyang , Zhou Junnian , Li Xuan , Yue Wen , Xi Jiafei , Zhang Zhaojun , Fang Xiangdong , Pei Xuetao
Published: April 2023

Ex vivo RBC production generates unsatisfactory expansion, β-globin expression, and maturation of erythroid cells. The underlying mechanisms behind these limitations and ex vivo terminal erythropoiesis from different origins are largely unexplained. In this study, we mapped an atlas of ex vivo terminally differentiated cells from umbilical cord blood mononuclear cells (UCBMNs) and pluripotent stem cells (PSCs), and observed the differential regulatory dynamics of erythropoiesis from these two origins at a single-cell resolution. We detected the presence of hematopoietic stem progenitor cells (HSPCs), erythroid progenitor (e.g., CFU-E), and non-erythroid cells (e.g., macrophages) in the terminal populations. We observed that UCBMN-derived erythropoiesis is more active than PSC-derived erythropoiesis in terms of the cell cycle, stress erythropoiesis, and autophagy at single cell resolution, which may provide new insights into the limitations in cell expansion, globin expression, and maturation in ex vivo RBC production, respectively. We verified that a stress-erythropoiesis-related gene,TRIB3, increases the expression of globin genes in ex vivo erythropoiesis. As the major unexpected component detected in terminally differentiated cells, CFU-E were further characterized as having high- or low- expansion capacity based on CD99 expression, which generally decreased over erythropoiesis. By inhibiting CD99 gene expression using antagonists, we increased reticulocyte production in the population. Heterogeneous CFU-Es also exist in bone marrow. Moreover, decreased CD99 expression mediates the interactions between macrophages and CFU-E during ex vivo erythropoiesis. Overall, our results provide a reference for facilitating the development of strategies to improve ex vivo RBC regeneration.

Highlights

We performed scRNA-seq and cell typing of late stage UCBMN- and PSC-derived cells

Stress erythropoiesis, autophagy and cell cycle related gene expression different in two origins

CD99highprogenitor cells are a proliferating colony forming unit erythroid subpopulation
Immediate effect of quadri-pulse stimulation on human brain microstructures
Authors: Kimura Ikko , Hayashi Masamichi J , Amano Kaoru
Published: April 2023

Several studies have implied that human brain microstructures can change immediately after a behavioral training. However, since widespread regions are involved in behavioral training, it remains unclear whether the microstructure in the living human brain changes immediately after the change in activity of a specific brain area. Hence, we aimed to examine whether the microstructures in the human brain change after the increase and decrease in the specific brain activity by repetitive transcranial magnetic stimulation, namely quadri-pulse stimulation (QPS). Right-handed healthy adults underwent both the excitatory (QPS5) and inhibitory (QPS50) QPS protocols over the left M1. Before and after QPS, diffusion MRI and resting-state fMRI scans were collected to detect any microstructural (fractional anisotropy [FA] and mean diffusivity [MD] values) and functional (functional connectivity between the bilateral M1) changes after QPS5 and QPS50. As a result, we observed no statistically significant change in FA or MD values after either QPS5 or QPS50 in cerebral cortex. This suggests that the brain activity change in widespread area is required to induce microstructural change immediately.
Detect+Track: Robust and flexible software tools for improved tracking and behavioural analysis of fish
Authors: Dutta Abhishek , Pérez-Campanero Natalia , Taylor Graham K. , Zisserman Andrew , Newport Cait
Published: April 2023

Developments in automated animal behavioural analysis software are increasing the efficiency of data collection and improving the standardization of behavioural measurements. There are now several open-source tools for tracking laboratory animals, but often these are only accurate under limited conditions (e.g. uniform lighting and background, uncluttered scenes, unobstructed focal animal). Tracking fish presents a particular challenge for these tools because movement at the water’s surface introduces significant noise. Partial occlusion of the focal animal can also be troublesome, particularly when tracking the whole organism. But identifying the position of an animal is only part of the task – analysing the movement of the animal relative to their environment and experimental context is often what provides information about their behaviour. Therefore, the automated detection of physical objects and boundaries would also be beneficial, but this feature is not commonly incorporated into existing tracking software. Here we describe a video processing method that uses a range of computer vision algorithms (e.g. object detector and tracker, optical flow, parallel plane homology) and computational geometry techniques (e.g. Voronoi tessellation) to analyse the movement behaviour of fish in response to experimental stimuli. A behavioural experiment, which involved tracking a fish’s trajectory through a field of obstacles, motivated our development of a set of tools that: (1) measure an animal’s trajectory, (2) record obstacle position, and (3) detect when the fish passed through ‘virtual gates’ between adjacent obstacles and/or the aquarium wall. We have introduced a novel Detect+Track approach that significantly enhances the accuracy and robustness of animal tracking, overcoming some of the limitations of existing tools and providing a more reliable solution for complex experimental conditions. Our workflow is divided into several discrete steps, and provides a set of modular software building blocks that can be adapted to analyse other experimental designs. A detailed tutorial is provided, together with all the data and code required to reproduce our results.
Cell-cycle regulator Whi5 shapes proliferative heterogeneity in clonal populations
Authors: Delgado-Román Irene , García-Marcelo María José , Delgado-Ramos Lidia , Chávez Sebastián , Muñoz-Centeno Mari Cruz
Published: April 2023

Clonal populations exhibit phenotypic variation in spite of being composed of genetically identical cells under equal environmental conditions. Proliferation rate also shows this heterogeneity, but the mechanisms underlying are still poorly understood. Cell cycle regulation controls proliferative capacity and previous transcriptomic studies revealed that budding yeast microcolonies with low proliferation rates display high levels of the G1-S transition inhibitor Whi5. In this work, we combined single cell microencapsulation with confocal microscopy to study heterogeneity in clonal cultures. We found that most slow-growing microcolonies are founded by young mother cells with a very short number of cell division cycles. This reduction in proliferation capacity is related to the expression levels of Whi5, which increases with the number of mother cell replication rounds since early stages. Our results establish that the proliferative structure of a cell population is cumulatively shaped in every mitotic cycle from the very first division, when a mother cell has the opportunity to increase Whi5 expression levels and to develop a slow proliferating lineage.
Host use preference of fruit flies (Diptera: Tephritidae) among selected cucurbitaceous vegetables in Morogoro, Eastern-Central Tanzania
Authors: Tarimo Petronila , Kabota Sija , Mwatawala Maulid , Majubwa Ramadhan , Kudra Abdul , Virgilio Massimiliano , Jordaens Kurt , De Meyer Marc
Published: April 2023

Fruit flies (Diptera, Tephritidae) represent a major threat to cucurbit production in Tanzania. They cause huge economic losses on cucurbit crops. Information on the infestation and yield loss caused by fruit flies on cucurbit crops is vital in designing a sound fruit fly management program. We identified the fruit fly species attacking cucurbit crops (Cucurbitales: Cucurbitaceae): cucumber (Cucumis sativusL.), watermelon (Citrullus lanatus[Thunb.] Matsum. & Nakai) and squash (Cucurbita moschataD.) in the Morogoro region and quantified their incidence and infestation per crop species. A weekly sampling of ten fruits per crop was repeated twice per plot for each zone for five consecutive weeks between March and October 2020. The number of fruit flies emerging from the collected fruits was quantified and infestation per crop species was determined. We observed significant differences in infestation between cucurbit species, across zones and seasons. Squash showed the highest infestation ofZeugodacus cucurbitae(Coquillet) in both zones followed by watermelon. Watermelon was highly infested byDacus vertebratus(Bigot) in the plateau zone followed by squash. Squash showed the highest infestation ofDacus ciliatusLoew in the plateau zone followed by watermelon. Cucumber had the lowest infestation for all fruit fly species.
DNA-m6A calling and integrated long-read epigenetic and genetic analysis with fibertools
Authors: Jha Anupama , Bohaczuk Stephanie C. , Mao Yizi , Ranchalis Jane , Mallory Benjamin J. , Min Alan T. , Hamm Morgan O. , Swanson Elliott , Dubocanin Danilo , Finkbeiner Connor , Li Tony , Whittington Dale , Noble William Stafford , Stergachis Andrew B. , Vollger Mitchell R.
Published: April 2023

Long-read DNA sequencing has recently emerged as a powerful tool for studying both genetic and epigenetic architectures at single-molecule and single-nucleotide resolution. Long-read epigenetic studies encompass both the direct identification of native cytosine methylation as well as the identification of exogenously placed DNAN6-methyladenine (DNA-m6A). However, detecting DNA-m6A modifications using single-molecule sequencing, as well as co-processing single-molecule genetic and epigenetic architectures, is limited by computational demands and a lack of supporting tools. Here, we introducefibertools, a state-of-the-art toolkit that features a semi-supervised convolutional neural network for fast and accurate identification of m6A-marked bases using PacBio single-molecule long-read sequencing, as well as the co-processing of long-read genetic and epigenetic data produced using either PacBio or Oxford Nanopore sequencing platforms. We demonstrate accurate DNA-m6A identification (>90% precision and recall) along >20 kilobase long DNA molecules with a ∼1,000-fold improvement in speed. In addition, we demonstrate thatfibertoolscan readily integrate genetic and epigenetic data at single-molecule resolution, including the seamless conversion between molecular and reference coordinate systems, allowing for accurate genetic and epigenetic analyses of long-read data within structurally and somatically variable genomic regions.
Conservation of dichromatin organization along regional centromeres
Authors: Dubocanin Danilo , Hartley Gabrielle A. , Cortes Adriana E. Sedeno , Mao Yizi , Hedouin Sabrine , Ranchalis Jane , Agarwal Aman , Logsdon Glennis A. , Munson Katherine M. , Real Taylor , Mallory Benjamin J. , Eichler Evan E. , Biggins Sue , O’Neill Rachel J. , Stergachis Andrew B.
Published: April 2023

The focal attachment of the kinetochore to the centromere is essential for genome maintenance, yet the highly repetitive nature of satellite regional centromeres, such as those in humans, limits our understanding of their chromatin organization. We demonstrate that single-molecule chromatin fiber sequencing (Fiber-seq) can uniquely co-resolve kinetochore and surrounding chromatin architectures along point centromeres, revealing largely homogeneous single-molecule kinetochore occupancy along each chromosome. In contrast, extension of Fiber-seq to regional satellite centromeres exposed marked per-molecule heterogeneity in their chromatin organization. Regional CENP-A-marked centromere cores uniquely contain a dichotomous chromatin organization (dichromatin) composed of compacted nucleosome arrays punctuated with highly accessible chromatin patches. CENP-B occupancy phases dichromatin to the underlying alpha-satellite repeat within centromere cores, but is not necessary for dichromatin formation. Centromere core dichromatin is a conserved feature between humans despite the marked divergence of their underlying alpha-satellite organization and is similarly a conserved feature along regional centromeres that lack satellite repeats in gibbon. Overall, the chromatin organization of regional centromeres is defined by marked per-molecule heterogeneity, likely buffering kinetochore attachment against sequence and structural variability within regional centromeres.

Highlights

Dichotomous accessible and compacted chromatin (dichromatin) marks centromere cores

Highly accessible chromatin patches punctuate sites of kinetochore attachment

Dichromatin can form irrespective of CENP-B occupancy

Conservation within centromeres is mediated at the level of chromatin, not DNA
A mouse model for environmentally induced and reversible circadian arrhythmia using gradual exposure to a fragmented day-night cycle
Authors: Richardson Melissa E. S. , Browne Chérie-Akilah , Huerta Mazariegos Citlali I.
Published: April 2023

Arrhythmia is considered the most disrupted state of the biological circadian clock, and usually occurs when circadian regulatory genes are rendered non-functional, or the master clock (Suprachiasmatic Nucleus) is ablated. Since clock gene expression is aligned by the external solar day-night cycle to exhibit a 24-hour rhythm, we hypothesized that ill-timed light and dark exposure could negatively impact endogenous circadian clock function in mice. In this study, we present an environmentally driven approach to induce arrhythmia in mice that is also reversible. Using the previously characterized fragmented day-night cycle (FDN) where the 8-hour night is split into four 2-hour fragments and equally distributed across the 24-hour day, we show that mice gradually exposed to the FDN for 1 month lose their circadian rhythmicity. Furthermore, subsequent exposure to constant light or constant dark conditions does not yield typical circadian rhythms, but instead, reveals circadian arrhythmia. Finally, we show that the arrhythmic locomotion phenotype is reversible with one week of reintroduction to a 12 hr day-12 hr night cycle. This is the first study to show how the light-dark environment induces arrhythmia of an intact circadian clock and how it can be reversed.
Tailored chemical reactivity probes for systemic imaging of aldehydes in fibroproliferative diseases
Authors: Ma Hua , Zhou Iris Y. , Chen Y. Iris , Rotile Nicholas J. , Ay Ilknur , Akam Eman , Wang Huan , Knipe Rachel , Hariri Lida P. , Zhang Caiyuan , Drummond Matthew , Pantazopoulos Pamela , Moon Brianna F. , Boice Avery T. , Zygmont Samantha E. , Weigand-Whittier Jonah , Sojoodi Mozhdeh , Gonzalez-Villalobos Romer A. , Hansen Michael K. , Tanabe Kenneth K. , Caravan Peter
Published: April 2023

During fibroproliferation, protein-associated extracellular aldehydes are formed by the oxidation of lysine residues on extracellular matrix proteins to form the aldehyde allysine. Here we report three Mn(II)-based, small molecule magnetic resonance (MR) probes that contain α-effect nucleophiles to target allysine in vivo and report on tissue fibrogenesis. We used a rational design approach to develop turn-on probes with a 4-fold increase in relaxivity upon targeting. The effects of aldehyde condensation rate and hydrolysis kinetics on the performance of the probes to detect tissue fibrogenesis noninvasively in mouse models were evaluated by a systemic aldehyde tracking approach. We showed that for highly reversible ligations, off-rate was a stronger predictor of in vivo efficiency, enabling histologically validated, three-dimensional characterization of pulmonary fibrogenesis throughout the entire lung. The exclusive renal elimination of these probes allowed for rapid imaging of liver fibrosis. Reducing the hydrolysis rate by forming an oxime bond with allysine enabled delayed phase imaging of kidney fibrogenesis. The imaging efficacy of these probes, coupled with their rapid and complete elimination from the body, make them strong candidates for clinical translation.
HB21/40/53 promote inflorescence arrest through ABA accumulation at the end of flowering
Authors: Sánchez-Gerschon Verónica , Ferrándiz Cristina , Balanzà Vicente
Published: April 2023

Flowers are produced by the activity of the inflorescence meristem after the floral transition. In plants with indeterminate inflorescences, as Arabidopsis, the final number of flowers produced by the inflorescence meristem will depend on two main factors, the rate of flower production by the meristem and the duration of the phase of inflorescence meristem activity. The end of flowering, understood as the moment when the inflorescence stops the production of new flowers, is associated with the meristem proliferative arrest. At this time point, the meristem ceases to initiate new floral primordia and the unpollinated flowers already formed arrest their development.

It has been known for a long time that fruit/seed production induces inflorescence meristem arrest, but the mechanisms controlling this process were elusive. During the last years, the regulation of the end of flowering has started to be elucidated in Arabidopsis. The meristem arrest at the end of flowering is controlled at the genetic level by the FRUITFULL-APETALA2 (FUL-AP2) pathway, that modulates meristem activity. The meristem arrest has been also shown to be controlled at the hormonal level. It has been proposed that auxin could mediate the fruit/seed effect to the meristem. Cytokinins regulation and response have been also proposed as important factors controlling the meristem activity at the end of flowering. Finally, it has been also described that arrested meristems at the end of flowering resembles dormant meristem at the transcriptomic level.

Previously, we have shown that the FUL-AP2 pathway controls the expression of the homeodomain leucine zipper transcription factorHOMEOBOX PROTEIN 21(HB21), a gene involved in the establishment of bud axillary dormancy. In this work we characterize the role ofHB21in the control of the proliferative arrest associated with the end of flowering. We observed thatHB21, together withHB40andHB53, accumulate in the inflorescence apexes at the end of flowering promoting the cessation of inflorescence meristem activity. We also show thatHB21induction of in young apexes is sufficient to induce flower and meristem arrest, likely mediated by an increase in ABA responses. Thus, our work confirms the parallelism proposed between dormant meristems and the arrested meristem at the end of flowering, which appear to be regulated by common pathways, and propose ABA as a new regulator in the control of inflorescence meristem arrest.
An isoform-resolution transcriptomic atlas of colorectal cancer from long-read single-cell sequencing
Authors: Li Zhongxiao , Zhang Bin , Chan Jia Jia , Tabatabaeian Hossein , Tong Qing Yun , Chew Xiao Hong , Fan Xiaonan , Driguez Patrick , Chan Charlene , Cheong Faith , Wang Shi , Siew Bei En , Tan Ian Jse-Wei , Lee Kai-Yin , Lieske Bettina , Cheong Wai-Kit , Kappei Dennis , Tan Ker-Kan , Gao Xin , Tay Yvonne
Published: April 2023

Colorectal cancer (CRC) is the second leading cause of cancer death worldwide. In recent years, short-read single-cell RNA sequencing (scRNA-seq) has been instrumental in deciphering tumor cell heterogeneities. However, these studies only enable gene-level expression quantification but neglect alterations in transcript structures, which arise from alternative end processing or splicing, and are frequently observed in cancer. In this study, we integrated short- and long-read scRNA-seq of CRC patient samples to build the first isoform-resolution CRC transcriptomic atlas. We identified 394 dysregulated transcript structures in tumor epithelial cells, including 299 resulting from various combinations of multiple splicing events. Secondly, we characterized genes and isoforms associated with epithelial lineages and subpopulations that exhibit distinct prognoses. Finally, we built an algorithm that integrated novel peptides derived from predicted ORFs of recurrent tumor-specific transcripts with mass spectrometry data and identified a panel of recurring neoepitopes that may aid the development of neoantigen-based cancer vaccines.
Metabolic predictors of response to immune checkpoint blockade therapy
Authors: Shorer Ofir , Yizhak Keren
Published: April 2023

Metabolism of immune cells in the tumor microenvironment (TME) plays a critical role in cancer patient response to immune checkpoint inhibitors (ICI). Yet, a metabolic characterization of immune cells in the TME of patients treated with ICI is lacking. To bridge this gap we performed a semi-supervised analysis of ∼1700 metabolic genes using single-cell RNA-seq data of >1 million immune cells from ∼230 tumor and blood samples treated with ICI. When clustering cells based on their metabolic gene expression, we found that similar immunological states are found in different metabolic states. Most importantly, we found metabolic states that are significantly associated with patient response. We then built a metabolic predictor based on a dozen gene signature which significantly differentiates between responding and non-responding patients across different cancer types (AUC = 0.8-0.86). Taken together, our results demonstrate the importance of metabolism in predicting patient response to ICI.
Mammary microvessels are sensitive to menstrual cycle sex hormones
Authors: Moccia Carmen , Cherubini Marta , Fortea Marina , Akinbote Akinola , Padmanaban Prasanna , Sastre Violeta Beltran , Haase Kristina
Published: April 2023

The mammary gland is a highly vascularized organ that is influenced by sex hormones including estrogen (E2) and progesterone (P4). Beyond whole-organism studies in rodents or 2D monocultures, hormonal interactions and their effects on the breast microvasculature remains largely understudied. Recent methods to generate 3D microvessels on-chip have enabled direct observation of complex vascular processes; however, these models often use non-tissue-specific cell types, such as HUVEC and fibroblasts from various sources. Here, novel mammary-specific microvessels are generated by co-culturing primary breast endothelial cells and fibroblasts under optimized culture conditions. These microvessels are mechano-sensitive (to interstitial flow) and require endothelial-stromal interactions to develop fully perfusable vessels. These mammary-specific microvessels are also responsive to exogenous stimulation by sex hormones. When treated with combined E2 and P4, corresponding to the four phases of the menstrual cycle (period, follicular, ovular, and luteal), vascular remodeling and barrier function are altered in a phase-dependent manner. The presence of high E2 (ovulation) promotes vascular growth and remodeling, corresponding to high depletion of proangiogenic factors, whereas high P4 concentrations (luteal) promote vascular regression. The effects of combined E2 and P4 hormones are not only dose-dependent but also tissue-specific, as is shown by similarly treating non-tissue-specific HUVEC microvessels.
Cues for predictive eye movements in naturalistic scenes
Authors: Goettker Alexander , Borgerding Nils , Leeske Linus , Gegenfurtner Karl R.
Published: April 2023

We previously compared following of the same trajectories with eye movements, but either as an isolated target or embedded in a naturalistic scene, in this case the movement of a puck in an ice hockey game. We observed that the oculomotor system was able to leverage the contextual cues available in the naturalistic scene to produce predictive eye movements. In this study we wanted to assess which factors are critical for achieving this predictive advantage by manipulating four factors: the expertise of the viewers, the amount of available peripheral information, and contextual and kinematic cues. The more peripheral information became available (by manipulating the area of the video that was visible), the better the predictions of all observers. However, expert ice hockey fans were consistently more accurate and better at predicting than novices and also benefitted more from additional peripheral information. Artificial contextual cues about the position of the players did not lead to a predictive advantage, whereas impairing the causal structure of kinematic cues by playing the video in reverse led to a severe impairment. When the videos were flipped vertically to introduce more difficult kinematic cues, predictive behavior was comparable to when observers watching the original videos. Together, these results demonstrate that when contextual information is available in naturalistic scenes, the oculomotor system is successfully integrating them, and is not only relying on low-level information about the target trajectory. Critical factors for successful prediction seem to be the amount of available information, experience with the stimuli and the availability of intact kinematic cues for player movements.
Intrinsic functional connections determine how curiosity and prediction errors enhance memory
Authors: Eschmann Kathrin C. J. , Valji Ashvanti , Graham Kim S. , Lawrence Andrew D. , Gruber Matthias J.
Published: April 2023

Individuals differ in the way they seek information, acquire knowledge, and form memories. Neural finger-prints of intrinsic functional connectivity distinguish between individuals and predict inter-individual differences in task performance. Both curiosity – the desire to acquire new information – and information prediction errors (IPEs) – the mismatch between information and previous expectations – enhance memory but differ considerably between individuals. The present study assessed whether inter-individual differences in functional connectivity measured using resting-state fMRI determine the extent to which individuals benefit from memory-enhancing effects of curiosity and IPEs. We found a double dissociation between individual differences in mesolimbic functional connectivity, which accounted for curiosity-driven but not IPE-related memory enhancements, and individual differences in cingulo-hippocampal functional connectivity, which predicted IPE-driven but not curiosity-related memory enhancements. These novel findings on how inter-individual differences in dissociable intrinsic functional networks determine memory enhancements stress the need to account for these differences in theoretical frameworks of curiosity and memory.
Mapping host-microbe transcriptional interactions by dual perturb-seq
Authors: Butterworth Simon , Kordova Kristina , Chandrasekaran Sambamurthy , Thomas Kaitlin K. , Torelli Francesca , Lockyer Eloise J. , Edwards Amelia , Goldstone Robert , Koshy Anita A. , Treeck Moritz
Published: April 2023

Intracellular pathogens and other endosymbionts reprogram host cell transcription to suppress immune responses and recalibrate biosynthetic pathways. This reprogramming is critical in determining the outcome of infection or colonisation. Here, we combine pooled CRISPR knockout screening with dual host–microbe single-cell RNA-sequencing to identify the molecular mediators of these transcriptional interactions, a method we term dual perturb-seq. Applying dual perturb-seq to the intracellular pathogenToxoplasma gondii, we are able to identify previously uncharacterised effector proteins and directly infer their function from the transcriptomic data. We show thatTgGRA59 contributes to the export of other effector proteins from the parasite into the host cell and identify a novel effector,TgSOS1, that is necessary for sustained host STAT6 signalling and thereby contributes to parasite immune evasion and persistence. Together, this work demonstrates a novel tool that can be broadly adapted to interrogate host-microbe transcriptional interactions and reveal mechanisms of infection and immune evasion.
Experience Shapes Chandelier Cell Function and Structure in the Visual Cortex
Authors: Seignette Koen , Jamann Nora , Papale Paolo , Terra Huub , Porneso Ralph P. O. , de Kraker Leander , van der Togt Chris , van der Aa Maaike , Neering Paul , Ruimschotel Emma , Roelfsema Pieter R. , Montijn Jorrit S. , Self Matthew W. , Kole Maarten H. P. , Levelt Christiaan N.
Published: April 2023

Detailed characterization of interneuron types in primary visual cortex (V1) has greatly contributed to understanding visual perception, yet the role of chandelier cells (ChCs) in visual processing remains poorly characterized. Using viral tracing we found that V1 ChCs predominantly receive monosynaptic input from local layer 5 pyramidal cells and higher-order cortical regions. Two-photon calcium imaging and convolutional neural network modelling revealed that ChCs are visually responsive but weakly selective for stimulus content. In mice running in a virtual tunnel, ChCs respond strongly to events known to elicit arousal, including locomotion and visuomotor mismatch. Repeated exposure of the mice to the virtual tunnel was accompanied by reduced visual responses of ChCs and structural plasticity of ChC boutons and axon initial segment length. Finally, ChCs only weakly inhibited pyramidal cells. These findings suggest that ChCs provide an arousal-related signal to layer 2/3 pyramidal cells that may modulate their activity and/or gate plasticity of their axon initial segments during behaviorally relevant events.
Thermoregulatory consequences of growing up during a heatwave or a cold snap
Authors: Persson Elin , Cuív Ciarán Ó , Nord Andreas
Published: April 2023

Changes in environmental temperature during the developmental period can affect growth, metabolism, and temperature tolerance of the offspring. We know little about whether such changes remain to adulthood, which is important to understand the links between climate change, development, and fitness. We investigated if phenotypic consequences of the thermal environment in early life remained in adulthood in two studies on Japanese quail (Coturnix japonica). Birds were raised under simulated heatwave-, cold snap-or control conditions, from hatching until halfway through the growth period, and then in a common garden until reproductively mature. We measured biometric and thermoregulatory (metabolic heat production [MHP], evaporative water and heat loss [EWL, EHL] and body temperature) responses to variation in submaximal air temperature at the end of the thermal acclimation period and in adulthood. Warm birds had lower MHP than control birds at the end of the thermal acclimation period and, in the warmest temperature studied (40°C), also had higher evaporative cooling capacity compared to controls. No analogous responses were recorded in cold birds, though they had higher EWL than controls in all but the highest test temperature. None of the effects found at the end of the heatwave-or cold snap period remained until adulthood. This implies that chicks exposed to higher temperatures could be more prepared to counter heat stress as juveniles, but that they do not enjoy any advantages of such developmental conditions when facing high temperatures as adults. Conversely, cold temperature does not seem to confer any priming effects in adolescence.
Worldwide population genomics reveal long-term stability of the mitochondrial chromosome composition in a keystone marine plant
Authors: Khachaturyan Marina , Reusch Thorsten B. H. , Dagan Tal
Published: April 2023

Mitochondrial genomes (mitogenomes) of flowering plants are comprised of multiple chromosomes. Their copy number and composition can be dynamic within and among individual plants due to uneven replication of the chromosomes and homologous recombination. Nonetheless, despite their functional importance, the level of mitogenome conservation within species remains understudied. Whether the ontogenetic variation translates to evolution of mitogenome composition over generations is currently unknown. Here we show that the mitochondrial chromosome composition of the seagrassZostera marinais conserved among worldwide populations that diverged ca 350,000 years ago. Using long-read sequencing we characterized theZ. marinamitochondrial genome and inferred the repertoire of recombination-induced configurations of its eight chromosomes. To characterize the chromosome composition worldwide and study its evolution we examined the mitogenome inZ. marinameristematic region sampled in 16 populations from the Pacific and Atlantic oceans. Our results reveal a striking similarity in the chromosome copy number suggesting stable equal proportions among distantly related populations and a high conservation of the mitochondrial genome within the plant germline, despite a notable variability during individual ontogenesis. Our study supplies a link between observations of dynamic mitogenomes at the level of plant individuals and long-term mitochondrial evolution.

Significance statement

Extensive studies on evolution of plant mitochondria in individual plants revealed great variability of the mitogenome architecture across tissues, however, data on the mitochondrion evolution at the population level is still scarce. We show that the mitochondrial genome architecture in a keystone marine plant,Zostera marina, remained conserved over ca. 350,000 years worldwide. We suggest that the extreme conservation of theZ. marinamitochondria is a manifestation of streamlined mitochondria inheritance over plant generations, e.g., via a plant germline.