bluuebunny/crossref_metadata_2025 huggingface dataset:
A Crispr Knockout Screen Identifies Foxf1 as a Suppressor of Colorectal Cancer Metastasis that Acts through Reduced mTOR Signalling
Authors: Lee Lennard , Woolley Connor , Starkey Thomas , Freeman-Mills Luke , Bassett Andrew , Franchini Fanny , Wang Lai Mun , Lewis Annabelle , Arnold Roland , Tomlinson Ian
Published: April 2019

Introduction

A greater understanding of molecular mechanisms underlying metastasis is necessary for development of new strategies to prevent and treat cancer.

Methods

We performed a genome-wide CRISPR/Cas9 knockout screen in MC38 colorectal cancer (CRC) cells transplanted orthotopically into mice to identify genes that promote metastasis. We undertook focussed molecular analyses to identify mechanisms underlying metastasis.

Results

The screen identified several gene knockouts over-represented in lung metastases, including Dptor (mTOR signalling) and Foxf1 (gastrointestinal tumour predisposition). We validate that loss of Foxf1 promotes metastasis, increased Foxf1 expression restrained cellular migration in-vitro and human CRC metastases express lower Foxf1 than paired primary tumours. Analysis of gene expression changes downstream of Foxf1 identified increased mTOR signalling as a possible mechanism of metastasis caused by Foxf1 loss, consistent with Dptor identification. We confirmed this mechanism demonstrating that mTOR inhibitor sirolimus reduced lung metastasis burden in xenografts.

Conclusion

Mesenchymal Foxf1 plays a major role in intestinal development. We have shown for the first time, through an unbiased genetic screen, that reduced epithelial Foxf1 results in raised mTOR signalling and metastasis.

Authorship statement

Lennard Lee-study concept and design, acquisition of data, analysis, interpretation of data, drafting of the manuscript, statistical analysis and obtained funding. Connor Woolley-acquisition of data, analysis and interpretation of data. Thomas Starkey-acquisition of data, analysis, interpretation of data, drafting of the manuscript. Luke Freeman-Mills-interpretation of data. Andrew Bassett-technical and material support. Fanny Fanchini-technical support. Lai Mun Wang-acquisition of data and study supervision. Annabelle Lewis-study supervision. Roland Arnold-analysis, interpretation of data, statistical analysis. Ian Tomlinson-study supervision and critical revision of the manuscript.

Conflict of Interest

The authors whose names are listed above declare that they have no conflict of interest.
Discovery of functional alternatively splicedPKMtranscripts in human cancers
Authors: Li Xiangyu , Zhang Cheng , Kim Woonghee , Arif Muhammad , Gao Chunxia , Hober Andreas , Kotol David , Strandberg Linnéa , Forsström Björn , Sivertsson Åsa , Oksvold Per , Grøtli Morten , Sato Yusuke , Kume Haruki , Ogawa Seishi , Boren Jan , Nielsen Jens , Uhlen Mathias , Mardinoglu Adil
Published: April 2019

The association of pyruvate kinase muscle type (PKM) with survival of cancer patients is controversial. Here, we focus on different transcripts ofPKMand investigate the association between their mRNA expression and the clinical survival of the patients in 25 different cancers. We find that the transcript encoding PKM2, and three other functional transcripts are prognostic in multiple cancers. Our integrative analysis shows that the functions of these four transcripts are highly conservative in different cancers. Next, we validate the prognostic effect of these transcripts in an independent kidney renal clear-cell carcinoma (KIRC) cohort and identify a prognostic signature which could distinguish high- and low-risk KIRC patients. Finally, we reveal the functional role of alternatively splicedPKMtranscripts in KIRC, and discover the protein products of different transcripts ofPKM. Our analysis demonstrated that alternatively spliced transcripts of not onlyPKMbut also other genes should be considered in cancer studies, since it may enable the discovery and targeting of the right protein product for development of the efficient treatment strategies.
PopTargs: A database for studying population evolutionary genetics of human microRNA target sites
Authors: Hatlen Andrea , Helmy Mohab , Marco Antonio
Published: April 2019

There is an increasing interest in the study of polymorphic variants at gene regulatory motifs, including microRNA target sites. Understanding the effects of selective forces at specific microRNA target sites, together with other factors like expression levels or evolutionary conservation, requires the joint study of multiple datasets. We have compiled information from multiple sources and compare it with predicted microRNA target sites to built a comprehensive database for the study of microRNA targets in human populations. PopTargs is a web-based tool that allows the easy extraction of multiple datasets and the joint analyses of them, including allele frequencies, ancestral status, population differentiation statistics and site conservation. The user can also compare the allele frequency spectrum between two groups of target sites, and conveniently produce plots. The database can be easily expanded as new data becomes available and the raw database as well as code for creating new custom made databases are available for downloading. We also describe a few illustrative examples.

Availability and implementation

Poptargs is available athttp://poptargs.essex.ac.uk

Contact

amarco.bio@gmail.com
The impact of population variation in the analysis of microRNA target sites
Authors: Helmy Mohab , Hatlen Andrea , Marco Antonio
Published: April 2019

The impact of population variation in the analysis of regulatory interactions is an underdeveloped area. MicroRNA target recognition occurs via pairwise complementarity. Consequently, a number of computational prediction tools have been developed to identify potential target sites, that can be further validated experimentally. However, as microRNA target predictions are done mostly considering a reference genome sequence, target sites showing variation among populations are neglected. Here we study variation at microRNA target sites in human populations and quantify their impact in microRNA target prediction. We found that African populations carry a significant number of potential microRNA target sites that are not detectable in the current human reference genome sequence. Some of these targets are conserved in primates and only lost in Out-of-Africa populations. Indeed, we identified experimentally validated microRNA/transcript interactions that are not detected in standard microRNA target prediction programs, yet they have segregating target alleles abundant in non-European populations. In conclusion, here we show that ignoring population diversity may leave out regulatory elements essential to understand disease and gene expression, particularly neglecting populations of African origin.
Experience-dependent coding of frequency-modulated trajectories by offsets in auditory cortex
Authors: Chong Kelly K , Dunlap Alex G , Kacsoh Dorottya B , Liu Robert C
Published: April 2019

Frequency modulations are an inherent feature of many behaviorally relevant sounds, including vocalizations and music. Changing trajectories in a sound’s frequency often conveys meaningful information, which can be used to differentiate sound categories, as in the case of intonations in tonal languages. However, it is not clear what features of the neural responses in what parts of the auditory cortical pathway might be more important for conveying information about behaviorally relevant frequency modulations, and how these responses change with experience. Here we uncover tuning to subtle variations in frequency trajectories in mouse auditory cortex. Surprisingly, we found that auditory cortical responses could be modulated by variations in a pure tone trajectory as small as 1/24th of an octave. Offset spiking accounted for a significant portion of tuned responses to subtle frequency modulation. Offset responses that were present in the adult A2, but not those in Core auditory cortex, were plastic in a way that enhanced the representation of an acquired behaviorally relevant sound category, which we illustrate with the maternal mouse paradigm for natural communication sound learning. By using this ethologically inspired sound-feature tuning paradigm to drive auditory responses in higher-order neurons, our results demonstrate that auditory cortex can track much finer frequency modulations than previously appreciated, which allows A2 offset responses in particular to attune to the pitch trajectories that distinguish behaviorally relevant, natural sound categories.
GWAS of brain volume on 54,407 individuals and cross-trait analysis with intelligence identifies shared genomic loci and genes
Authors: Jansen Philip R , Nagel Mats , Watanabe Kyoko , Wei Yongbin , Savage Jeanne E , de Leeuw Christiaan A , van den Heuvel Martijn P , van der Sluis Sophie , Posthuma Danielle
Published: April 2019

The phenotypic correlation between human intelligence and brain volume (BV) is considerable (r≈0.40), and has been shown to be due to shared genetic factors1. To further examine specific genetic factors driving this correlation, we present genomic analyses of the genetic overlap between intelligence and BV using genome-wide association study (GWAS) results. First, we conducted the largest BV GWAS meta-analysis to date (N=54,407 individuals), followed by functional annotation and gene-mapping. We identified 35 genomic loci (27 novel), implicating 362 genes (346 novel) and 23 biological pathways for BV. Second, we used an existing GWAS for intelligence (N=269,867 individuals2), and estimated the genetic correlation (rg) between BV and intelligence to be 0.23. We show that the rg is driven by physical overlap of GWAS hits in 5 genomic loci. We identified 67 shared genes between BV and intelligence, which are mainly involved in important signaling pathways regulating cell growth. Out of these 67 we prioritized 32 that are most likely to have functional impact. These results provide new information on the genetics of BV and provide biological insight into BV’s shared genetic etiology with intelligence.
Somatic Genetic Aberrations in Benign Breast Disease and the Risk of Subsequent Breast Cancer
Authors: Zeng Zexian , Vo Andy , Li Xiaoyu , Shidfar Ali , Saldana Paulette , Xuei Xiaoling , Luo Yuan , Khan Seema A. , Clare Susan E.
Published: April 2019

It is largely unknown how the risk of development of breast cancer is transduced by somatic genetic alterations. To address this lacuna of knowledge and acknowledging that benign breast disease (BBD) is an established risk factor for breast cancer, we established a case-control study: The Benign Breast & Cancer Risk (BBCAR) Study. Cases are women with BBD who developed subsequent invasive breast cancer (IBC) at least 3 years after the biopsy and controls are women with BBD who did not develop IBC (median follow-up 16.6 years). We selected 135 cases and individually matched controls (1:2) to cases based on age and type of benign disease: non-proliferative or proliferation without atypia. Whole exome sequencing was performed on DNA from the benign lesions and from subsets with available germline DNA or tumor DNA. Although the number of cases and controls with copy number variation data is limited, several amplifications and deletions are exclusive to the cases. In addition to two known mutational signatures, a novel signature was identified that is significantly (p=0.007) associated with triple negative breast cancer. The somatic mutation rate in benign lesions is similar to that of invasive breast cancer and does not differ between cases and controls. Two mutated genes are significantly associated with time to the diagnosis of breast cancer, and mutations shared between the benign biopsy tissue and the breast malignancy for the ten cases for which we had matched pairs were identified. BBD tissue is a rich source of clues to breast oncogenesis.

One Sentence Summary

Genetic aberrations in benign breast lesions distinguish breast cancer cases from controls and predict cancer risk.
Exercise enhances motor skill learning by neurotransmitter switching in the adult midbrain
Authors: Li Hui-quan , Spitzer Nicholas C.
Published: April 2019

Physical exercise promotes motor skill learning in normal individuals and those with neurological disorders but its mechanism of action is unclear. We found that one week of voluntary wheel running enhances the acquisition of motor skills in adult mice. One week of running also induces switching from ACh to GABA expression in neurons in the caudal pedunculopontine nucleus (cPPN). The switching neurons make projections to the substantia nigra (SN), ventral tegmental area (VTA) and ventrolateral-ventromedial nuclei of the thalamus (VL-VM), which regulate acquisition of motor skills. Use of viral vectors to override transmitter switching blocks the beneficial effect of running on motor skill learning. We suggest that neurotransmitter switching provides the basis by which sustained running benefits motor skill learning, presenting a new target for clinical treatment of movement disorders.
The user’s guide to comparative genomics with EnteroBase, including case studies on transmissions of micro-clades ofSalmonella, the phylogeny of ancient and modernYersinia pestisgenomes, and the core genomic diversity of allEscherichia
Authors: Zhou Zhemin , Alikhan Nabil-Fareed , Mohamed Khaled , Fan Yulei ,   the Agama Study Group, Achtman Mark
Published: April 2019

EnteroBase is an integrated software environment which supports the identification of global population structures within several bacterial genera that include pathogens. Here we provide an overview on how EnteroBase works, what it can do, and its future prospects. EnteroBase has currently assembled more than 300,000 genomes from Illumina short reads fromSalmonella, Escherichia, Yersinia, Clostridiodes, Helicobacter, Vibrio, andMoraxella, and genotyped those assemblies by core genome Multilocus Sequence Typing (cgMLST). Hierarchical clustering of cgMLST sequence types allows mapping, a new bacterial strain to predefined population structures at multiple levels of resolution within a few hours after uploading its short reads. Case study 1 illustrates this process for local transmissions ofSalmonella entericaserovar Agama between neighboring social groups of badgers and humans. EnteroBase also supports SNP calls from both genomic assemblies and after extraction from metagenomic sequences, as illustrated by case study 2 which summarizes the microevolution ofYersinia pestisover the last 5,000 years of pandemic plague. EnteroBase can also provide a global overview of the genomic diversity within an entire genus, as illustrated by case study 3 which presents a novel, global overview of the population structure of all of the species, subspecies and clades withinEscherichia.
A High Dose Tango Intervention for People with Parkinson’s disease (PwPD)
Authors: Rabinovich Débora B. , Garreto Nélida , Arakaki Tomoko , DeSouza Joseph FX
Published: April 2019

Background

Dance has been used extensively to help supplement ongoing therapies for people with PD, most commonly on a weekly or biweekly basis. A daily dose, however, may provide additional benefits. This study examines the dose effect of a dance intervention delivered within a clinic for movement disorders in which PwPD are paired with experienced studio tango dancers.

Objective

The current study aims to examine the dose effects of daily dance for PwPD on motor and non-motor functions directly within a movement disorders clinic.

Design

within-subject, pre-post-intervention, mixed-methods evaluation including UPDRS III.

Setting

The intervention was held at the Movement Disorders Department of a General Hospital in Buenos Aires, Argentina over two-weeks.

Subjects

The class had 21 people in total attendance per class. Two were expert tango dancers and instructors, nine were advanced tango dancers (volunteers), two were caregivers and eight were people with mild and moderate severity [Hoehn and Yahr (H&Y) scale 1-2] idiopathic Parkinson’s disease.

Intervention

Ten dance lessons, each 90-minutes daily within a two-week period.

Outcome measures

The Movement Disorders Society Unified Parkinson’s Disease Rating Scale (UPDRS) part III was used for pre- and post-motor assessment. Psychological questionnaires, a Likert scale examining symptoms, and a pictorial scale were used to assess non-motor aspects. Semi-structured interviews were conducted to assess the impact of the dance intervention on participants’ experience.

Results

Our study found a significant 18% amelioration in motor symptoms as measured by UPDRS III. We also found improvements on activities of daily living (ADL), sleep, Psychological Needs variables - post high dose dance intervention in Likert Scale.

Conclusions

A high dose short-term tango intervention for PwPD improved motor and non-motor aspects of PD such as ADL and sleep with high levels of adherence (97.5%) and enjoyment reported by participants. This dance intervention also improved participant’s perception of their own skills. The frequency or dosage of dance in rehabilitation suggests that an increased dose from once per week to 5 times per week can ameliorate many symptoms of PD and could be used as a short-term intervention.
A glycosaminoglycan extract from Portunus pelagicus inhibits BACE1, the β secretase implicated in Alzheimer’s disease
Authors: Mycroft-West Courtney J. , Cooper Lynsay C. , Devlin Anthony J. , Procter Patricia , Guimond Scott Eric , Guerrini Marco , Fernig David G. , Lima Marcelo A. , Yates Edwin A. , Skidmore Mark A.
Published: April 2019

Therapeutic options for Alzheimer’s disease, the most common form of dementia, are currently restricted to palliative treatments. The glycosaminoglycan heparin, widely used as a clinical anticoagulant, has previously been shown to inhibit the Alzheimer’s disease-relevant β-secretase 1 (BACE1). Despite this, the deployment of pharmaceutical heparin for the treatment of Alzheimer’s disease is largely precluded by its potent anticoagulant activity. Furthermore, ongoing concerns regarding the use of mammalian sourced heparins, primarily due to prion diseases and religious beliefs, hinder the deployment of alternative heparin based therapeutics. A marine-derived, heparan sulphate-containing glycosaminoglycan extract isolated from the crabPortunus pelagicus, was identified to inhibit human BACE1 with comparable bioactivity to that of mammalian heparin (IC50= 1.85 μg.mL-1(R2= 0.94) and 2.43 μg.mL-1(R2= 0.93), respectively) possessing highly attenuated anticoagulant activities. The results from several structural techniques suggest that the interactions between BACE1 and the extract fromP. pelagicusare complex and distinct from those of heparin.
Raven Calls Indicate Sender’s Neural State
Authors: Huang Zixuan , Wang Zhilong , Xie Jun , Mirt Greg , Yan Chengying , Zhong Jing , Deng Xianli , Liu Fangfang , Zhou Chunlin , Xu Fan
Published: April 2019

Vocal communication accounts for dominantly percentage within animal species. The information of vocal samples contains not only the amplitude of objects, but also the emotional states behind it. However, to extract the emotion state behind the sound remains controversial. Here we introduce an artificial network method, the Back Propagation Neural Network, BPNN, to classify the emotional states behind the sound. The results disclosed the behaviour categories, including alarm, flight, begging and singing which has been successfully classified. This artificial intelligence classification may aid us to distinguish the ecological categories via animal vocal communication and to discover its significance of evolution and nature.
Pan-genomic and Polymorphic Driven Prediction of Antibiotic Resistance inElizabethkingia
Authors: Naidenov Bryan , Willyerd Karyn , Lim Alexander , Torres Nathanial J , Johnson William L. , Hwang Hong Jin , Hoyt Peter , Gustafson John , Chen Charles
Published: April 2019

TheElizabethkingiaare a genetically diverse genus of emerging pathogens that exhibit multidrug resistance to a range of common antibiotics. Two representative species,Elizabethkingia bruunianaandElizabethkingia meningoseptica, were phenotypically tested to determine minimum inhibitory concentrations for five antibiotics. Ultra-long read sequencing with Oxford Nanopore Technologies and subsequentde novoassembly produced complete, gapless circular genomes for each strain. Alignment based annotation with Prokka identified 5,480 features inE. bruunianaand 5,203 features inE. meningoseptica, where none of these identified genes or gene combinations corresponded to observed phenotypic resistance values. Pan-genomic analysis, performed with an additional 19Elizabethkingiastrains, identified a core-genome size of 2,658,537 bp, 32 uniquely identifiable intrinsic chromosomal antibiotic resistance core-genes and 77 antibiotic resistance pan-genes. Using core-SNPs and pan-genes in combination with six machine learning algorithms, binary classification of clindamycin and vancomycin resistance achieved f1 scores of 0.94 and 0.84 respectively. Performance on the more challenging multiclass problem for fusidic acid, rifampin and ciprofloxacin resulted in f1 scores of 0.70, 0.75 and 0.54 respectively.
CXCL5-mediated recruitment of neutrophils into the peritoneal cavity ofGdf15-deficient mice protects against abdominal sepsis
Authors: Santos Isa , Colaço Henrique G. , Neves-Costa Ana , Seixas Elsa , Velho Tiago R. , Pedroso Dora , Barros André , Martins Rui , Carvalho Nuno , Payen Didier , Weis Sebastian , Yi Hyon-Seung , Shong Minho , Moita Luís Ferreira
Published: April 2019

Sepsis is a life-threatening organ dysfunction condition caused by a dysregulated host response to an infection. Here we report that the circulating levels of growth-differentiation factor-15 (GDF15) are strongly increased in septic shock patients and correlate with mortality. In mice, we find that peptidoglycan is a potent ligand that signals through the TLR2-Myd88 axis for the secretion of GDF15 and thatGdf15-deficient animals are protected against abdominal sepsis due to increased chemokine CXC ligand 5 (CXCL5)-mediated recruitment of neutrophils into the peritoneum leading to better local bacterial control. Our results identify GDF15 as a potential target to improve sepsis treatment. Its inhibition should increase neutrophil recruitment to the site of infection and consequently lead to better pathogen control and clearance.
Two-Color Spatial Cumulant Analysis Detects Heteromeric Interactions between Membrane Proteins
Authors: Foust D.J. , Godin A.G. , Ustione A. , Wiseman P.W. , Piston D.W.
Published: April 2019

Fluorescence fluctuation spectroscopy (FFS) can be used to measure the aggregation of fluorescently labeled molecules and is typically performed using time series data. Spatial intensity distribution analysis (SpIDA) and fluorescence moment image analysis (FMIA) are established tools for measuring molecular brightnesses from single-color images collected with laser scanning microscopes. We have extended these tools for analysis of two-color images to resolve heteromeric interactions between molecules labeled with spectrally distinct chromophores. We call these new methods two-color SpIDA (2c-SpIDA) and two-color spatial cumulant analysis (2c-SpCA). To implement these techniques on a hyperspectral imaging system, we developed a spectral shift filtering (SSF) technique to remove artifacts due to intrinsic crosstalk between detector bins. We determined that 2c-SpCA provides better resolution from samples containing multiple fluorescent species, hence this technique was carried forward to study images of living cells. We used fluorescent heterodimers labeled with EGFP and mApple to quantify the effects of resonance energy transfer and incomplete maturation of mApple on brightness measurements. We show that 2c-SpCA can detect the interaction between two components of trimeric G-protein complexes. Thus 2c-SpCA presents a robust and computationally expedient means of measuring heteromeric interactions in cellular environments.

Statement of Significance

Fluorescence fluctuation spectroscopy (FFS) techniques determine biophysical parameters from samples containing fluorescently labeled biomolecules by considering the statistical nature of fluorescent signals measured with photodetectors. The present study introduces two-color spatial cumulant analysis (2c-SpCA) to the canon of FFS techniques. 2c-SpCA analyzes pixel-value data of two-color images collected with laser scanning fluorescence microscopes. We show that 2c-SpCA can determine several biophysical parameters in living cells including Forster resonance energy transfer efficiency, the dark state fraction of fluorescent proteins, and heteromerization between distinctly labeled proteins. In comparison to existing techniques, 2c-SpCA requires very few image frames for analysis, minimal computations, and can be applied to images of fixed tissue samples.
Response of Bat Activity to Land-Cover and Land-Use Change in Savannas is Scale-, Season-, and Guild-Specific
Authors: Shapiro Julie Teresa , Monadjem Ara , Röder Timo , McCleery Robert A.
Published: April 2019

Tropical savannas are biomes of global importance that are under severe pressure from anthropogenic change, including land-cover and land-use change. Bats, the second-most diverse group of mammals, are critical to ecosystem functioning, but may be vulnerable to such anthropogenic stresses. However, there is little information on the response of savanna bats to land-cover and land-use change, especially in Africa. This limits our ability to develop conservation strategies for bats and maintain the ecosystem functions and services they provide in this biome. Using acoustic monitoring, we measured how guild-specific (aerial, edge, and clutter forager) bat activity responded to both fine-scale metrics of vegetation structure and landscape-scale metrics of land-cover composition and configuration across the wet and dry seasons in a savanna in southern Africa undergoing rapid land-cover and land-use change. We found that all three guilds responded more strongly to landscape metrics than fine-scale vegetation structure, although the specific metrics varied between guilds. Aerial and edge bats responded most strongly to the percent savanna cover and savanna fragmentation in both seasons while clutter bats responded to percent rural cover in the wet season and percent water cover in the dry. All three guilds responded more strongly to the landscape in the dry season than the wet season. Our results show it is possible to conserve bats, and the ecosystem services they can provide, in savannas undergoing anthropogenic land-use and land-cover change but strategies to do so must consider foraging guild, large spatial scales, and seasonal variation in bat activity.

Highlights

Bats in savannas respond to land-cover and land-use change on large spatial scales

Landscape had a greater influence on bat activity in the dry season than the wet

Aerial and edge forager activity responded to savanna cover and fragmentation

Clutter forager activity was best explained by rural and water cover

Minimizing fragmentation and maintaining water promotes bat activity in modified savannas
Rapid 3D enhanced resolution microscopy reveals the dynamics of cortical dendritic spinules
Authors: Zaccard CR , Myczek K , Martin-de-Saavedra MD , Penzes P
Published: April 2019

Dendritic spinules are thin, membranous protrusions formed by neuronal dendritic spines that are not adequately resolved by diffraction-limited light microscopy. Hence, our understanding of spinules is inferred predominantly from fixed-tissue electron microscopy (EM). Super-resolution modalities have enabled live-cell nanoscopic imaging, but their utility for fast, time-lapse, volumetric imaging has been restricted. Herein, we utilized rapid structured illumination microscopy (SIM) and ‘enhanced resolution’ confocal microscopy to study spatiotemporal spinule dynamics in live cultured cortical pyramidal neurons. Spinules on mushroom spines typically recurred at the same topographical locations and most were short-lived, originating near simple post-synaptic densities (PSDs), while a subset was long-lived and elongated, emerging from complex PSDs. Ca2+ puncta within spinules synchronized with spine head transients and Ca2+ depletion drastically decreased spinule number. Moreover, we uncovered evidence of differential Ca2+-mediated regulation of short-lived and long-lived spinules. Thus, we identified unique spinule classes divergent in lifespan, dynamics, morphology, relationship to the PSD, and regulation. These data suggest distinct synaptic functions of spinule classes, informing future studies, while demonstrating a new application for enhanced resolution microscopy.
Optimizing oleaginous yeast cell factories for flavonoids and hydroxylated flavonoids biosynthesis
Authors: Lv Yongkun , Koffas Mattheos , Zhou Jingwen , Xu Peng
Published: April 2019

Plants possess myriads of secondary metabolites with a broad spectrum of health-promoting benefits. Up to date, plant extraction is still the primary route to produce high-value natural products, which inherently suffers from economics and scalability issues. Heterologous production in microbial host is considered as a feasible approach to overcoming these limitations. Flavonoid and its hydroxylated derivatives represent a diversified family of bioactive compounds, most prominently known as antioxidant and anti-aging agents. Oleaginous yeast is rich in hydrophobic lipid bodies and spatially-organized organelles, which provides the ideal environment for the regioselectivity and stereoselectivity of many plant-specific enzymes. In this report, we validated thatY. lipolyticais a superior platform for heterologous production of high-value flavonoids and hydroxylated flavonoids. By modular construction and characterization, we determined the rate-limiting steps for efficient flavonoids biosynthesis inY. lipolytica. We evaluated various precursor pathways and unleashed the metabolic potential ofY. lipolyticato produce flavonoids, including the supply of acetyl-CoA, malonyl-CoA and chorismate. Coupled with the optimized chalcone synthase module and the hydroxylation module, our engineered strain produced 252.4 mg/L naringenin, 134.2 mg/L eriodictyol and 110.5 mg/L taxifolin from glucose. Collectively, these findings demonstrate our ability to harness oleaginous yeast as microbial workhorse to expand nature’s biosynthetic potential, enabling us to bridge the gap between drug discovery and natural product manufacturing.
A Rab GTPase protein FvSec4 is necessary for fumonisin B1 biosynthesis and virulence inFusarium verticillioides
Authors: Yan Huijuan , Huang Jun , Zhang Huan , Shim Won Bo
Published: April 2019

Rab GTPases are responsible for a variety of membrane trafficking and vesicular transportation in fungi. But the role of Rab GTPases inFusarium verticillioides, one of the key corn pathogens worldwide, remains elusive. These Small GTPases in fungi, particularly those homologous toSaccharomyces cerevisiaeSec4, are known to be associated with protein secretion, vesicular trafficking, secondary metabolism and pathogenicity. Here, we characterized the molecular functions of FvSec4 by generating a null mutant and learned that it is important for vegetative growth, hyphal branching, and conidiation. Interestingly, the mutation did not impair the expression of key conidiation-related genes. Meanwhile, the mutant did not show any defect in sexual development, including perithecia production. GFP-FvSec4 localized to growing hyphal tips, and raised the possibility that FvSec4 is involved in protein trafficking and endocytosis. The mutant exhibited defect in corn stalk rot virulence and also significant alteration of fumonisn B1 production. The mutation led to more sensitivity to oxidative and cell wall stress agents, and defects in carbon utilization. Gene complementation fully restored the defects in the mutant demonstrating that FvSec4 plays important role in these functions. Taken together, our data indicate that FvSec4 plays important roles inF. verticillioideshyphal development, virulence, mycotoxin production and stresses response. Further study is needed to characterize whether the mutation in FvSec4 leads to altered vesicle trafficking and protein secretion, which ultimately impactF. verticillioidesphysiology and virulence.
Molecular signatures of non-typeable Haemophilus influenzae lung adaptation in paediatric chronic lung disease
Authors: Aziz Ammar , Sarovich Derek S. , Nosworthy Elizabeth , Beissbarth Jemima , Chang Anne B. , Smith-Vaughan Heidi , Price Erin P. , Harris Tegan M.
Published: April 2019

Non-typeable Haemophilus influenzae (NTHi), an opportunistic pathogen of the upper airways of healthy children, can infect the lower airways, driving chronic lung disease. However, the molecular basis underpinning NTHi transition from a commensal to a pathogen is not clearly understood. Here, we performed comparative genomic and transcriptomic analyses of 12 paired, isogenic NTHi strains, isolated from the nasopharynx (NP) and bronchoalveolar lavage (BAL) of 11 children with chronic lung disease, to identify convergent molecular signatures associated with lung adaptation. Comparative genomic analyses of the 12 NP-BAL pairs demonstrated that five were genetically identical, with the remaining seven differing by only 1 to 3 mutations. Within-patient transcriptomic analyses identified between 2 and 58 differentially expressed genes in 8 of the 12 NP-BAL pairs, including pairs with no observable genomic changes. Whilst no convergence was observed at the gene level, functional enrichment analysis revealed significant under-representation of differentially expressed genes belonging to Coenzyme metabolism, Function unknown, Translation, ribosomal structure and biogenesis Cluster of Orthologous Groups categories. In contrast, Carbohydrate transport and metabolism, Cell motility and secretion, Intracellular trafficking and secretion, and Energy production categories were over-represented. This observed trend amongst genetically-unrelated NTHi strains provides evidence of convergent transcriptional adaptation of NTHi to paediatric airways that deserves further exploration. Understanding the pathoadaptative mechanisms that NTHi employs to infect and persist in the lower paediatric airways is essential for devising targeted diagnostics and treatments aimed at minimising disease severity, and ultimately, preventing NTHi lung infections and subsequent chronic lung disease in children.
Hallmarks of slow translation initiation revealed in mitochondrially localizing mRNA sequences
Authors: Poulsen Thomas M. , Imai Kenichiro , Frith Martin C. , Horton Paul
Published: April 2019

The mRNA of some, but not all, nuclear encoded mitochondrial proteins localize to the periphery of mitochondria. Previous studies have shown that both the nascent polypeptide chain and an mRNA binding protein play a role in this phenomenon, and have noted a positive correlation between mRNA length and mitochondrial localization. Here, we report the first investigation into the relationship between mRNA translation initiation rate and mRNA mitochondrial localization. Our results indicate that translation initiation promoting factors such as Kozak sequences are associated with cytosolic localization, while inhibiting factors such as 5′ UTR secondary structure correlate with mitochondrial localization. Moreover, the frequencies of nucleotides in various positions of the 5′ UTR show higher correlation with localization than the 3′ UTR. These results indicate that mitochondrial localization is associated with slow translation initiation. Interestingly this may help explain why short mRNAs, which are thought to initiate translation rapidly, seldom localize to mitochondria. We propose a model in which translating mRNA has reduced mobility and tends not to reach mitochondria. Finally, we explore this model with a simulation of mRNA diffusion using previously estimated translation initiation probabilities, confirming that our model can produce localization values similar to those measured in experimental studies.
Role of human induced pluripotent stem cell-derived spinal cord astrocytes in the functional maturation of motor neurons in a multielectrode array system
Authors: Taga Arens , Dastgheyb Raha , Habela Christa , Joseph Jessica , Richard Jean-Philippe , Gross Sarah K. , Lauria Giuseppe , Lee Gabsang , Haughey Norman , Maragakis Nicholas J.
Published: April 2019

The ability to generate human induced pluripotent stem cell (hiPSC)-derived neural cells displaying region-specific phenotypes is of particular interest for modeling central nervous system (CNS) biology in vitro. We describe a unique method by which spinal cord hiPSC-derived astrocytes (hiPSC-A) are cultured with spinal cord hiPSC-derived motor neurons (hiPSC-MN) in a multielectrode array (MEA) system to record electrophysiological activity over time. We show that hiPSC-A enhance hiPSC-MN electrophysiological maturation in a time-dependent fashion. The sequence of plating, density, and age in which hiPSC-As are co-cultured with MN, but not their respective hiPSC line origin, are factors that influence neuronal electrophysiology. When compared to co-culture with mouse primary spinal cord astrocytes, we observe an earlier and more robust electrophysiological maturation in the fully human cultures, suggesting that the human origin is relevant to the recapitulation of astrocyte/motor neuron cross-talk. Finally, we test pharmacological compounds on our MEA platform and observe changes in electrophysiological activity which confirm hiPSC-MN maturation. These findings are supported by immunocytochemistry and real time PCR studies in parallel cultures demonstrating human astrocyte mediated changes in the structural maturation and protein expression profiles of the neurons. Interestingly, this relationship is reciprocal and co-culture with neurons influences astrocyte maturation as well. Taken together these data indicate that in a human in vitro spinal cord culture system, astrocytes alter hiPSC-MN maturation in a time-dependent and species specific manner and suggest a closer approximation of in vivo conditions.

Main Points

We developed a method for the co-culture of human iPSC-A/MN for multielectrode array recordings.

The morphological, molecular, pharmacological, and electrophysiological characterization of the co-cultures suggests bidirectional maturation.
Modeling the language of life – Deep Learning Protein Sequences
Authors: Heinzinger Michael , Elnaggar Ahmed , Wang Yu , Dallago Christian , Nechaev Dmitrii , Matthes Florian , Rost Burkhard
Published: April 2019

Background

One common task in Computational Biology is the prediction of aspects of protein function and structure from their amino acid sequence. For 26 years, most state-of-the-art approaches toward this end have been marrying machine learning and evolutionary information. The retrieval of related proteins from ever growing sequence databases is becoming so time-consuming that the analysis of entire proteomes becomes challenging. On top, evolutionary information is less powerful for small families, e.g. for proteins from theDark Proteome.

Results

We introduce a novel way to represent protein sequences as continuous vectors (embeddings) by using the deep bi-directional model ELMo taken from natural language processing (NLP). The model has effectively captured the biophysical properties of protein sequences from unlabeled big data (UniRef50). After training, this knowledge is transferred to single protein sequences by predicting relevant sequence features. We refer to these new embeddings asSeqVec(Sequence-to-Vector) and demonstrate their effectiveness by training simple convolutional neural networks on existing data sets for two completely different prediction tasks. At the per-residue level, we significantly improved secondary structure (for NetSurfP-2.0 data set: Q3=79%±1, Q8=68%±1) and disorder predictions (MCC=0.59±0.03) over methods not using evolutionary information. At the per-protein level, we predicted subcellular localization in ten classes (for DeepLoc data set: Q10=68%±1) and distinguished membrane-bound from water-soluble proteins (Q2= 87%±1). All results built upon the embeddings gained from the new toolSeqVecneither explicitly nor implicitly using evolutionary information. Nevertheless, it improved over some methods using such information. Where the lightning-fastHHblitsneeded on average about two minutes to generate the evolutionary information for a target protein,SeqVeccreated the vector representation on average in 0.03 seconds.

Conclusion

We have shown that transfer learning can be used to capture biochemical or biophysical properties of protein sequences from large unlabeled sequence databases. The effectiveness of the proposed approach was showcased for different prediction tasks using only single protein sequences.SeqVecembeddings enable predictions that outperform even some methods using evolutionary information. Thus, they prove to condense the underlying principles of protein sequences. This might be the first step towards competitive predictions based only on single protein sequences.

Availability

SeqVec:https://github.com/mheinzinger/SeqVecPrediction server:https://embed.protein.properties
Crosslinking actin networks produces compressive force
Authors: Ma Rui , Berro Julien
Published: April 2019

Actin has been shown to be essential for clathrin-mediated endocytosis in yeast. However, actin polymerization alone is likely insufficient to produce enough force to deform the membrane against the huge turgor pressure of yeast cells. In this paper, we used Brownian dynamics simulations to demonstrate that crosslinking of a meshwork of non-polymerizing actin filaments is able to produce compressive forces. We show that the force can be up to thousands of piconewtons if the crosslinker has a high stiffness. The force decays over time as a result of crosslinker turnover, and is a result of converting chemical binding energy into elastic energy.
Ultraviolet light degrades the mechanical and structural properties of human stratum corneum
Authors: Lipsky Z.W. , German G. K.
Published: April 2019

Prolonged exposure of human skin to sunlight causes photodamage, which induces the early onset of wrinkles and increased tissue fragility. While solar ultraviolet (UV) light is considered to have the most damaging effect, the UV range that is most harmful remains a topic of significant debate. In this study, we take a first step towards elucidating biomechanical photoageing effects by quantifying how exposure to different UV ranges and dosages impacts the mechanical and structural properties of human stratum corneum (SC), the most superficial skin layer. Mechanical testing reveals that irradiation of isolated human SC to UVA (365 nm), UVB (302 nm), or UVC (265 nm) light with dosages of up to 4000 J/cm2notably alters the elastic modulus, fracture stress, fracture strain, and work of fracture. For equivalent incident dosages, UVC degrades SC the greatest. However, upon discounting reflected and transmitted components of the incident light, a generalized scaling law relating the photonic energy absorbed by the SC to the energy cost of tissue fracture emerges. This relationship indicates that no one UV range is more damaging than another. Rather, the magnitude of absorbed UV energy governs the degradation of tissue mechanical integrity. Subsequent structural studies are performed to elucidate the cause of this mechanical degradation. UV absorption scales with the spatial dispersion of desmoglein 1 (Dsg 1), a component of corneocyte cell-cell junctions, away from intercellular sites. Combining both scaling laws, we establish a mechanical-structural model capable of predicting UV induced tissue mechanical integrity from Dsg 1 dispersion.

Statement of Significance

Photoageing from the sun can produce early onset of skin wrinkles and an increase in tissue fragility that heightens the risk of rupture. While solar ultraviolet (UV) light is considered to have the most damaging effect, the UV range that is most harmful remains a topic of significant debate. In this study, we elucidate photoageing effects by quantifying how exposure to different UV ranges and dosages impacts the mechanical and structural properties of human stratum corneum (SC), the most superficial skin layer. Results establish a mechanical-structural model that relates the amount of UV energy absorbed by the tissue, irrespective of UV range, to the energy cost of tissue fracture and spatial dispersion of desmoglein 1.
Time to monitor livestock carcasses for biodiversity conservation and public health
Authors: Mateo‐Tomás Patricia , Olea Pedro P. , López‐Bao José Vicente
Published: April 2019

Law enforcement and integration of environmental issues into other policies able to affect species and ecosystems are cornerstones for the effective protection of biodiversity. We illustrate the necessity of monitoring and improving such enforcement and integration through the example of the European sanitary policies for managing livestock carcasses after the “mad cow disease” outbreak while supporting scavengers’ conservation.

Continuous updates of EU sanitary regulations for reconciling scavenger conservation and public health have led to actions whose implementation and monitoring rely on delineating scavenger feeding zones and estimating livestock mortality and scavenger feeding requirements (SFRs).

However, the lack of clear and homogeneous criteria to calculate SFRs results in remarkable variations (e.g. of up to >450% in some regions). Although we offer a prospective approach to improve SFR calculations, we show here the high levels of uncertainty in these estimates based on the imperfect information available regarding scavengers’ ecology.

Synthesis and applications. The multiple socioeconomic and ecological impacts of sanitary crises such as “mad cow disease” underpin the need for high‐level political commitments to guarantee public health, food security and biodiversity conservation. To effectively integrate scavenger conservation into sanitary policies, the shortfalls here detected when estimating scavenger feeding requirements should be addressed. We recommend policymakers establish systematic on‐ground carcass monitoring for detecting implementation gaps (e.g. food shortages compromising scavenger conservation, potential risks of disease transmission), and to modify EU sanitary regulations accordingly. This monitoring would increase knowledge on scavenging ecology and epidemiological surveillance while enhancing cross‐compliance. Lessons learned in Europe may help to integrate conservation issues into other sectorial policies worldwide.