bluuebunny/crossref_metadata_2025_split huggingface dataset:
A two-color haploid genetic screen identifies novel host factors involved in HIV latency
Authors: Röling Michael D , Mollapour Sisakht Mahsa , Ne Enrico , Moulos Panagiotis , Stoszko Mateusz , De Crignis Elisa , Bodmer Helen , Kan Tsung Wai , Akbarzadeh Maryam , Harokopos Vaggelis , Hatzis Pantelis , Palstra Robert-Jan , Mahmoudi Tokameh 
Published: January 2021

AbstractTo identify novel host factors as putative targets to reverse HIV latency, we performed an insertional mutagenesis genetic screen in a latently HIV-1-infected pseudo-haploid KBM7 cell line (Hap-Lat). Following mutagenesis, insertions were mapped to the genome and bioinformatic analysis resulted in the identification of 69 candidate host genes involved in maintaining HIV-1 latency. A select set of candidate genes was functionally validated using shRNA mediated depletion in latent HIV-1 infected J-Lat A2 and 11.1 T cell lines. We confirmed ADK, CHD9, CMSS1, EVI2B, EXOSC8, FAM19A, GRIK5, IRF2BP2, NF1, and USP15 as novel host factors involved in the maintenance of HIV latency. Chromatin immunoprecipitation assays indicated that CHD9, a Chromodomain Helicase DNA-binding protein, maintains HIV latency via direct association with the HIV 5’LTR, and its depletion results in increased histone acetylation at the HIV-1 promoter, concomitant with HIV-1 latency reversal. FDA-approved inhibitors 5-Iodotubercidin, Trametinib, and Topiramate, targeting ADK, NF1, and GRIK5, respectively were characterized for their latency reversal potential. While 5-Iodotubercidin exhibited significant cytotoxicity in both J-Lat and primary CD4+ T cells, Trametinib reversed latency in J-Lat cells but not in latently HIV-1-infected primary CD4+ T cells. Crucially, Topiramate reversed latency in cell-line models and latently infected primary CD4+ T cells, without inducing T cell activation or significant toxicity. Thus, using an adaptation of a haploid forward genetic screen, we identified novel and druggable host factors contributing to HIV-1 latency.ImportanceA reservoir of latent HIV-1-infected cells persists in the presence of combination antiretroviral therapy (cART), representing a major obstacle for viral eradication. Reactivation of the latent HIV-1 provirus is part of curative strategies which aim to promote clearance of the infected cells. Using a two-color haploid screen, we identified 69 candidate genes as latency maintaining host factors and functionally validated a subset of 10 of those in additional T-cell based cell line models of HIV-1 latency. We further demonstrated that CHD9 is associated with HIV-1’s promoter, the 5’LTR while this association is lost upon reactivation. Additionally, we characterized the latency reversal potential of FDA compounds targeting ADK, NF1, and GRIK5 and identify the GRIK5 inhibitor Topiramate as a viable latency reversal agent with clinical potential.
Filamentation inCandida albicansis modulated by adaptive translation of farnesol signalling genes
Authors: Oliveira Carla , Guimarães Ana Rita , Correia Inês , Sousa Inês , Poim Ana , Rocha Sílvia M. , Moura Gabriela , Santos Manuel A. S. , Bezerra Ana Rita 
Published: January 2021

AbstractThe complex biology of the human pathogenCandida albicansis reflected in its remarkable ability to proliferate in numerous body sites, adapt to drastic changes in the environment, form various types of colonies and grow in yeast, pseudo-hyphal and hyphal forms. Much has been learnt in recent years about the relevance of this phenotypic plasticity, but the mechanisms that support it are still not fully understood. We have demonstrated that atypical translation of the CUG codon is a source of unexpected morphological diversity. The CUG codon is translated as both leucine (Leu) (~3%) and serine (Ser) (~97%) in normal growth conditions, but Ser/Leu levels change in response to stress. Remarkably, recombinantC. albicansstrains incorporating between 20% and 99% of Leu at CUG sites display a diverse array of phenotypes and produce colonies of variable morphology containing a mixture of yeast, pseudohyphal and hyphal cells. In this work we investigate the role of the CUG codon in the yeast-hypha transition. Our data show that increasing incorporation levels of Leu at CUG sites trigger hyphal initiation under non-inducing conditions by reducing farnesol production, and increasing the degradation of the Nrg1 hyphal repressor. We propose that dual CUG Ser/Leu translation triggers filamentation via the Nrg1 pathway.ImportanceThe unique translation of the CUG codon as both Ser (~97%) and Leu (~3%) plays a key role in the production of high genomic and phenotypic diversity inC. albicans. The molecular mechanisms that support such diversity are poorly understood. Here, we show that increased Leu incorporation at CUG sites induce hyphae formation in media whereC. albicansnormally grows in the yeast form. The data show that increasing Leu at CUG sites triggers the degradation of the hyphal repressor Nrg1, allowing for full expression of hyphal genes. Since filamentation is important for invasion of host tissues, this work shows how the atypical translation of a single codon may play a critical role in the virulence of all fungi of the CTG clade.
iGEM comes of age: trends in its research output
Authors: Jainarayanan Ashwin K. , Galanis Anastasios , Nakara Amatullah Mustafa , Kundlatsch Guilherme E. , Rubio-Sánchez Roger 
Published: January 2021

AbstractThe international Genetically Engineered Machine (iGEM) is an educational benchmark in synthetic biology. Eighteen years after its inception, it has also catalysed the infusion of synthetic biology with interdisciplinary fundamental and translational research as well as with inspired young scientists. Here, we communicate a quantitative analysis of compiled published work associated to iGEM projects, highlighting trends in their dissemination and versatility. As iGEM comes of age, we anticipate it will continue to revolutionise, alongside SynBio, several disciplines of science and industries through the development of synthetic biological systems towards a sustainable future.
Independent control of mean and noise by convolution of gene expression distributions
Authors: Gerhardt Karl P. , Rao Satyajit D. , Olson Evan J. , Igoshin Oleg A. , Tabor Jeffrey J. 
Published: January 2021

AbstractGene expression noise can reduce cellular fitness or facilitate processes such as alternative metabolism, antibiotic resistance, and differentiation. Unfortunately, efforts to study the impacts of noise have been hampered by a scaling relationship between noise and expression level from a single promoter. Here, we use theory to demonstrate that mean and noise can be controlled independently by expressing two copies of a gene from separate inducible promoters in the same cell. We engineer low and high noise inducible promoters to validate this result inEscherichia coli, and develop a model that predicts the experimental distributions. Finally, we use our method to reveal that the response of a promoter to a repressor is less sensitive with higher repressor noise and explain this result using a law from probability theory. Our approach can be applied to investigate the effects of noise on diverse biological pathways or program cellular heterogeneity for synthetic biology applications.
Prophage-encoded hotspots of bacterial immune systems
Authors: Rousset François , Dowding Julien , Bernheim Aude , Rocha Eduardo P.C. , Bikard David 
Published: January 2021

AbstractThe arms race between bacteria and phages led to the emergence of a variety of genetic systems used by bacteria to defend against viral infection, some of which were repurposed as powerful biotechnological tools. While numerous defense systems have been identified in genomic regions termed defense islands, it is believed that many more remain to be discovered. Here, we show that P2- like prophages and their P4-like satellites have genomic hotspots that represent a significant source of novel anti-phage systems. We validate the defense activity of 14 systems spanning various protein domains and describe PARIS, an abortive infection system triggered by a phage-encoded anti-restriction protein. Immunity hotspots are present across prophages of distant bacterial species, highlighting their biological importance in the competition between bacteria and phages.
A new method for determining ribosomal DNA copy number shows differences betweenSaccharomyces cerevisiaepopulations
Authors: Sharma Diksha , Hermann-Le Denmat Sylvie , Matzke Nicholas J. , Hannan Katherine , Hannan Ross D. , O’Sullivan Justin M. , Ganley Austen R. D. 
Published: January 2021

AbstractRibosomal DNA genes (rDNA) encode the major ribosomal RNAs (rRNA) and in eukaryotic genomes are typically present as one or more arrays of tandem repeats. Species have characteristic rDNA copy numbers, ranging from tens to thousands of copies, with the number thought to be redundant for rRNA production. However, the tandem rDNA repeats are prone to recombination-mediated changes in copy number, resulting in substantial intra-species copy number variation. There is growing evidence that these copy number differences can have phenotypic consequences. However, we lack a comprehensive understanding of what determines rDNA copy number, how it evolves, and what the consequences are, in part because of difficulties in quantifying copy number. Here, we developed a genomic sequence read approach that estimates rDNA copy number from the modal coverage of the rDNA and whole genome to help overcome limitations in quantifying copy number with existing mean coverage-based approaches. We validated our method using strains of the yeastSaccharomyces cerevisiaewith previously-determined rDNA copy numbers, and then applied our pipeline to investigate rDNA copy number in a global sample of 788 yeast isolates. We found that wild yeast have a mean copy number of 92, consistent with what is reported for other fungi but much lower than in laboratory strains. We also show that different populations have different rDNA copy numbers. These differences can partially be explained by phylogeny, but other factors such as environment are also likely to contribute to population differences in copy number. Our results demonstrate the utility of the modal coverage method, and highlight the high level of rDNA copy number variation within and between populations.Author summaryThe ribosomal RNA gene repeats (rDNA) form large tandem repeat arrays in most eukaryote genomes. Their tandem arrangement makes the rDNA prone to copy number variation, and there is increasing evidence that this copy number variation has phenotypic consequences. However, difficulties in measuring rDNA copy number hamper investigation into rDNA copy number dynamics and their significance. Here we developed a novel bioinformatics method for measuring rDNA copy number from whole genome sequence data that is based on the modal sequence read coverage. We established parameters for optimal performance of the method and validated it using yeast strains of known rDNA copy numbers. We then applied the method to a dataset of almost 800 global yeast isolates and demonstrate that yeast populations have different rDNA copy numbers that partially correlate with phylogeny. Our work provides a simple and accurate method for determining rDNA copy number that leverages the growing number of whole genome datasets, and highlights the dynamic nature of rDNA copy number.
The fitness costs and benefits of trisomy of eachCandida albicanschromosome
Authors: Yang Feng , Jiang Yuan-ying , Cao Yong-bing , Berman Judith 
Published: January 2021

AbstractCandida albicansis a prevalent human fungal pathogen. Rapid genomic change, due to aneuploidy, is a common mechanism that facilitates survival from multiple types of stresses including the few classes of available antifungal drugs. The stress survival of aneuploids occurs despite the fitness costs attributed to most aneuploids growing under idealized lab conditions. Systematic study of the aneuploid state inC. albicanshas been hindered by the lack of a comprehensive collection of aneuploid strains. Here, we describe a collection of diploidC. albicansaneuploid strains, each carrying one extra copy of each chromosome, all from the same genetic background. We tested the fitness of this collection under several physiological conditions including shifts in pH, low glucose, oxidative stress, temperature, high osmolarity, membrane stress and cell wall stress. We found that, most aneuploids, under most conditions, were less fit than their euploid parent, yet there were specific conditions under which specific aneuploid isolates provided a fitness benefit relative to the euploid parent strain. Importantly, this fitness benefit was attributable to the change in the copy number of specific chromosomes. Thus,C. albicanscan tolerate aneuploidy of each chromosome and some aneuploids confer improved growth under conditions that the yeast encounters in its host niches.
Structure of an Hsp90-immunophilin complex reveals cochaperone recognition of the client-maturation state
Authors: Lee Kanghyun , Thwin Aye C. , Tse Eric , Gates Stephanie N. , Southworth Daniel R. 
Published: January 2021

SummaryThe Hsp90 chaperone promotes the folding and activation of hundreds of client proteins in the cell through an ATP-dependent conformational cycle guided by distinct cochaperone regulators. The FKBP51 immunophilin binds Hsp90 with its tetratricopeptide repeat (TPR) domain and catalyzes peptidyl-prolyl isomerase (PPIase) activity during the folding of kinases, nuclear receptors and tau. Here we have determined the cryo-EM structure of the human Hsp90:FKBP51:p23 complex to 3.3 Å that, together with mutagenesis and crosslinking analysis, reveals the basis for cochaperone binding to Hsp90 during client maturation. A helix extension in the TPR functions as a key recognition element, interacting across the Hsp90 C-terminal dimer interface presented in the closed, ATP conformation. The PPIase domain is positioned along the middle domain, adjacent Hsp90 client binding sites, while a single p23 makes stabilizing interactions with the N-terminal dimer. With this architecture, FKBP51 could thereby act on specific client residues presented during Hsp90-catalyzed remodeling.
Stress fiber strain recognition by the LIM protein testin is cryptic and mediated by RhoA
Authors: Sala Stefano , Oakes Patrick W. 
Published: January 2021

AbstractThe actin cytoskeleton is a key regulator of mechanical processes in cells. The family of LIM domain proteins have recently emerged as important mechanoresponsive cytoskeletal elements capable of sensing strain in the actin cytoskeleton. The mechanisms regulating this mechanosensitive behavior, however, remain poorly understood. Here we show that the LIM domain protein testin is peculiar in that despite the full-length protein primarily appearing diffuse in the cytoplasm, the C-terminal LIM domains alone recognize focal adhesions and strained actin while the N-terminal domains alone recognize stress fibers. Phosphorylation mutations in the dimerization regions of testin, however, reveal its mechanosensitivity and cause it to relocate to focal adhesions and sites of strain in the actin cytoskeleton. Finally, we demonstrate activated RhoA causes testin to adorn stress fibers and become mechanosensitive. Together, our data show that testin’s mechanoresponse is regulated in cells and provide new insights into LIM domain protein recognition of the actin cytoskeleton mechanical state.
Different resource allocation in aBacillus subtilispopulation displaying bimodal motility
Authors: Syvertsson Simon , Wang Biwen , Staal Jojet , Gao Yongqiang , Kort Remco , Hamoen Leendert W. 
Published: January 2021

ABSTRACTTo cope with sudden changes in their environment, bacteria can use a bet-hedging strategy by dividing the population into cells with different properties. This so-called bimodal or bistable cellular differentiation is generally controlled by positive feedback regulation of transcriptional activators. Due to the continuous increase in cell volume, it is difficult for these activators to reach an activation threshold concentration when cells are growing exponentially. This is one reason why bimodal differentiation is primarily observed from the onset of the stationary phase when exponential growth ceases. An exception is the bimodal induction of motility inBacillus subtilis, which occurs early during exponential growth. Several mechanisms have been put forward to explain this, including double negative-feedback regulation and the stability of the mRNA molecules involved. In this study, we used fluorescence-assisted cell sorting to compare the transcriptome of motile and non-motile cells and noted that expression of ribosomal genes is lower in motile cells. This was confirmed using an unstable GFP reporter fused to the strong ribosomalrpsDpromoter. We propose that the reduction in ribosomal gene expression in motile cells is the result of a diversion of cellular resources to the synthesis of the chemotaxis and motility systems. In agreement, single-cell microscopic analysis showed that motile cells are slightly shorter than non-motile cells, an indication of slower growth. We speculate that this growth rate reduction can contribute to the bimodal induction of motility during exponential growth.IMPORTANCETo cope with sudden environmental changes, bacteria can use a bet-hedging strategy and generate different types of cells within a population, so called bimodal differentiation. For example, aBacillus subtilisculture can contain both motile and non-motile cells. In this study we compared the gene expression between motile and non-motile cells. It appeared that motile cells express less ribosomes. To confirm this, we constructed a ribosomal promoter fusion that enabled us to measure expression of this promoter in individual cells. This reporter fusion confirmed our initial finding. The re-allocation of cellular resources from ribosome synthesis towards synthesis of the motility apparatus results in a reduction in growth. Interestingly, this growth reduction has been shown to stimulate bimodal differentiation.
Bacterial “galactosemia” is caused by cytoplasmic interference of an essential cell wall biosynthesis glycosyltransferase
Authors: Habib Cameron , Mueller Anna , Liu Ming , Zhu Jun , Schneider Tanja , Garner Ethan , Sun Yingjie , Chai Yunrong 
Published: January 2021

AbstractBacterial galactosemia or “galactose death,” triggered by incomplete galactose metabolism, was first discovered inEscherichia coliandSalmonellasix decades ago, and later in many other microorganisms, yet the mechanism for the toxicity and subsequent cell death remains unclear. InBacillus subtilis,galactosemia is manifested by a buildup of uridine-diphosphate-galactose (UDP-Gal) and a strong toxicity phenotype characterized by cell shape abnormality and rapid cell lysis. Here we present evidence that inB. subtilis,the toxicity is due to inhibition of cell wall biosynthesis through interference of the essential glycosyltransferase MurG that carries out lipid II synthesis from lipid I and uridine-diphosphate-N-acetyl-glucosamine (UDP-GlcNAc). Single-molecule imaging reveals real-time inhibition of cell wall biosynthesis and MurG activities in cells exhibiting toxicity. We further show thatin vitro, MurG is able to utilize UDP-Gal as a substrate generating a “toxic” lipid II, causing a potential poisoning effect on peptidoglycan crosslinking. Evidence also suggests a similar mechanism inVibrio choleraeandStaphylococcus aureus. Lastly, a strong synergistic lethality was seen inS. aureuswild-type cells treated with both galactose and sub-lethal doses of cell-wall antibiotics. Our study provides mechanistic explanation of the toxicity associated with bacterial galactosemia and its potential application in antibacterial solutions.SignificanceGalactosemia is a potentially fatal genetic disorder due to incomplete galactose metabolism, found in both eukaryotic and prokaryotic organisms. The molecular mechanisms of galactosemia-associated toxicity remain unclear in all cases. Here we present evidence that in the bacteriumBacillus subtilis, the toxicity is due to interference of an essential glycosyltransferase, MurG, which concerts lipid I to lipid II during peptidoglycan biosynthesis, by a nucleotide sugar derived from galactose metabolism. This interference leads to a halt of cell wall biosynthesis and structural defects causing rapid cell lysis. Our evidence also suggests a similar mechanism in other bacteria such asStaphylococcus aureusandVibrio cholerae.Our study may help solve the long-time puzzle of bacterial galactosemia first uncovered six decades ago.
Modeling post-logging height growth of black spruce forests by combining airborne LiDAR and historical forestry maps in eastern Canadian boreal forest
Authors: Bour Batistin , Danneyrolles Victor , Boucher Yan , Fournier Richard A. , Guindon Luc 
Published: January 2021

AbstractIncrease in forest disturbance due to land use as well as climate change has led to an expansion of young forests worldwide, which affects global carbon dynamics and forest management. In this study, we present a novel method that combines a single airborne LiDAR acquisition and historical harvesting maps to model height growth of post-logged black spruce-dominated forests in a 1700 km2eastern Canadian boreal landscape. We developed a random forest model where forest height is a function of stand age, combined with environmental variables. Our results highlight the strong predictive power of this model: least-square regression between predicted and observed height of our validation dataset was very close to the 1:1 relation and strongly supported by validation metrics (R2= 0.75; relative RMSE = 19%). Moreover, our findings indicated an ecological gradient responsible for differences in height growth at the landscape scale, with better growth rates on mesic slopes compared to badly drained soils on flat lands. With the increased availability of LiDAR data, this method is promising since it can be applied to forests across the globe that are affected by stand-replacing disturbances.