bluuebunny/crossref_metadata_2025 huggingface dataset:
Integrated protein-protein interaction and RNA interference screens reveal novel restriction and dependency factors for a tick-borne flavivirus in its human host
Authors: Sourisseau Marion , Unterfinger Yves , Lemasson Manon , Caignard Gregory , Piumi François , Grot Axel , Moutailler Sara , Vitour Damien , Coulpier Muriel , Lacour Sandrine A. , Richardson Jennifer
Published: November 2022

In Europe, tick-borne encephalitis virus (TBEV) is responsible for severe neurological disease in humans. Like other viruses, TBEV is an obligate intracellular life form whose survival requires subversion of metabolic processes and evasion of anti-viral pathways. This feat is achieved in no small part by binary interactions between dedicated viral proteins and host proteins. Such protein-protein interactions (PPI) constitute molecular determinants of critical pathobiologic traits of viruses, including host-range, zoonotic potential and virulence, and represent realistic targets for anti-viral therapies.

To shed light on the pathobiology of TBEV in human, we have resolved the network of PPI established with its human host by interaction proteomics. A high-throughput screen for virus-host PPI was performed involving the complete set of open reading frames of TBEV and the cDNA libraries ofHomo sapiens, by means of yeast two-hybrid methodology. We have discovered a large set of virus-host protein-protein interactions concerning 42 different human proteins directly interacting with nine viral proteins. Many of these human interactors have never been linked in the literature to viral infection.

The functional significance of the host interactors in viral infection as viral dependency or restriction factors was then characterizedin vitroby RNA interference, and their function inferred by bioinformatic analysis. Approximately 40% of the identified human proteins have a significative impact on TBEV viral replication. These are engaged in many biological processes, whose involvement in viral infection is expected for many, but enigmatic for some. Further work will be necessary to gain molecular understanding of how these biological processes support or restrict TBEV replication, and whether they constitute viral vulnerabilities that can be exploited therapeutically.
SOX9 elongates cell cycle phases and biases fate decisions in human intestinal stem cells
Authors: Burclaff Joseph , Bliton R. Jarrett , Breau Keith A , Cotton Michael J , Hinesley Caroline M , Ok Meryem T , Sweet Caden W , Zheng Anna , Bankaitis Eric D , Ariel Pablo , Magness Scott T
Published: November 2022

Background and Aims

The transcription factor SOX9 is expressed in many stem/progenitor cell populations and has biphasic correlations with proliferation rates across different biological systems. In murine intestinal crypts, distinct Sox9 levels mark three phenotypically different cell types, with lowest levels marking rapidly-dividing transit amplifying (TA) cells, intermediate levels marking intestinal stem cells (ISCs), and highest levels marking slowly-dividing label retaining secretory precursors. SOX9 expression levels and the impact of these levels on cell cycle and stem cell activity have not been characterized for humans.

Methods

Monolayers of primary human ISCs isolated from healthy organ donors were engineered with stable SOX9-knockout (KO) and/or SOX9-overexpression (OE) genomic modifications to assess the impact of SOX9 levels on proliferative capacity by DNA content analysis, cell cycle phase length by live imaging for a PIP-FUCCI reporter, stem cell activity via organoid formation assays, and cell fate after ISC differentiation tracked via qPCR.

Results

SOX9 was expressed at diverse levels in human intestinal crypt lineages in vivo, repressed proliferation in human ISC monolayers, and predominantly lengthened G1 by >40% with lesser lengthening of S and G2/M phases. Overexpression of SOX9 caused slower proliferation yet increased organoid forming efficiency. Higher SOX9 levels biased ISC differentiation towards tuft cell and follicle-associated epithelium fates while loss of SOX9 biased cells toward absorptive enterocyte, goblet cell, BEST4+cell, and enteroendocrine cell fates.

Conclusions

SOX9 is a master regulator of stem cell activity in human ISCs, lengthening the cell cycle, promoting stemness, and altering differentiation fate. Interestingly, differences are noted between species, highlighting the importance of analyzing regulatory mechanisms in primary healthy human cells.
Three decades of increasing fish biodiversity across the north-east Atlantic and the Arctic Ocean
Authors: Gordó-Vilaseca Cesc , Stephenson Fabrice , Coll Marta , Lavin Charles , Costello Mark John
Published: November 2022

Observed range shifts of numerous species support predictions of climate change models that species will shift their distribution northwards into the Arctic and sub-Arctic seas due to ocean warming. However, how this is affecting overall species richness is unclear. Here we analyse scientific research trawl surveys from the North Sea to the Arctic Ocean collected from 1994 to 2020, including 193 fish species. We found that demersal fish species richness at the local scale has doubled in some Arctic regions, including the Barents Sea, and increased at a lower rate at adjacent regions in the last three decades, followed by an increase in species richness and turnover at a regional scale. These changes in biodiversity paralleled an increase in sea bottom temperature. Within the study area, Arctic species’ probability of occurrence generally declined over time. However, the increase of species from southern latitudes, together with an increase of some Arctic species, ultimately led to an enrichment of the Arctic and sub-Arctic marine fauna due to increasing water temperature consistent with climate change.

Significance Statement

Global modelling studies suggest increased species arrivals from lower latitudes and local expirations at high latitudes due to global warming. Our analysis of 20,670 standardized scientific trawl surveys with 193 fish species from the north-east Atlantic and Arctic Oceans found an increase in species richness in the region parallel to an increase in sea bottom temperature. Some Arctic species declined in probability of occurrence over time, but some increased. This, together with the increase of southern-latitude species led to an enrichment of the Arctic and sub-Arctic marine fauna attributed to climate change.
The DNA binding protein BCL6 regulates NFκB-controlled endothelial inflammatory gene expression
Authors: Acevedo Adriana Franco , Mack Julia J. , Valenzuela Nicole M
Published: November 2022

Background

NFκB drives acute vascular inflammation by activating gene expression programs in endothelial cells to promote leukocyte recruitment. Numerous negative feedback regulators of NFκB activation have been defined that promote resolution of inflammation. However, the identities of endogenous suppressors of NFκB transcription are less clear. In macrophages, the transcriptional repressor BCL6 was shown to substantially overlap with NFκB-driven genes and influence the response to LPS. We identified that the DNA binding protein BCL6 was expressed in endothelial cells. Although the role of BCL6 in adaptive immune cells has been characterized, how BCL6 modifies transcription in endothelial cells has not been studied.

Objective

Based on prior knowledge that BCL6 represses part of the LPS-induced transcriptome in macrophages, we asked whether BCL6 regulated endothelial pro-inflammatory state by direct interaction with NFκB.

Methods

We analyzed public datasets of RNA and ChIP-Seq, probed BCL6 expression in human tissue, and tested BCL6 knockdown, overexpression and pharmacological manipulation on TNFα induced gene expressionin vitrousing human primary endothelium isolated from the heart.

Results

We demonstrate that the DNA binding protein BCL6 is basally expressed in the endothelium, with chromatin marks reflective of a superenhancer, and is particularly enriched in aortic endothelial cells (ECs) compared with ECs from other organs. Although basal expression was relatively low, BCL6 was rapidly upregulated in cardiac endothelium stimulated with TNFα, through direct action of NFκB. The BCL6 consensus DNA binding motif overlaps with that of NFκB. BCL6 target genes included endothelial pro-inflammatory chemokines and adhesion molecules, as well as NFκB-related genes themselves. BCL6 knockdown and the degrading BCL6 inhibitor BI-3802 augmented the endothelial cell response to TNFα. Surprisingly, antagonism of the BTB domain of BCL6 with small molecules 79-6, FX1 or BI-3812, blocked leukocyte adherence and accordingly suppressed both NFκB transcriptional activity as well as the expression of many genes in response to TNFα. Lastly, we show that HDAC activity is increased by TNFα, and can be reduced in the presence of BTB domain inhibitors.

Conclusions

Our results demonstrate that BCL6 is a repressor of NFκB-driven gene expression and inflammation in cardiac endothelial cells. These findings indicate that targeting of BCL6 may enhance vascular inflammation resolution.
Chytrids-conveyed long-chain polyunsaturated fatty acids to Daphnia alleviate the detrimental effect of heat when combined with limiting dietary organic matter quantity and nutritional quality
Authors: Abonyi András , Pilecky Matthias , Rasconi Serena , Ptacnik Robert , Kainz Martin J.
Published: November 2022

Global warming enhances the dominance of poorly palatable PUFA-deprived bloom-forming cyanobacteria. Chytrid fungal parasites increase herbivory and dietary access to polyunsaturated fatty acids (PUFA) across the phytoplankton-zooplankton interface. Little is known however about the role chytrids may play in compensating for the decrease of algae-derived PUFA under global warming scenarios. We tested experimentally the combined effects of water temperature increase and the presence of chytrids withDaphnia magnaas the consumer and the cyanobacteriumPlanktothrix rubescensas the main diet. We hypothesised that the diet including chytrids would enhanceDaphniafitness due to increased PUFA transfer irrespective of water temperature. Chytrid-infected diet significantly increasedDaphniasurvival, somatic growth, and reproduction, irrespective of water temperature. The PUFA content ofDaphniafeeding on the chytrid-infected diet was unaffected by heat at the onset of the first successful reproduction. Carbon stable isotopes of fatty acids highlighted preferential n-3 PUFA upgrading by chytrids and an ~3x higher endogenous n-3 PUFA conversion compared with n-6 PUFA byDaphnia, irrespective of water temperature. Diet including chytrids enhanced the retention of eicosapentaenoic acid (EPA; 20:5n-3) and arachidonic acid (ARA; 20:4n-6) inDaphnia. The heat did not decrease EPA and even increased ARA retention by enhanced endogenous bioconversion inDaphniawhen feeding on the chytrid-infected diet. We conclude that chytrids supportDaphniafitness at higher water temperatures via increased n-3 and n-6 PUFA retention and preferential n-3 PUFA bioconversion. Thus, they help function pelagic ecosystems with PUFA availability at the phytoplankton-zooplankton interface in a warmer climate.
An improved Erk biosensor reveals oscillatory Erk dynamics driven by mitotic erasure during early development
Authors: Wilcockson Scott G. , Guglielmi Luca , Rodriguez Pablo Araguas , Amoyel Marc , Hill Caroline S.
Published: November 2022

Erk signaling dynamics elicit distinct cellular responses in a variety of contexts. The early zebrafish embryo is an ideal model to explore the role of Erk signaling dynamicsin vivo, as a gradient of activated diphosphorylated Erk (P-Erk) is induced by Fgf signaling at the blastula embryonic margin. Here we describe an improved Erk-specific biosensor which we term modified Erk Kinase Translocation Reporter (modErk-KTR). We demonstrate the utility of this biosensorin vitroand in developing zebrafish andDrosophilaembryos. Moreover, we show that Fgf/Erk signaling is dynamic and coupled to tissue growth during both early zebrafish andDrosophiladevelopment. Signaling is rapidly extinguished just prior to mitosis, which we refer to as mitotic erasure, inducing periods of inactivity, thus providing a source of heterogeneity in an asynchronously dividing tissue. Our modified reporter and transgenic lines represent an important resource for interrogating the role of Erk signaling dynamicsin vivo.
Transcriptomic landscape of early hair follicle and epidermal development
Authors: Sulic Ana-Marija , Roy Rishi Das , Papagno Verdiana , Lan Qiang , Saikkonen Riikka , Jernvall Jukka , Thesleff Irma , Mikkola Marja L
Published: November 2022

Morphogenesis of ectodermal organs, such as hair, tooth, and mammary gland, starts with the formation of local epithelial thickenings, or placodes, but it remains to be determined how distinct cell types and differentiation programs are established during ontogeny. Here, we use bulk and single-cell transcriptomics and pseudotime modelling to address these questions in developing hair follicles and epidermis, and produce a comprehensive transcriptomic profile of cellular populations in the hair placode and interplacodal epithelium. We report previously unknown cell populations and marker genes, including early suprabasal and genuine interfollicular basal markers, and propose the identity of suprabasal progenitors. By uncovering four different hair placode cell populations organized in three spatially distinct areas, with fine gene expression gradients between them, we posit early establishment of cell fates. This work is accompanied by a readily accessible online tool to stimulate further research on skin appendages and their progenitors.
Glucocorticoid involvement in reproductive biology
Authors: Mullins Linda J. , Morley Steven D. , Kenyon Christopher J. , Mullins John J.
Published: November 2022

Oestrogen and progesterone play essential roles in the release of mature oocytes, the priming and cycling of the uterine lining, and the maintenance of mammalian pregnancy. Progesterone is synthesizedde novoat the embryo implantation site in the mouse, during decidualization of the endometrium. During early stages of pregnancy, the locally produced progesterone is thought to act as an immunosuppressant, preventing rejection of the fetal allograft at the fetal-maternal interface. However, both uterine natural killer cells and dendritic cells express glucocorticoid receptor rather than progesterone receptor. The importance of glucocorticoids in early pregnancy is inferred from the presence of steroid receptors and the 11β-hydroxysteroid dehydrogenase enzymes, which modulate corticosterone action in the decidua, the trophoblast, the placenta, and the fetus. 11β-hydroxylase is the last enzyme in the metabolism of cholesterol to corticosterone and, in a mouse model of 11β-hydroxylase deficiency, complications of reproduction suggested its requirement for normal ovulation and uterine cell turnover. We present evidence that, in this model, folliculogenesis occurs normally but ovulation is inhibited, and abnormal uterine cell turnover ultimately leads to adenomyosis. Ovaries respond to a superovulation protocol by releasing oocytes and forming corpora lutea, and homozygous null blastocysts are capable of implantation, but the pregnancy is not maintained. We show that glucocorticoid is produced locally at the implantation site in control animals, revealing wide involvement of glucocorticoids in reproductive biology.
Body mass decline in a Mediterranean community of solitary bees supports the size shrinking effect of climatic warming
Authors: Herrera Carlos M. , Núñez Alejandro , Valverde Javier , Alonso Conchita
Published: November 2022

The long-known, widely documented inverse relationship between body size and environmental temperature (“temperature-size rule”) has recently led to predictions of body size decline following current climatic warming (“size shrinking effect”). For keystone pollinators such as wild bees, body shrinking in response to warming can have pervasive effects on pollination processes, but there is still little evidence of the phenomenon because adequate tests require controlling for climate-linked confounding factors (e.g., urbanization, land use change). This paper tests the shrinking effect in a diverse community of solitary bees from well-preserved habitats in the core of a large nature reserve undergoing fast climatic warming but not disturbances or habitat changes. Long-term variation in mean body mass was evaluated using data from 1,186 individual bees (108 species, 25 genera, 6 families) sampled over 1990-2022. Climate of the region warmed at a fast rate during this period and changes in bee body mass verified expectations from the size shrinking effect. Mean individual body mass of the regional community of solitary bees declined significantly, shrinking being particularly intense for female individuals and large-bodied species. As a consequence, the pollination and mating systems of many bee-pollinated plants in the region are likely undergoing important alterations.
A Virus-Packageable CRISPR System Identifies Host Dependency Factors Across Multiple HIV-1 Strains
Authors: Montoya Vanessa R. , Ready Trine M. , Felton Abby , Fine Sydney R. , OhAinle Molly , Emerman Michael
Published: November 2022

At each stage of the HIV life cycle, host cellular proteins are hijacked by the virus to establish and enhance infection. We adapted the virus packageable HIV-CRISPR screening technology at a genome-wide scale to comprehensively identify host factors that affect HIV replication in a human T cell line. Using a smaller, targeted HIV Dependency Factor (HIVDEP) sub-library, we then performed screens across multiple HIV strains representing different clades and with different biological properties to define which T cell host factors are strain-specific versus which ones are important across all HIV strains and different clades. Nearly 90% genes selected across multiple host pathways validated in subsequent assays as bona fide host dependency factors including numerous proteins not previously reported to play role in HIV biology such as UBE2M, MBNL1, FBXW7, PELP1, SLC39A7, and others. Our ranked list of screen hits across multiple viral strains form a resource of HIV dependency factors for future investigation of host proteins involved in HIV biology.
Methylglyoxal controls tomato fruit ripening by regulating ethylene biosynthesis
Authors: Gambhir Priya , Raghuvanshi Utkarsh , Parida Adwaita Prasad , Kujur Stuti , Sharma Shweta , Sopory Sudhir K. , Kumar Rahul , Sharma Arun Kumar
Published: November 2022

Methylglyoxal (MG), a toxic compound produced as a byproduct in several cellular processes such as respiration and photosynthesis, is well investigated for its deleterious effects, mainly through glycation of proteins during plant stress responses. However, very little is known about its impact on fruit ripening. In the present study, we report that MG levels are maintained at high level in green tomato fruits, which declines during fruit ripening inspite of a respiratory burst during this transition. We demonstrate that this decline is mainly mediated by glutathione-dependent MG detoxification pathway and primarily catalyzed by glyoxalase enzyme encoded bySlGLY14gene.SlGLYI4is a direct target of MADS-RIN and is induced during fruit ripening. Silencing of this gene leads to drastic MG overaccumulation at ripening-stages in the transgenic fruits and interferes with the ripening process. Further investigations show that MG plausibly glycates and inhibits key enzymes such as methionine synthase (MS) and S-adenosyl methionine synthase (SAMS) of ethylene biosynthesis pathway, thereby indirectly affecting fruit pigmentation and cell was metabolism. MG overaccumulation in several non-ripening or inhibited- ripening tomato mutant fruits suggests the tightly regulated MG detoxification process is crucial for normal ripening program. Overall, we underpin aSlGLYI4-mediated novel regulatory mechanism of MG detoxification controlling fruit ripening in tomato.
Broad compatibility between yeast UAS elements and core promoters and identification of promoter elements that determine cofactor specificity
Authors: Schofield Jeremy A. , Hahn Steven
Published: November 2022

Three general classes of yeast protein-coding genes are distinguished by their dependence on the transcription cofactors TFIID, SAGA and Mediator (MED) Tail, but little is known about whether this dependence is determined by the core promoter, Upstream activation sites (UASs), or other gene features. It is also unclear whether UASs can broadly activate transcription from the different promoter classes or whether efficient transcription requires matching UASs and promoters of similar gene class. Here we measure transcription and cofactor specificity for tens of thousands of UAS-core promoter combinations. We find that <5% of UASs display strong core promoter specificity while most UASs can broadly activate promoters regardless of regulatory class. However, we find that matching UASs and promoters from the same gene class is generally important for optimal expression. From examining the cofactor dependence of this large UAS-promoter set, we find that sensitivity to rapid depletion of MED Tail or SAGA is dependent on the identity of both UAS and promoter while dependence on TFIID localizes to only the core promoter. Our results explain why transcription factor-mediated MED recruitment to the UAS does not always result in Tail-dependent transcription and highlight the role of TATA and TATA-like promoter sequences in MED Tail function.
CrusTome: A transcriptome database resource for large-scale analyses across Crustacea
Authors: Pérez-Moreno Jorge L. , Kozma Mihika T. , DeLeo Danielle M. , Bracken-Grissom Heather D. , Durica David S. , Mykles Donald L.
Published: November 2022

Transcriptomes from non-traditional model organisms often harbor a wealth of unexplored data. Examining these datasets can lead to clarity and novel insights in traditional systems, as well as to discoveries across a multitude of fields. Despite significant advances in DNA sequencing technologies and in their adoption, access to genomic and transcriptomic resources for non-traditional model organisms remains limited. Crustaceans, for example, being amongst the most numerous, diverse, and widely distributed taxa on the planet, often serve as excellent systems to address ecological, evolutionary, and organismal questions. While they are ubiquitously present across environments, and of economic and food security importance, they remain severely underrepresented in publicly available sequence databases. Here, we present CrusTome, a multi-species, multi-tissue, transcriptome database of 201 assembled mRNA transcriptomes (189 crustaceans, 30 of which were previously unpublished, and 12 ecdysozoan outgroups) as an evolving, and publicly available resource. This database is suitable for evolutionary, ecological, and functional studies that employ genomic/transcriptomic techniques and datasets. CrusTome is presented in BLAST and DIAMOND formats, providing robust datasets for sequence similarity searches, orthology assignments, phylogenetic inference, etc., and thus allowing for straight-forward incorporation into existing custom pipelines for high-throughput analyses. In addition, to illustrate the use and potential of CrusTome, we conducted phylogenetic analyses elucidating the identity and evolution of the Cryptochrome Photolyase Family of proteins across crustaceans.
Positive selection drivescis-regulatory evolution across the threespine stickleback Y chromosome
Authors: Shaw Daniel E. , Naftaly Alice Shanfelter , White Michael A.
Published: November 2022

Allele-specific gene expression evolves rapidly on heteromorphic sex chromosomes. Over time, the accumulation of mutations on the Y chromosome leads to widespread loss of gametolog expression, relative to the X chromosome. It remains unclear if expression evolution on degrading Y chromosomes is primarily driven by mutations that accumulate through processes of selective interference, or if positive selection can also favor the downregulation of coding regions on the Y chromosome that contain deleterious mutations. Identifying the relative rates ofcis-regulatory sequence evolution across Y chromosomes has been challenging due to the limited number of reference assemblies. The threespine stickleback (Gasterosteus aculeatus) Y chromosome is an excellent model to identify how regulatory mutations accumulate on Y chromosomes due to its intermediate state of divergence from the X chromosome. A large number of Y-linked gametologs still exist across three differently aged evolutionary strata to test these hypotheses. We found that putative enhancer regions on the Y chromosome exhibited elevated substitution rates and decreased polymorphism when compared to non-functional sites, like intergenic regions and synonymous sites. This suggests that manycis-regulatory regions are under positive selection on the Y chromosome. This divergence was correlated with X-biased gametolog expression, indicating the loss of expression from the Y chromosome may be favored by selection. Our findings provide evidence that Y-linkedcis-regulatory regions exhibit signs of positive selection quickly after the suppression of recombination and allow comparisons with recent theoretical models that suggest the rapid divergence of regulatory regions may be favored to mask deleterious mutations on the Y chromosome.
The structure of a Type III-A CRISPR-Cas effector complex reveals conserved and idiosyncratic contacts to target RNA and crRNA among Type III-A systems
Authors: Paraan Mohammadreza , Nasef Mohamed , Chou-Zheng Lucy , Khweis Sarah A. , Schoeffler Allyn J. , Hatoum-Aslan Asma , Stagg Scott M. , Dunkle Jack A.
Published: November 2022

Type III CRISPR-Cas systems employ multiprotein effector complexes bound to small CRISPR RNAs (crRNAs) to detect foreign RNA transcripts and elicit a complex immune response that leads to the destruction of invading RNA and DNA. Type III systems are among the most widespread in nature, and emerging interest in harnessing these systems for biotechnology applications highlights the need for detailed structural analyses of representatives from diverse organisms. We performed cryo-EM reconstructions of the Type III-A Cas10-Csm effector complex fromS. epidermidisbound to an intact, cognate target RNA and identified two oligomeric states, a 276 kDa complex and a 318 kDa complex. 3.1 Å density for the well-ordered 276 kDa complex allowed construction of atomic models for the Csm2, Csm3, Csm4 and Csm5 subunits within the complex along with the crRNA and target RNA. We also collected small-angle X-ray scattering data which was consistent with the 276 kDa Cas10-Csm architecture we identified. Detailed comparisons between theS. epidermidisCas10-Csm structure and the well-resolved bacterial (S. thermophilus) and archaeal (T. onnurineus) Cas10-Csm structures reveal differences in how the complexes interact with target RNA and crRNA which are likely to have functional ramifications. These structural comparisons shed light on the unique features of Type III-A systems from diverse organisms and will assist in improving biotechnologies derived from Type III-A effector complexes.
A gene-level test for directional selection on gene expression
Authors: Colbran Laura L. , Ramos-Almodovar Fabian C. , Mathieson Iain
Published: November 2022

Most variants identified in human genome-wide association studies and scans for selection are non-coding. Interpretation of these variants’ effects and understanding of the way in which they contribute to phenotypic variation and adaptation in human populations is therefore limited by our understanding of gene regulation and by the difficulty in confidently linking non-coding variants to genes. To overcome this, we developed a gene-by-gene test for population-specific selection based on combinations of regulatory variants.

We extended theQXtest for polygenic selection to test for selection on regulatory variants for 17,388 protein-coding genes across 2,504 individuals. We identified 45 genes with significant evidence (FDR <0.1) for selection, includingFADS1,KHK,SULT1A2,ITGAM, and genes in the HLA region. We further confirm that significant selection signals do correspond to plausible population-level differences in predicted expression. However, we find that very few (0.2%) genes have strong evidence for directional, population-specific selection on the component of their expression that is predicted bycis-regulatory variants. While this is consistent with mostcis-regulatory variation evolving under genetic drift or stabilizing selection, it is also possible that any effects are smaller than we can detect, or that population-specific selection is driven by tissue-specific ortranseffects.

Our gene-levelQXscore is independent of other methods for detecting selection based on genomic variation, may therefore be useful when used in combination with more traditional selection tests to specifically identify selection on regulatory variation. Overall, our results demonstrate the utility of one approach to combining population-level information with functional data to understand the evolution of gene expression.
Comparative proteome analysis of differentSaccharomyces cerevisiaestrains during growth on sucrose and glucose
Authors: Rodrigues Carla Inês Soares , den Ridder Maxime , Pabst Martin , Gombert Andreas K. , Wahl Sebastian Aljoscha
Published: November 2022

Both the identity and the amount of a carbon source present in laboratory or industrial cultivation media have major impacts on the growth and physiology of a microbial species. In the case of the yeastSaccharomyces cerevisiae, sucrose is arguably the most important sugar used in industrial biotechnology, whereas glucose is the most common carbon and energy source used in research, with many well-known and described regulatory effects, e.g. glucose repression. Here we compared the label-free proteomes of exponentially growingS. cerevisiaecells in a defined medium containing either sucrose or glucose as the sole carbon source. For this purpose, bioreactor cultivations were employed, and three different strains were investigated, namely: CEN.PK113-7D (a common laboratory strain), UFMG-CM-Y259 (a wild isolate), and JP1 (an industrial bioethanol strain). These strains present different physiologies during growth on sucrose; some of them reach higher specific growth rates on this carbon source, when compared to growth on glucose, whereas others display the opposite behavior. It was not possible to identify proteins that commonly presented either higher or lower levels during growth on sucrose, when compared to growth on glucose, considering the three strains investigated here, except for one protein, named Mnp1 – a mitochondrial ribosomal protein of the large subunit, which had higher levels on sucrose than on glucose, for all three strains. Interestingly, following a Gene Ontology overrepresentation and KEGG pathway enrichment analyses, an inverse pattern of enriched biological functions and pathways was observed for the strains CEN.PK113-7D and UFMG-CM-Y259, which is in line with the fact that whereas the CEN.PK113-7D strain grows faster on glucose than on sucrose, the opposite is observed for the UFMG-CM-Y259 strain.
Stronger between-clan than within-clan contests and their ecological correlates in a non-territorial, fission-fusion species, the Asian elephant
Authors: Gautam Hansraj , Vidya T.N.C.
Published: November 2022

The authors have withdrawn this manuscript owing to duplicate posting. Therefore, the authors do not wish this work to be cited as reference for the project. If you have any questions, please contact the corresponding author.

The correct manuscript can be found athttps://doi.org/10.1101/754515.
Mapping of the podocin proximity-dependent proteome reveals novel components of the kidney podocyte foot process
Authors: Gerlach Gary F. , Imseis Zachary H. , Cooper Shamus L. , Santos Anabella N. , O’Brien Lori L.
Published: November 2022

The unique architecture of glomerular podocytes is integral to kidney filtration. Interdigitating foot processes extend from the podocyte cell body, wrap around fenestrated capillaries, and form specialized junctional complexes termed slit diaphragms to create a molecular sieve. However, the full complement of proteins which maintain foot process integrity, and how this localized proteome changes with disease, remains to be elucidated. Proximity-dependentbiotinidentification (BioID) enables the identification of spatially localized proteomes. To this end, we developed a novelin vivoBioID knock-in mouse model. We utilized the slit diaphragm protein podocin (Nphs2) to create a podocin-BioID fusion. Podocin-BioID localizes to the slit diaphragm and biotin injection leads to podocyte-specific protein biotinylation. We isolated the biotinylated proteins and performed mass spectrometry to identify proximal interactors. Gene ontology analysis of 54 proteins specifically enriched in our podocin-BioID sample revealed ‘cell junctions’, ‘actin binding’, and ‘cytoskeleton organization’ as top terms. Known foot process components were identified and we further uncovered two novel proteins: the tricellular junctional protein Ildr2 and the CDC42 and N-WASP interactor Fnbp1l. We confirmed Ildr2 and Fnbp1l are expressed by podocytes and partially colocalize with podocin. Finally, we investigated how this proteome changes with age and uncovered a significant increase in Ildr2. This was confirmed by immunofluorescence on human kidney samples and suggests altered junctional composition may preserve podocyte integrity. Together, these assays have led to new insights into podocyte biology and supports the efficacy of utilizing BioIDin vivoto interrogate spatially localized proteomes in health, aging, and disease.
Magnetic Levitation and Sorting of Neoplastic Circulating Cell Hybrids
Authors: Liang Kaitlyn , Yaman Sena , Patel Ranish K. , Parappilly Michael S. , Walker Brett S. , Wong Melissa H. , Durmus Naside Gozde
Published: November 2022

Circulating hybrid cells (CHCs) are a novel, rare cell population that harbor tumor and immune cell phenotypes and genotypes and are detectible in peripheral blood. Several recent reports implicated CHCs in the metastatic cascade and found their enumeration to provide better prognostic value than conventionally-defined circulating tumor cells (CTCs). However, methods for isolation and enrichment of CHCs are not well-studied or established. Here, we developed an ultrasensitive, antigen-independent platform leveraging the principles of magnetic levitation for the detection and isolation of disseminated neoplastic CHCs. For the first time, we demonstrate that CHCs can be magnetically focused to different levitation heights, under various paramagnetic conditions using a static levitation system, and we quantified the biophysical properties of CHCs (i.e., levitation heights). In addition, we investigated whether magnetic levitation approach can be combined with the affinity-based strategies to enrich CHCs under the magnetic field. Using clinical samples from breast and colorectal cancer patients, we demonstrated that neoplastic cells can be sorted with a magnetic levitation-based sorting device, without relying on any surface markers. Overall, we demonstrated the feasibility of the magnetic levitation method for unbiased enrichment of rare neoplastic-immune hybrid cells from peripheral blood specimens from cancer patients. This approach can be expanded to more clinical samples and cancer types to unprecedentedly explore the biology of rare neoplastic cells and develop metastasis-tailored therapies broadly impacting personalized and precision clinical treatments.
A retrograde GCN2/eIF2α, but ATF4 independent, mechanism maintains synaptic strength under acute amino acid scarcity at the NMJ
Authors: Kauwe Grant , Mori Megumi , Liao Edward H. , Scott Gary , Haghighi A. Pejmun
Published: November 2022

Neuronal response to nutrient availability plays an important role in the maintenance of cellular homeostasis and behavioral response to the environment in higher eukaryotes. However, we know little about how neuronal function is influenced by acute changes in nutrients at high resolution. Taking advantage of powerful fly genetics and the amenability of theDrosophilalarval neuromuscular junction (NMJ), we have investigated the synaptic response to acute amino acid restriction. Our findings indicate that the presence of general control nonderepressible 2 (GCN2) and phosphorylation of its target eukaryotic initiation factor 2 alpha (eIF2α) are essential for the ability of the NMJ to maintain normal neurotransmitter output when the larvae are deprived of amino acids. Surprisingly, activating transcription factor 4 (ATF4), which normally acts downstream of GCN2/eIF2α, appears dispensable in this regulation. Furthermore, we show that GCN2/eIF2α dependent cascade acts retrogradely from muscle back to motoneuron to adjust synaptic release. These results provide a mechanistic insight into the intricate regulation of synaptic strength through the action of GCN2 when organisms are faced with amino acid scarcity.
Pupil size reflects activation of subcortical ascending arousal system nuclei during rest
Authors: Lloyd Beth , de Voogd Lycia D. , Mäki-Marttunen Verónica , Nieuwenhuis Sander
Published: November 2022

Neuromodulatory nuclei that are part of the ascending arousal system (AAS) play a crucial role in regulating cortical state and optimizing task performance. Pupil diameter, under constant luminance conditions, is increasingly used as an index of activity of these AAS nuclei. Indeed, task-based functional imaging studies in humans have begun to provide evidence of stimulus-driven pupil-AAS coupling. However, whether there is such a tight pupil-AAS coupling during rest is not clear. To address this question, we examined simultaneously acquired resting-state fMRI and pupil-size data from 74 participants, focusing on six AAS nuclei: the locus coeruleus, ventral tegmental area, substantia nigra, dorsal and median raphe nuclei, and cholinergic basal forebrain. Activation in all six AAS nuclei was optimally correlated with pupil size at 0-to 2-second lags, suggesting that spontaneous pupil changes were almost immediately followed by corresponding BOLD-signal changes in the AAS. These results suggest that spontaneous changes in pupil size that occur during states of rest can be used as a noninvasive general index of activity in AAS nuclei. Importantly, the nature of pupil-AAS coupling during rest appears to be vastly different from the relatively slow canonical hemodynamic response function that has been used to characterize task-related pupil-AAS coupling.
Patterning precision under non-linear morphogen decay and molecular noise
Authors: Adelmann Jan A. , Vetter Roman , Iber Dagmar
Published: November 2022

Morphogen gradients can instruct cells about their position in a patterned tissue. Non-linear morphogen decay has been suggested to increase gradient precision by reducing the sensitivity to variability in the morphogen source. Here, we use cell-based simulations to quantitatively compare the positional error of gradients for linear and non-linear morphogen decay. While we confirm that non-linear decay reduces the positional error close to the source, the reduction is very small for physiological noise levels. Far from the source, the positional error is much larger for non-linear decay in tissues that pose a flux barrier to the morphogen at the boundary. In light of this new data, a physiological role of morphogen decay dynamics in patterning precision appears unlikely.
Development of a high-throughput starch digestibility assay reveals wide variation among the A. E. Watkins wheat landrace collection
Authors: Zafeiriou Petros , Savva George M. , Ahn-Jarvis Jennifer H. , Warren Frederick J. , Pasquariello Marianna , Griffiths Simon , Seung David , Hazard Brittany A.
Published: November 2022

Breeding for less digestible starch in wheat can improve the health impact of bread and other wheat foods. Based on an establishedin vitrostarch digestibility assay by Edwards et al. (2019) we developed a high-throughput assay to measure starch digestibility in hydrothermally processed samples for use in forward genetic approaches. Digestibility of purified starch from maize and wheat was measured using both methods and produced comparable results. Using the high-throughput assay, we estimated starch digestibility of 118 wheat landraces from the core Watkins collection and found wide variation across lines and elite UK varieties, (20% to 40% and 31% to 44% starch digested after 90 minutes respectively). Sieved flour fractions and purified starch for selected lines showed altered starch digestibility profiles compared with wholemeal flour, suggesting that matrix properties of flour rather than intrinsic properties of starch granules conferred the low starch digestibility observed.
Peptide-mediated inhibition of the transcriptional regulator Elongin BC induces apoptosis in cancer cells
Authors: Fischer Sabrina , Trinh Van Tuan , Simon Clara , Weber Lisa Marie , Formé Ignasi , Nist Andrea , Bange Gert , Abendroth Frank , Stiewe Thorsten , Steinchen Wieland , Liefke Robert , Vázquez Olalla
Published: November 2022

Inhibition of protein-protein interactions (PPIs) via designed peptides is an effective strategy to interfere with their biological functions. The Elongin BC heterodimer (ELOB/C) is involved in transcription elongation and protein turnover by PPIs that involve the so-called BC-box. ELOB and ELOC are commonly upregulated in cancer and essential for cancer cell growth, making them attractive drug targets. However, no strategy has been established to inhibit their functions in cells, so far. Here, we report a peptide that mimics a high-affinity BC-box and tightly binds to the ELOB/C dimer (kD= 0.45 ± 0.03 nM). Our peptide blocks the association of ELOB/C with its interaction partners, both in vitro and in the cellular environment. Cancer cells treated with this peptide inhibitor show decreased cell viability, altered cell cycle and increased apoptosis. Therefore, our work proposes that blocking the BC-box binding pocket of ELOB/C is a feasible strategy to impair the function of the ELOB/C heterodimer and inhibit cancer cell growth. Our peptide inhibitor promises novel mechanistic insights into the biological function of the ELOB/C dimer and offers a starting point for therapeutics linked to ELOB/C dysfunction.
Shared requirement for MYC upstream super-enhancer region in tissue regeneration and cancer
Authors: Sur Inderpreet , Zhao Wenshuo , Zhang Jilin , Pilström Margareta Kling , Rozell Björn , Webb Anna T. , Cheng Huaitao , Ristimäki Ari , Katajisto Pekka , Enge Martin , Rannikmae Helena , de la Roche Marc , Taipale Jussi
Published: November 2022

Cancer has been characterized as a wound that does not heal. Malignant cells are morphologically distinct from normal proliferating cells, but have extensive similarities to tissues undergoing wound healing and/or regeneration. The mechanistic basis of this similarity has, however, remained enigmatic. Here we show that the genomic region upstream of Myc, which is required for intestinal tumorigenesis, is also required for intestinal regeneration after radiation damage. The region is also critical for growth of adult intestinal cells in 3D organoid culture, indicating that culture conditions that recapitulate most aspects of normal tissue architecture still reprogram normal cells to proliferate using a mechanism similar to that employed by cancer cells. Our results uncover a genetic link between tissue regeneration and tumorigenesis, and establish that normal tissue renewal and regeneration of tissues after severe damage are mechanistically distinct.

Myc regulatory region links regeneration and cancer
Modest transcriptomic response to polyploidization in allohexaploid wheat synthetics
Authors: Banouh Meriem , Armisen David , Bouguennec Annaig , Huneau Cecile , Sow Mamadou Dia , Pont Caroline , Salse Jerome , Civan Peter
Published: November 2022

Bread wheat is a recent allohexaploid (genomic constitution AABBDD) that emerged through a hybridization between tetraploidTriticum turgidum(AABB) and diploidAegilops tauschii(DD) <10,000 years ago. The hexaploidization can be re-created artificially, producing synthetic wheat that has been used to study immediate genomic responses to polyploidization. Here we produced several synthetic wheats from alternative parental genotypes and reciprocal crosses, and examined transcriptomes from two different tissues and successive generations. We did not detect a massive reprogramming in gene expression, with only ∼1% of expressed genes showing significant differences compared to their lower-ploidy parents. Most of the differential expression is located on the D subgenome, without consistency in the direction of the expression change. Leaves and developing endosperm show distinct patterns of homoeologous expression bias, and almost non-overlapping sets of differentially expressed genes, implying that the polyploidization-triggered reprogramming is not effectuated through permanent (epi)genetic changes. While 0-3 families of transposable elements (TEs) became upregulated in wheat synthetics, we did not detect any significant association between TEs and the expression of nearby genes. We conclude that the modest tissue-specific and partially genotype-specific transcriptomic response to polyploidization is likely caused by rare incompatibilities of parental regulomes, and we discuss the pitfalls of transcriptomic comparisons across ploidy levels that can inflate the de-regulation signal.
A primary sensory cortical interareal feedforward inhibitory circuit for tacto-visual integration
Authors: Weiler Simon , Rahmati Vahid , Isstas Marcel , Wutke Johann , Stark Andreas Walter , Franke Christian , Geis Christian , Witte Otto W. , Hübener Mark , Bolz Jürgen , Margrie Troy W. , Holthoff Knut , Teichert Manuel
Published: November 2022

Tactile sensation and vision are often both utilized for the exploration of objects that are within reach though it is not known whether or how these two distinct sensory systems might combine such information. Here in mice we find that stimulation of the contralateral whisker array suppresses visually evoked activity in a subarea of primary visual cortex (VISp) whose visual space covers the whisker search space. This is mediated by local fast spiking interneurons that receive a direct cortico-cortical input predominantly from layer 6 of the primary somatosensory barrel cortex (SSp-bfd). These data demonstrate functional convergence within and between two primary sensory cortical areas for multisensory object detection and recognition.