PaperPlayer biorxiv molecular biology

Link to bioRxiv paper:

http://biorxiv.org/cgi/content/short/2020.08.06.239343v1?rss=1

Authors: Castro-Cruz, M., Lembo, F., Borg, J.-P., Trave, G., Vincentelli, R., Zimmermann, P.

Abstract:

PSD95-disc large-zonula occludens (PDZ) domains are globular modules of 80-90 amino acids that co-evolved with multicellularity. They commonly bind to carboxy-terminal sequences of a plethora of membrane-associated proteins and influence their trafficking and signaling. We previously built a PDZ resource (PDZome) allowing to unveil human PDZ interactions by Yeast two-hybrid. Yet, this resource is partial according to the current knowledge on the human PDZ proteome. Here we built the PDZome 2.0 library for Yeast two-hybrid, based in a PDZ library manually curated from online resources. The PDZome2.0 contains 305 individual clones (266 PDZ domains in isolation and 39 tandems), for which all boundaries have been designed based on available PDZ structures. Using as bait the E6 oncoprotein from HPV16, a known promiscuous PDZ interactor, we show that PDZome 2.0 outperforms the previous resource.

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Link to bioRxiv paper:

http://biorxiv.org/cgi/content/short/2020.08.06.239418v1?rss=1

Authors: de Wolf, B., Oghabian, A., Akinyi, M. V., Hanks, S., Tromer, E. C., van Hooff, J., van Voorthuijsen, L., van Rooijen, L. E., Verbeeren, J., Uijttewaal, E. C. H., Baltissen, M. P. A., Yost, S., Piloquet, P., Vermeulen, M., Snel, B., Isidor, B., Rahman, N., Frilander, M. J., Kops, G. J. P. L.

Abstract:

Aneuploidy is the leading cause of miscarriage and congenital birth defects, and a hallmark of cancer. Despite this strong association with human disease, the genetic causes of aneuploidy remain largely unknown. Through exome sequencing of patients with constitutional mosaic aneuploidy, we identified biallelic truncating mutations in CENATAC (CCDC84). We show that CENATAC is a novel component of the minor (U12-dependent) spliceosome that promotes splicing of a specific, rare minor intron subtype. This subtype is characterized by AT-AN splice sites and relatively high basal levels of intron retention. CENATAC depletion or expression of disease mutants resulted in excessive retention of AT-AN minor introns in ~100 genes enriched for nucleocytoplasmic transport and cell cycle regulators, and caused chromosome segregation errors. Our findings reveal selectivity in minor intron splicing with a specific impact on the chromosome segregation process, and show how defects herein can cause constitutional aneuploidy.

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Link to bioRxiv paper:

http://biorxiv.org/cgi/content/short/2020.08.06.239251v1?rss=1

Authors: Aughey, G. N., Delandre, C., Southall, T. D., Marshall, O. J.

Abstract:

Targeted DamID (TaDa) is an increasingly popular method of generating cell-type specific DNA binding profiles in vivo. Although sensitive and versatile, TaDa requires the generation of new transgenic fly lines for every protein that is profiled, which is both time-consuming and costly. Here, we describe the FlyORF-TaDa system for converting an existing FlyORF library of inducible open reading frames (ORFs) to TaDa lines via a genetic cross, with recombinant progeny easily identifiable by eye colour. Profiling the binding of the H3K36me3-associated chromatin protein MRG15 in larval neural stem cells using both FlyORF-TaDa and conventional TaDa demonstrates that new lines generated using this system provide accurate and highly-reproducible DamID binding profiles. Our data further show that MRG15 binds to a subset of active chromatin domains in vivo. Courtesy of the large coverage of the FlyORF library, the FlyORF-TaDa system enables the easy creation of TaDa lines for 74% of all transcription factors and chromatin modifying proteins within the Drosophila genome.

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Link to bioRxiv paper:

http://biorxiv.org/cgi/content/short/2020.08.05.238543v1?rss=1

Authors: Bates, T. A., Chuong, C., Hawks, S. A., Rai, P., Salgado, R. M., Duggal, N. K., Weger-Lucarelli, J. A.

Abstract:

Usutu virus (USUV; genus Flavivirus; family Flaviviridae) is a mosquito-borne, positive-sense RNA virus that is currently causing significant die-offs in numerous bird species throughout Europe and has caused infections in humans. Currently, there are no molecular clones for USUV, hence, hindering studies on the pathogenesis and transmission of USUV. In this report, we demonstrate the development and characterization of infectious clones for two modern strains of USUV isolated from Europe and Africa. We show that the infectious clone-derived viruses replicated similarly to the parental strains in both mammalian and insect cells. Additionally, we observed similar levels of replication and pathogenesis in two mouse models. This reverse genetics system will aid the scientific community in studying and developing USUV infection, transmission, diagnostics, and vaccines.

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Link to bioRxiv paper:

http://biorxiv.org/cgi/content/short/2020.08.06.240192v1?rss=1

Authors: Osipiuk, J., Azizi, S.-A., Dvorkin, S., Endres, M., Jedrzejczak, R., Jones, K. A., Kathayat, R. S., Lisnyak, V. G., Maki, S. L., Kang, S., Kim, Y., Nicolaescu, V., Taylor, C. A., Tesar, C., Zhang, Y.-A., Zhou, Z., Randall, G., Michalska, K., Snyder, S. A., Dickinson, B. C., Joachimiak, A.

Abstract:

The number of new cases world-wide for the COVID-19 disease is increasing dramatically, while efforts to contain Severe Acute Respiratory Syndrome Coronavirus 2 is producing varied results in different countries. There are three key SARS-CoV-2 enzymes potentially targetable with antivirals: papain-like protease (PLpro), main protease (Mpro), and RNA-dependent RNA polymerase. Of these, PLpro is an especially attractive target because it plays an essential role in several viral replication processes, including cleavage and maturation of viral polyproteins, assembly of the replicasetranscriptase complex (RTC), and disruption of host viral response machinery to facilitate viral proliferation and replication. Moreover, this enzyme is conserved across different coronaviruses and promising inhibitors have already been discovered for its SARS-CoV variant. Here we report a substantive body of structural, biochemical, and virus replication studies that identify several inhibitors of the enzyme from SARS-CoV-2 in both wild-type and mutant forms. These efforts include the first structures of wild-type PLpro, the active site C111S mutant, and their complexes with inhibitors, determined at 1.60-2.70 Angstroms. This collection of structures provides fundamental molecular and mechanistic insight to PLpro, and critically, illustrates details for inhibitors recognition and interactions. All presented compounds inhibit the peptidase activity of PLpro in vitro, and some molecules block SARS-CoV-2 replication in cell culture assays. These collated findings will accelerate further structure-based drug design efforts targeting PLpro, with the ultimate goal of identifying high-affinity inhibitors of clinical value for SARS-CoV-2.

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Link to bioRxiv paper:

http://biorxiv.org/cgi/content/short/2020.08.05.237651v1?rss=1

Authors: Hoffmann, M., Hofmann-Winkler, H., Smith, J. C., Krueger, N., Sorensen, L. K., Sogaard, O. S., Hasselstrom, J. B., Winkler, M., Hempel, T., Raich, L., Olsson, S., Yamazoe, T., Yamatsuta, K., Mizuno, H., Ludwig, S., Noe, F., Sheltzer, J. M., Kjolby, M., Poehlmann, S.

Abstract:

Antiviral therapy is urgently needed to combat the coronavirus disease 2019 (COVID-19) pandemic, which is caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). The protease inhibitor camostat mesylate inhibits SARS-CoV-2 infection of lung cells by blocking the virus-activating host cell protease TMPRSS2. Camostat mesylate has been approved for treatment of pancreatitis in Japan and is currently being repurposed for COVID-19 treatment. However, potential mechanisms of viral resistance as well as camostat mesylate metabolization and antiviral activity of metabolites are unclear. Here, we show that SARS-CoV-2 can employ TMPRSS2-related host cell proteases for activation and that several of them are expressed in viral target cells. However, entry mediated by these proteases was blocked by camostat mesylate. The camostat metabolite GBPA inhibited the activity of recombinant TMPRSS2 with reduced efficiency as compared to camostat mesylate and was rapidly generated in the presence of serum. Importantly, the infection experiments in which camostat mesylate was identified as a SARS-CoV-2 inhibitor involved preincubation of target cells with camostat mesylate in the presence of serum for 2 h and thus allowed conversion of camostat mesylate into GBPA. Indeed, when the antiviral activities of GBPA and camostat mesylate were compared in this setting, no major differences were identified. Our results indicate that use of TMPRSS2-related proteases for entry into target cells will not render SARS-CoV-2 camostat mesylate resistant. Moreover, the present and previous findings suggest that the peak concentrations of GBPA established after the clinically approved camostat mesylate dose (600 mg/day) will result in antiviral activity.

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Link to bioRxiv paper:

http://biorxiv.org/cgi/content/short/2020.08.04.236638v1?rss=1

Authors: Wolf, J. F., Kriss, K. D., MacAulay, K. M., Munro, K., Patterson, B. R., Shafer, A. B. A.

Abstract:

Individuals exhibit differences in their microbial composition that have important implications for both population dynamics and ecological processes. The gut microbiome of animals can vary by age, reproductive status, diet, and habitat quality, and directly influences an individuals health and fitness. Likewise, variation in an individuals home range can lead to differences in reproductive behaviour, feeding strategies, and fitness. Ungulates (hooved mammals) exhibit species-specific microbiomes and habitat use patterns that differ by season, sex, and age-class, leading to variation among individuals occupying a similar geographic area. Here, we combined fecal microbiome and movement data to assess the relationship between space use and the gut microbiome in a specialist and a generalist ungulate. We captured and GPS radiocollared 24 mountain goats (Oreamnos americanus) and 34 white-tailed deer (Odocoileus virginianus). During captures we collected fresh fecal samples and conducted high-throughput sequencing of the fecal microbiome (i.e. 16S rRNA gene) to quantify gut microbial diversity. We generated Brownian Bridge Movement Models from the GPS location data to estimate core (50%) and home range (95%) sizes and calculated the proportion of use for several important habitat types. An increase in Firmicutes to Bacteroides ratios corresponded to an increase in core range area in both species. In mountain goats we observed a negative relationship between gut diversity and use of both escape terrain and treed habitat, both critical features for this alpine specialist. There were no relationships between habitat use and the gut microbiome in the more generalist white-tailed deer. We hypothesize that larger Firmicutes to Bacteroides ratios confer body size or fat advantages that allow for larger home ranges and that relationships between gut diversity and disproportionate use of particular habitats is stronger in mountain goats due to their restricted niche relative to white-tailed deer. This is the first study to relate core range size to gut diversity in wild ungulates and is an important proof of concept that advances the potential type of information that can be gleaned from non-invasive sampling.

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Link to bioRxiv paper:

http://biorxiv.org/cgi/content/short/2020.08.04.235887v1?rss=1

Authors: Thevkar-Nagesh, P., Rawal, S., Zahr, T., Fisher, E. A., Garabedian, M.

Abstract:

Acyl-CoA synthetase 1 (ACSL1) is an enzyme that converts fatty acids to acyl-CoA-derivatives for use in both lipid catabolism and lipid synthesis, including of arachidonic acid mediators that promote inflammation. ACSL1 has also been linked to the pro-atherosclerotic effects of diabetes in mice. ACSL1 expression has been reported to be upregulated in monocytes and macrophages by hyperglycemia, as well as enhanced by inflammatory stimuli, yet surprisingly little is known about the mechanisms underlying its transcriptional regulation. Herewe show that increased Acsl1 mRNA expression in mouse macrophages by hyperglycemia is via transcription initiation such that nascent ACSL1 RNA and Acsl1 promoter activity are increased. We further demonstrate that the hyperglycemic-dependent induction of Acsl1 mRNA is governed by the glucose-sensing transcription factor, Carbohydrate Response Element Binding Protein (CHREBP), since the hyperglycemic upregulation of Acsl1 mRNA is lost in mouse bone marrow derived macrophages (BMDMs) from Chrebp knock out mice. In addition, we show that LPS treatment of mouse BMDMs increased Acsl1 mRNA, and this is attenuated by an NF-kappa Binhibitor that blocks p65 subunit binding to DNA. We further show that LPS treatment increased ACSL1 protein abundance and stimulated ACSL1 protein localization to membranes where it likely exerts its activity. Using an ACSL1 reporter gene containing the promoter and 1.6 Kb of upstream regulatory region, which contain multiple predicted CHREBP and NF-kappa B(RELA) binding sites conserved between the human and mouse ACSL1 gene, we found a synergistic increase of ACSL1 promoter activity when CHREBP and RELA were co-expressed. Thus, we have identified pathways controlling the expression of ACSL1 by hyperglycemia and inflammation through CHREBP and NF-kappa B.

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Link to bioRxiv paper:

http://biorxiv.org/cgi/content/short/2020.08.05.237586v1?rss=1

Authors: Trieu-Duc, V., Iwasaki, Y., Oshima, K., Nikaido, M., Chiu, M.-T., Okada, N.

Abstract:

Territorial defense involves frequent aggressive confrontations with competitors, but little is known about how brain-transcriptomic profiles change between individuals competing for territory establishment. Our previous study elucidated that brain-transcriptomic synchronization occurs in a pair-specific manner between two males of the fish Betta splendens during fighting, reflecting a mutual assessment process between them at the level of gene expression. Here we evaluated how the brain-transcriptomic profiles of opponents change immediately after shifting their social status (i.e., the winner/loser has emerged) and 30 min after this shift. We showed that unique and carryover hypotheses can be adapted to this system, in which changes in the expression of certain genes are unique to different fighting stages and in which the expression patterns of certain genes are transiently or persistently changed across all fighting stages. Interestingly, the specificity of the brain-transcriptomic synchronization of a pair during fighting was gradually lost after fighting ceased, because of the decrease in the variance in gene expression across all individuals, leading to the emergence of a basal neurogenomic state. Strikingly, this unique state was more basal than the state that existed in the before-fighting group and resulted in the reduced and consistent expression of genes across all individuals. In spite of the consistent and basal overall gene expression in each individual in this state, expression changes for genes related to metabolism, learning and memory, and autism still differentiated losers from winners. The fighting system using male B. splendens thus provides a promising platform for investigating neurogenomic states of aggression in vertebrates.

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Link to bioRxiv paper:

http://biorxiv.org/cgi/content/short/2020.08.03.235234v1?rss=1

Authors: Manacorda, C. A., Gudesblat, G., Sutka, M., Alemano, S., Peluso, F., Oricchio, P., Baroli, I., asurmendi, s.

Abstract:

In this work the effects of TuMV infection on stomatal closure and water balance were studied in Arabidopsis. Thermal imaging analyses showed that TuMV-infected plants had consistently higher foliar temperature than mock treated controls. Non-destructive time-course experiments revealed that this differential phenotype was stable during both daytime and nighttime. This effect was due to reduced gas exchange in TuMV-infected plants, as observed through stomatal conductance and stomatal aperture assays in systemic leaves. Measurements of daily water consumption and initial dehydration rate further proved that TuMV infection reduced water loss. Salicylic acid (SA) and abscisic acid (ABA) contents were increased in TuMV-infected plants. In addition, the expression of ABI2, involved in ABA signaling, was enhanced, and ABCG40 (required for ABA transport into guard cells) was highly induced upon TuMV infection. Hypermorfic abi2-1 mutant plants, but no other ABA or SA biosynthetic, signaling or degradation mutants tested abolished both stomatal closure and low stomatal conductance phenotypes caused by TuMV. Therefore, not overall ABA levels, but localized differences in ABA import and perception in guard cells, are likely to be responsible for stomatal closure observed under TuMV infection. Plants simultaneously subjected to drought and viral stresses showed higher mortality rates than their mock-inoculated drought stressed counterparts, consistent with down-regulation of drought-responsive gene RD29A, both in short and long day conditions. Our findings indicate that in spite of stomatal closure triggered by TuMV, additional phenomena cause compromised drought tolerance of TuMV-infected Arabidopsis plants.

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