PaperPlayer biorxiv molecular biology

Link to bioRxiv paper:

http://biorxiv.org/cgi/content/short/2020.07.27.223032v1?rss=1

Authors: Pajak, J., Dill, E., White, M. A., Kelch, B. A., Jardine, P., Arya, G., Morais, M. C.

Abstract:

Double-stranded DNA viruses package their genomes into pre-assembled protein capsids using virally-encoded ATPase ring motors. While several structures of isolated monomers (subunits) from these motors have been determined, they provide little insight into how subunits within a functional ring coordinate their activities to efficiently generate force and translocate DNA. Here we describe the first atomic-resolution structure of a functional ring form of a viral DNA packaging motor and characterize its atomic-level dynamics via long timescale molecular dynamics simulations. Crystal structures of the pentameric ATPase ring from bacteriophage ascc{varphi}28 show that each subunit consists of a canonical N-terminal ASCE ATPase domain connected to a 'vestigial' nuclease domain by a small lid subdomain. The lid subdomain closes over the ATPase active site and engages in extensive interactions with a neighboring subunit such that several important catalytic residues are positioned to function in trans. The pore of the ring is lined with several positively charged residues that can interact with DNA. Simulations of the ATPase ring in various nucleotide- and DNA-bound states provide information about how the motor coordinates sequential nucleotide binding, hydrolysis, and exchange around the ring. Simulations also predict that the ring adopts a helical structure to track DNA, consistent with recent cryo-EM reconstruction of the {varphi}28 packaging ATPase. Based on these results, an atomistic model of viral DNA packaging is proposed wherein DNA translocation is powered by stepwise helical-to-planar ring transitions that are tightly coordinated by ATP binding, hydrolysis, and release.

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Link to bioRxiv paper:

http://biorxiv.org/cgi/content/short/2020.07.27.222760v1?rss=1

Authors: Mosig, R. A., Castaneda, A. N., Deslauriers, J. C., Frazier, L. P., He, K. L., Maghzian, N., Pokharel, A., Schrier, C. T., Zhu, L., Koike, N., Tyson, J. J., Green, C. B., Takahashi, J. S., Kojima, S.

Abstract:

Circadian transcriptome studies identified a novel transcript at the Period2 (Per2) locus, which we named Per2AS. Per2AS is a long non-coding RNA transcribed from the antisense strand of Per2, and is expressed rhythmically and anti-phasic to Per2 mRNA. Previously, we mathematically tested the hypothesis that Per2AS and Per2 mutually inhibit each other's expression by forming a double negative feedback loop, and found that Per2AS expands the oscillatory domain. In this study, we have experimentally tested this prediction by perturbing the expression of Per2AS in mouse fibroblasts. We found that Per2AS represses Per2 pre-transcriptionally in cis and regulates the amplitude of the circadian clock, but not period or phase. Unexpectedly, we also found that Per2 positively regulates Per2AS post-transcriptionally, indicating that Per2AS and Per2 form a single negative feedback loop. Because knock-down of Per2 does not recapitulate the phenotypes of Per2AS perturbation and Per2AS also activates Bmal1 in trans, we propose that Per2AS regulates the amplitude of the circadian clock without producing a protein by rewiring the molecular clock circuit.

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Link to bioRxiv paper:

http://biorxiv.org/cgi/content/short/2020.07.27.221887v1?rss=1

Authors: Ling, W.-L., Su, C. T.-T., Lua, W.-H., Poh, J.-J., Ng, Y.-L., Wipat, A., Gan, S. K.

Abstract:

Boosting the production of recombinant therapeutic antibodies is crucial in both academic and industry settings. In this work, we investigated the usage of varying signal peptides by antibody genes and their roles in recombinant transient production. Comparing myeloma and the native signal peptides of both heavy and light chains in 168 antibody permutation variants, we performed a systematic analysis, finding amino acids counts to be involved in antibody production to construct a model for predicting co-transfection transient recombinant antibody production rates using the HEK293 system. The findings also provide insights into the usage of the large repertoire of antibody signal peptides.

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Link to bioRxiv paper:

http://biorxiv.org/cgi/content/short/2020.07.24.220756v1?rss=1

Authors: Maksimov, P., Bergmann, H., Wassermann, M., Romig, T., Gottstein, B., Casulli, A., Conraths, F. J.

Abstract:

Infections with eggs of Echinococcus granulosus sensu lato (s.l.) can cause cystic echinococcosis in intermediate host animals and humans. Upon ingestion of viable eggs, oncospheres hatch from the eggs and subsequently develop into fluid-filled larval cysts, most frequently in the liver or the lungs. The slowly growing cysts progressively interfere with organ function. The risk of infection is determined by the host range of the parasite, its pathogenicity and other epidemiologically relevant parameters, which differ significantly among the five species within the E. granulosus s.l. complex. It is therefore essential to diagnose the correct species within E. granulosus s.l. to help understand specific disease epidemiology and to facilitate effective implementation of control measures. For this purpose, simple, fast and cost-effective typing techniques are needed. We developed quantitative Real-Time Polymerase Chain Reactions (qPCRs) and corresponding sequence-specific hydrolysis DNA probes to target polymorphic regions in the mitochondrial genome of E. granulosus s.l.. In a single-step typing approach, we distinguished E. granulosus s.l. members in four epidemiologically relevant subgroups. These were E. granulosus sensu stricto (G1, G3), E. equinus (G4), E. ortleppi (G5) and the E. canadensis cluster (G6 to G8 and G10). The technique also allowed identification and differentiation of these species from other Echinococcus or Taenia taxa for samples isolated from cysts or faeces. Single-step genotyping techniques for the molecular diagnosis of Echinococcus spp. by qPCRs may not only improve diagnostic performance, but also our knowledge on the epidemiology of the parasites and help controlling the various agents of cystic echinococcosis.

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Link to bioRxiv paper:

http://biorxiv.org/cgi/content/short/2020.07.24.220558v1?rss=1

Authors: Phillips, J. B., Ardell, D. H.

Abstract:

The evolution of tRNA multi-gene families remains poorly understood, exhibiting unusual phenomena such as functional conversions of tRNA genes through anticodon shift substitutions. We reannotated tRNA genes from twelve Drosophila species, incorporating previously identified ortholog sets to compare substitution rates across tRNA bodies at single-site resolution. All rapidly evolving sites fell in the same metal-ion-binding pocket at the interface of the two tRNA stacked helical domains. We applied our tRNA Structure-Function Mapper (tSFM) method independently to each Drosophila species and one outgroup species Musca domestica. We found that, although predicted tRNA structure-function maps are generally highly conserved in flies, one tRNA Class-Informative Feature (CIF) within the rapidly-evolving ion-binding pocket --- Cytosine 17 (C17), ancestrally informative for lysylation identity --- independently gained asparaginylation identity and substituted in parallel across tRNAAsn paralogs at least three times in evolution of the genus. In D. melanogaster, approximately 15 tRNALys and tRNAAsn genes co-occur in heterologous clusters, suggesting that both heterologous gene conversion and structural similarities of the closely related asparaginyl-tRNA synthetase (AsnRS) and lysyl-tRNA synthetase (LysRS) proteins may have played a role in these changes. A previously identified Asn-to-Lys anticodon shift substitution in D. ananassae may have arisen to compensate for the convergent and parallel gains of C17 in tRNAAsn paralogs of the D. ananassae lineage. Our results underscore the functional and evolutionary relevance of our tRNA structure-function map predictions and illuminate multiple genomic and structural factors contributing to rapid, parallel and compensatory evolution of tRNA multi-gene families.

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Link to bioRxiv paper:

http://biorxiv.org/cgi/content/short/2020.07.27.223420v1?rss=1

Authors: Ashoti, A., Limone, F., van Kranenburg, M., Alemany, A., Baak, M., Vivie, J., Piccioni, F., Creyghton, M., Eggan, K., Geijsen, N.

Abstract:

Facioscapulohumeral muscular dystrophy (FHSD), a fundamentally complex muscle disorder that thus far remains untreatable. As the name implies, FSHD starts in the muscles of the face and shoulder gridle. The main perturbator of the disease is the pioneer transcription factor DUX4, which is misexpressed in affected tissues due to a failure in epigenetic repressive mechanisms. In pursuit of unraveling the underlying mechanism of FSHD and finding potential therapeutic targets or treatment options, we performed an exhaustive genome-wide CRISPR/Cas9 phenotypic rescue screen to identify modulators of DUX4 cytotoxicity. We found no key effectors other than DUX4 itself, suggesting treatment efforts in FSHD should be directed towards its direct modulation. The screen did however reveal some rare and unexpected Cas9-induced genomic events, that may provide important considerations for planning future CRISPR/Cas9 knock-out screens.

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Link to bioRxiv paper:

http://biorxiv.org/cgi/content/short/2020.07.27.190561v1?rss=1

Authors: Moon, J., Park, J. H., Choi, Y., Lim, C.-W., Park, J.-M., Yu, S.-H., Kim, Y., Han, H. J., Kim, C.-H., Song, Y.-S., Kim, C.

Abstract:

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) causes coronavirus 2019 (COVID-19). No treatment is available. Micro-RNAs (miRNAs) in mesenchymal stem cell-derived extracellular vesicles (MSC-EVs) are potential novel therapeutic agents because of their ability to regulate gene expression by inhibiting mRNA. Thus, they may degrade the RNA genome of SARS-CoV-2. EVs can transfer miRNAs to recipient cells and regulate conditions within them. MSC-EVs harbor major therapeutic miRNAs that play important roles in the biological functions of virus-infected host cells. Here, we examined their potential impact on viral and immune responses. MSC-EVs contained 18 miRNAs predicted to interact directly with the 3' UTR of SARS-CoV-2. These EVs suppressed SARS-CoV-2 replication in Vero E6 cells. In addition, five major miRNAs suppressed virus activity in a luciferase reporter assay by binding the 3' UTR. MSC-EVs showed strong regenerative effects and potent anti-inflammatory activity which may prevent lethal cytokine storms. We confirmed that EVs regulated inflammatory responses by several cell types, including human brain cells that express the viral receptor ACE2, suggesting that the brain may be targeted by SARS-CoV-2. miRNAs in MSC-EVs have several advantages as therapeutic agents against SARS-CoV-2: 1) they bind specifically to the viral 3' UTR, and are thus unlikely to have side effects; 2) because the 3' UTR is highly conserved and rarely mutates, MSC-EV miRNAs could be used against novel variants arising during viral replication; and 3) unique cargoes carried by MSC-EVs can have diverse effects, such as regenerating damaged tissue and regulating immunity.

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Link to bioRxiv paper:

http://biorxiv.org/cgi/content/short/2020.07.27.220954v1?rss=1

Authors: Liu, X., Lou, L., Zhou, L.

Abstract:

Background: Coronavirus Disease 2019 (COVID-19) caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has spread around the world. Developing cardiac injury is a common condition in COVID-19 patients, but the pathogenesis remains unclear. Methods: The RNA-Seq dataset (GES150392) compared expression profiling of mock human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) and SARS-cov-2-infected hiPSC-CMs were obtained from Gene Expression Omnibus (GEO). We identified the differentially expressed genes (DEGs) between those two groups. Through gene set enrichment analysis (GSEA), Gene Ontology (GO) analysis, Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis, and CLINVAR human diseases analysis to identify the main effect of SARS-CoV-2 on cardiomyocytes. A protein-protein interaction (PPI) network was constructed to visualize interactions and functions of the hub genes. Results: A total of 1554 DEGs were identified (726 upregulated genes and 828 downregulated genes). Gene enrichment analysis shown that SARS-CoV-2 activate immuno-inflammatory responses via multiple signal pathways, including TNF, IL6-JAK-STAT3, IL2-STAT5, NF-{kappa}B, IL17, and Toll-like receptor signaling pathway in hiPSC-CMs. Whereas, the muscle contraction, cellular respiration and cell cycle of hiPSC-CMs were inhibited by SARS-CoV-2. CLINVAR human diseases analysis shown SRAS-Cov-2 infection was associated with myocardial infarction, cardiomyopathy and Limb-girdle muscular dystrophy. 15 hub genes were identified based on PPI network. Function analysis revealed that 11 upregulated hub genes were mainly enriched in cytokine activity, chemokine activity, Inflammatory response, leukocyte chemotaxis, and lipopolysaccharide-mediated signaling pathway. Furthermore, 4 downregulated hub genes were related to cell cycle regulation. Conclusion: The present study elucidates that the SARS-CoV-2 infection induced a strong defensive response in cardiomyocyte, leading to excessive immune inflammation, cell hypoxia, functional contractility reduction and apoptosis, ultimately result in myocardial injury.

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Link to bioRxiv paper:

http://biorxiv.org/cgi/content/short/2020.07.24.215095v1?rss=1

Authors: Poddar, A., Azam, M. S., Kayikcioglu, T., Bobrovskyy, M., Zhang, J., Ma, X., Labhsetwar, P., Fei, J., Singh, D., Luthey-Schulten, Z., Vanderpool, C. K., Ha, T.

Abstract:

Base-pairing interactions mediate intermolecular target recognition in many biological systems and applications, including DNA repair, CRISPR, microRNA, small RNA (sRNA) and antisense oligo therapies. Even a single base-pair mismatch can cause a substantial difference in biological activity but presently we do not yet know how the target search kinetics in vivo are influenced by single nucleotide level changes. Here, we used high-throughput sequencing to identify functionally relevant single point mutants of the bacterial sRNA, SgrS, and quantitative super-resolution microscopy to probe the mutational impact on the regulation of its primary target, ptsG mRNA. Our super-resolution imaging and analysis platform allowed us to further dissect mutational effects on SgrS lifetimes, and even subtle changes in the in vivo rates of target association, kon, and dissociation, koff. Mutations that disrupt Hfq binding and are distal to the mRNA annealing region still decreased kon and increased koff, providing an in vivo demonstration that Hfq directly facilitates sRNA-mRNA annealing. Single base-pair mismatches in the annealing region reduced kon by 24-31% and increased koff by 14-25%, extending the time it takes to find and destroy the target mRNA by about a third, depending on whether an AU or GC base-pair is disrupted. The effects of disrupting contiguous base-pairing are much more modest than that expected from thermodynamics, suggesting that Hfq also buffers base-pair disruptions.

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Link to bioRxiv paper:

http://biorxiv.org/cgi/content/short/2020.07.24.220640v1?rss=1

Authors: Bouska, M., Bai, H.

Abstract:

The spectrin cytoskeleton has been shown to be critical in diverse processes such as axon development and degeneration, myoblast fusion, and spermatogenesis. Spectrin can be modulated in a tissue specific manner through junctional protein complexes, however, it has not been shown that lncRNAs interact with and regulate spectrin. Here we provide evidence of a lncRNA that interacts with and {beta} Spectrin to regulate spermatogenesis and endosomal related activity in fat bodies of Drosophila. Protein-RNA and Protein-Protein biochemical analysis indicated the interaction between and {beta} Spectrin is modulated by the lncRNA CR45362. Immunocytochemistry revealed CR45362 is highly expressed in the basal testis while and {beta} Spectrin are clearly disrupted in this same region of CR45362 mutants. We genetically demonstrate -Spectrin and CR45362 deficiencies cause spermatid nuclear bundling defects with congruous changes of spectrin distribution and reduced Lysotracker staining in the fat body. Our data suggests lncRNA regulation of spectrin could provide cells with a repertoire of modulatory molecules to manipulate cell-type specific cytoskeletal and endosomal requirements.

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