PaperPlayer biorxiv genomics

Link to bioRxiv paper:

http://biorxiv.org/cgi/content/short/2020.07.23.217430v1?rss=1

Authors: Raghav, S., Ghosh, A., Turuk, J., Kumar, S., Jha, A., Madhulika, S., Priyadarshini, M., Biswas, V. K., Shyamli, P. S., Singh, B., Singh, N., Singh, D., Kiran, A., Smita, S., Sabat, J., Bhattacharya, D., Dash, R., Senapati, S., Beuria, T. K., Swain, R., Chattopadhyay, S., Syed, G. H., Dixit, A., Prasad, P., Pati, S., Parida, A.

Abstract:

COVID-19 that emerged as a global pandemic is caused by SARS-CoV-2 virus. The virus genome analysis during disease spread reveals about its evolution and transmission. We did whole genome sequencing of 225 clinical strains from the state of Odisha in eastern India using ARTIC protocol-based amplicon sequencing. Phylogenetic analysis identified the presence of all five reported clades 19A, 19B, 20A, 20B and 20C in the population. The analyses revealed two major routes for the introduction of the disease in India i.e. Europe and South-east Asia followed by local transmission. Interestingly, 19B clade was found to be much more prevalent in our sequenced genomes (17%) as compared to other genomes reported so far from India. The haplogroup analysis for clades showed evolution of 19A and 19B in parallel whereas the 20B and 20C appeared to evolve from 20A. Majority of the 19A and 19B clades were present in cases that migrated from Gujarat state in India suggesting it to be one of the major initial points of disease transmission in India during month of March and April. We found that with the time 20A and 20B clades evolved drastically that originated from central Europe. At the same time, it has been observed that 20A and 20B clades depicted selection of four common mutations i.e. 241 C>T (5UTR), P323L in RdRP, F942F in NSP3 and D614G in the spike protein. We found an increase in the concordance of G614 mutation evolution with the viral load in clinical samples as evident from decreased Ct value of spike and Orf1ab gene in qPCR. Molecular modelling and docking analysis identified that D614G mutation enhanced interaction of spike with TMPRSS2 protease, which could impact the shedding of S1 domain and infectivity of the virus in host cells.

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Link to bioRxiv paper:

http://biorxiv.org/cgi/content/short/2020.07.20.212100v1?rss=1

Authors: Ashton, J. M., Rehrauer, H., Myers, J., Myers, J., Zanche, M., Balys, M., Foox, J., Mason, C. E., Steen, R., Kuentzel, M., Aquino, C., Garcia-Reyero, N., Chittur, S. V.

Abstract:

Single-cell RNA sequencing (scRNA-seq) offers great new opportunities for increasing our understanding of complex biological processes. In particular, development of an accurate Human Cell Atlas is largely dependent on the rapidly advancing technologies and molecular chemistries employed in scRNA-seq. These advances have already allowed an increase in throughput for scRNA-seq from 96 to 80,000 cells on a single instrument run by capturing cells within nano-liter droplets. While this increase in throughput is critical for many experimental questions, a thorough comparison between microfluidic-, plate-, and droplet-based technologies or between multiple available platforms utilizing these technologies is largely lacking. Here, we report scRNA-seq data from SUM149PT cells treated with the histone deacetylase inhibitor TSA vs. untreated controls across several scRNA-seq platforms (Fluidigm C1, WaferGen iCell8, 10X Genomics Chromium Controller, and Illumina/BioRad ddSEQ). The primary goal of this project was to demonstrate RNA sequencing (RNA-seq) methods for profiling the ultra-low amounts of RNA present in individual cells, and this report discusses the results of the study as well as technical challenges/lesson learned and present general guidelines for best practices in sample preparation and analysis.

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Link to bioRxiv paper:

http://biorxiv.org/cgi/content/short/2020.07.21.213801v1?rss=1

Authors: Medini, H., Cohen, T., Mishmar, D.

Abstract:

Mitochondrial gene expression is pivotal to cell metabolism. Nevertheless, it is unknown whether it diverges within a given cell type. Here, we analysed single-cell RNA-seq experiments from [~]4600 human pancreatic alpha and beta cells, as well as [~]900 mouse beta cells. Cluster analysis revealed two distinct human beta cells populations, which diverged by mitochondrial (mtDNA) and nuclear DNA (nDNA)-encoded oxidative phosphorylation (OXPHOS) gene expression in healthy and diabetic individuals, and in newborn but not in adult mice. Insulin gene expression was elevated in beta cells with higher mtDNA gene expression in humans and in young mice. Such human beta cell populations also diverged in mt-RNA mutational repertoire, and in their selective signature, thus implying the existence of two previously overlooked distinct and conserved beta cell populations. While applying our approach to alpha cells, two sub-populations of cells were identified which diverged in mtDNA gene expression, yet these cellular populations did not consistently diverge in nDNA OXPHOS genes expression, nor did they correlate with the expression of glucagon, the hallmark of alpha cells. Thus, pancreatic beta cells within an individual are divided into distinct groups with unique metabolic-mitochondrial signature.

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Link to bioRxiv paper:

http://biorxiv.org/cgi/content/short/2020.07.21.213975v1?rss=1

Authors: Tvedte, E. S., Gasser, M., Sparklin, B. C., Michalski, J., Zhao, X., Bromley, R., Tallon, L. J., Sadzewicz, L., Rasko, D. A., Dunning Hotopp, J. C.

Abstract:

BackgroundThe newest generation of DNA sequencing technology is highlighted by the ability to sequence reads hundreds of kilobases in length, and the increased availability of long read data has democratized the genome sequencing and assembly process. PacBio and Oxford Nanopore Technologies (ONT) have pioneered competitive long read platforms, with more recent work focused on improving sequencing throughput and per-base accuracy. Released in 2019, the PacBio Sequel II platform advertises substantial enhancements over previous PacBio systems.

ResultsWe used whole-genome sequencing data produced by two PacBio platforms (Sequel II and RS II) and two ONT protocols (Rapid Sequencing and Ligation Sequencing) to compare assemblies of the bacteria Escherichia coli and the fruit fly Drosophila ananassae. Sequel II assemblies had higher contiguity and consensus accuracy relative to other methods, even after accounting for differences in sequencing throughput. ONT RAPID libraries had the fewest chimeric reads in addition to superior quantification of E. coli plasmids versus ligation-based libraries. The quality of assemblies can be enhanced by adopting hybrid approaches using Illumina libraries for bacterial genome assemblies or combined ONT and Sequel II libraries for eukaryotic genome assemblies. Genome-wide DNA methylation could be detected using both technologies, however ONT libraries enabled the identification of a broader range of known E. coli methyltransferase recognition motifs in addition to undocumented D. ananassae motifs.

ConclusionsThe ideal choice of long read technology may depend on several factors including the question or hypothesis under examination. No single technology outperformed others in all metrics examined.

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Link to bioRxiv paper:

http://biorxiv.org/cgi/content/short/2020.07.22.216499v1?rss=1

Authors: Galka-Marciniak, P., Urbanek-Trzeciak, M. O., Nawrocka, P. M., Kozlowski, P.

Abstract:

It is a well-known and intensively studied phenomenon that the levels of many miRNAs are differentiated in cancer. miRNA biogenesis and functional expression are complex processes orchestrated by many proteins cumulatively called miRNA biogenesis proteins. To characterize cancer somatic mutations in the miRNA biogenesis genes and investigate their potential impact on the levels of miRNAs, we analyzed whole-exome sequencing datasets of over 10,000 cancer/normal sample pairs deposited within the TCGA repository. We identified and characterized over 3,600 somatic mutations in 29 miRNA biogenesis genes and showed that some of the genes are overmutated in specific cancers and/or have recurrent hotspot mutations (e.g., SMAD4 in PAAD, COAD, and READ; DICER1 in UCEC; PRKRA in OV; and LIN28B in SKCM). We identified a list of miRNAs whose level is affected by particular types of mutations in either SMAD4, SMAD2, or DICER1 and showed that hotspot mutations in the RNase domains in DICER1 not only decrease the level of 5p-miRNAs but also increase the level of 3p-miRNAs, including many well-known cancer-related miRNAs. We also showed an association of the mutations with patient survival. Eventually, we created an atlas/compendium of miRNA biogenesis alterations providing a useful resource for different aspects of biomedical research.

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Link to bioRxiv paper:

http://biorxiv.org/cgi/content/short/2020.07.21.211367v1?rss=1

Authors: Iha, C., Varela, J. A., Avila, V., Jackson, C. J., Bogaert, K. A., Chen, Y., Judd, L. M., Wick, R. R., Holt, K. E., Pasella, M. M., Ricci, F., Repetti, S. I., Medina, M., Marcelino, V. R., Chan, C. X., Verbruggen, H.

Abstract:

The green alga Ostreobium is an important coral holobiont member, playing key roles in skeletal decalcification and providing photosynthate to bleached corals that have lost their dinoflagellate endosymbionts. Ostreobium lives in the corals skeleton, a low-light environment with variable pH and O2 availability. We present the Ostreobium nuclear genome and a metatranscriptomic analysis of healthy and bleached corals to improve our understanding of Ostreobiums adaptations to its extreme environment and its roles as a coral holobiont member. The Ostreobium genome has 11,735 predicted protein-coding genes and shows adaptations for life in low and variable light conditions and other stressors in the endolithic environment. It presents an unusually rich repertoire of light harvesting complex proteins, lacks many genes for photoprotection and photoreceptors, and has a large arsenal of genes for oxidative stress response. An expansion of extracellular peptidases suggests that Ostreobium may supplement its energy needs and compensate for its lysine auxotrophy by feeding on the organic skeletal matrix, and a diverse set of fermentation pathways allow it to live in the anoxic skeleton at night. Ostreobium depends on other holobiont members for biotin and vitamin B12, and our metatranscriptomes identify potential bacterial sources. Metatranscriptomes showed Ostreobium becoming a dominant agent of photosynthesis in bleached corals and provided evidence for variable responses among coral samples and different Ostreobium genotypes. Our work provides a comprehensive understanding of the adaptations of Ostreobium to its extreme environment and an important genomic resource to improve our comprehension of coral holobiont resilience, bleaching and recovery.

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Link to bioRxiv paper:

http://biorxiv.org/cgi/content/short/2020.07.22.212464v1?rss=1

Authors: Fellers, J. P., Sakthikumar, S., He, F., McRell, K., Bakkeren, G., Cuomo, C. A., Kolmer, J. A.

Abstract:

BackgroundThe wheat leaf rust fungus, Puccinia triticina Erikss. is a worldwide pathogen of tetraploid durum and hexaploid wheat. Many races of P. triticina differ for virulence to specific leaf rust resistance genes and are found in most wheat-growing regions of the world. Wheat cultivars with effective leaf rust resistance exert selection pressure on P. triticina populations for virulent race types. The objectives of this study were to examine whole-genome sequence data of 121 P. triticina isolates and to gain insight into race evolution. The collection included isolates comprising many different race phenotypes collected worldwide from common wheat in the U.S. and the European Union together with isolates from durum wheat. One isolate from the wild wheat relative Aegilops speltoides, and two from Ae. cylindrica were also included for comparison.

ResultsBased on 121,907 variants identified relative to the reference race 1-1 genome, the isolates were clustered into 11 major lineages with 100% bootstrap support. The isolates were also grouped based on variation in approximately 1400 secreted resistance interactor candidate proteins. In gene-coding regions, all groups had high ratios of non-synonymous to synonymous mutations and nonsense to readthrough mutations.

ConclusionsBased on total variation or variation in the secreted protein genes, isolates grouped the same indicating that variants were distributed across the entire genome. Our results suggest that recurrent mutation and selection play a major role in differentiation within the clonal lineages.

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Link to bioRxiv paper:

http://biorxiv.org/cgi/content/short/2020.07.21.215038v1?rss=1

Authors: Zapata, L., Caravagna, G., Williams, M., Lakatos, E., Abdul-Jabbar, K., Werner, B., Graham, T. A., Sottoriva, A.

Abstract:

Immunoediting is a major force during cancer evolution that selects for clones with low immunogenicity (adaptation), or clones with mechanisms of immune evasion (escape). However, quantifying immunogenicity in the cancer genome and how the tumour-immune coevolutionary dynamics impact patient outcomes remain unexplored. Here we show that the ratio of nonsynonymous to synonymous mutations (dN/dS) in the immunopeptidome quantifies tumor immunogenicity and differentiates between adaptation and escape. We analysed 8,543 primary tumors from TCGA and validated immune dN/dS as a measure of selection associated with immune infiltration in immune-adapted tumours. In a cohort of 308 metastatic patients that received immunotherapy, pre-treatment lesions in non-responders showed increased immune selection (dN/dS<1), whereas responders did not and instead harbour a higher proportion of genetic escape mechanisms. Ultimately, these findings highlight the potential of evolutionary genomic measures to predict clinical response to immunotherapy.

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Link to bioRxiv paper:

http://biorxiv.org/cgi/content/short/2020.07.22.215749v1?rss=1

Authors: Rousseau-Gueutin, M., Belser, C., Da Silva, C., Richard, G., Istace, B., Cruaud, C., Falentin, C., Boideau, F., Boutte, J., Delourme, R., Deniot, G., Engelen, S., Ferreira de Carvalho, J., Lemainque, A., Maillet, L., Morice, J., Wincker, P., Denoeud, F., Chevre, A.-M., Aury, J.-M.

Abstract:

BackgroundThe combination of long-reads and long-range information to produce genome assemblies is now accepted as a common standard. This strategy not only allow to access the gene catalogue of a given species but also reveals the architecture and organisation of chromosomes, including complex regions like telomeres and centromeres. The Brassica genus is not exempt and many assemblies based on long reads are now available. The reference genome for Brassica napus, Darmor-bzh, which was published in 2014, has been produced using short-reads and its contiguity was extremely low if compared to current assemblies of the Brassica genus.

FindingsHere, we report the new long-reads assembly of Darmor-bzh genome (Brassica napus) generated by combining long-reads sequencing data, optical and genetic maps. Using the PromethION device and six flowcells, we generated about 16M long-reads representing 93X coverage and more importantly 6X with reads longer than 100Kb. This ultralong-reads dataset allows us to generate one of the most contiguous and complete assembly of a Brassica genome to date (contigs N50 > 10Mb). In addition, we exploited all the advantages of the nanopore technology to detect modified bases and sequence transcriptomic data using direct RNA to annotate the genome and focus on resistance genes.

ConclusionUsing these cutting edge technologies, and in particular by relying on all the advantages of the nanopore technology, we provide the most contiguous Brassica napus assembly, a resource that will be valuable for the Brassica community for crop improvement and will facilitate the rapid selection of agronomically important traits.

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Link to bioRxiv paper:

http://biorxiv.org/cgi/content/short/2020.07.22.215582v1?rss=1

Authors: Smith, J., Lipkin, E., Soller, M., Fulton, J. E., Burt, D. W.

Abstract:

Mareks Disease (MD) represents a significant global economic and animal welfare issue. Mareks Disease Virus (MDV) is a highly contagious oncogenic and highly immune-suppressive alpha-herpes virus, which infects chickens, causing neurological effects and tumour formation. Though partially controlled by vaccination, MD continues to have a profound impact on animal health and on the poultry industry. Genetic selection provides an alternative and complementary method to vaccination. However, even after years of study, the genetic mechanisms underlying resistance to MDV remain poorly understood. The MHC is known to play a role in disease resistance, along with a handful of other. In this study, one of the largest to date, we used a multi-facetted approach to identify QTL regions (QTLR) influencing resistance to MDV, including an F6 population from a full-sib advanced intercross line (FSIL) between two elite commercial layer lines differing in resistance to MDV, RNA-seq information from virus challenged chicks, and genome wide association study (GWAS) from multiple commercial lines. Candidate genomic elements residing in the QTLR were further tested for association with offspring mortality in the face of MDV challenge in 8 pure lines of elite egg-layer birds. Thirty-eight QTLR were found on 19 chicken chromosomes. Candidate genes, miRNAs, lncRNAs and potentially functional mutations were identified in these regions. Association tests were carried out in 26 of the QTLR, using 8 pure lines of elite egg-layer birds. Numerous candidate genomic elements were strongly associated with MD resistance. Genomic regions significantly associated with resistance to MDV were mapped, and candidate genes identified. Various QTLR elements were shown to have strong genetic association with resistance. These results provide a large number of significant targets for mitigating the effects of MDV infection on both poultry health and the economy - whether by means of selective breeding, improved vaccine design or gene-editing technologies.

Author summaryMareks Disease has a huge impact on the global poultry industry in terms of both animal welfare and economic cost. For many years, researchers have sought to identify the genes underlying resistance to Mareks Disease Virus (MDV). However, this is a complex trait with each genetic locus having a small effect, so identifying causal genes and variants is no easy task. To date, it is known that the MHC confers differing susceptibility/resistance. A few other non-MHC genes have also been implicated in disease resistance, although based on experimental inbred lines and not representing real world commercial poultry. Using an F6 intercross population of birds with differences in MDV survival, we have identified many regions of the genome involved in resistance and highlighted candidate genes, miRNAs and lncRNA. Access to DNA from phenotyped birds spanning 15 generations of 8 elite commercial lines has provided a unique opportunity for us to show genetic association of markers in these transcripts with MDV survival. This genetic study, the largest to date, provides novel targets for mitigation of Mareks Disease within the poultry industry. This could be through selective breeding strategies, improved vaccine design or future gene editing technologies.

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