PaperPlayer biorxiv genetics

Link to bioRxiv paper:

http://biorxiv.org/cgi/content/short/2020.07.27.223560v1?rss=1

Authors: McClanahan, P. D., McCloskey, R. J., Ng, M., Raizen, D. M., Fang-Yen, C.

Abstract:

Ultracold preservation is widely used for storage of genetic stocks of Caenorhabditis elegans. Current cryopreservation protocols are vulnerable to refrigeration failures, which can result in the loss of stock viability due to damage during re-freezing. Here we present a method for preserving worms in a dehydrated and frozen form that retains viability after multiple freeze/thaw cycles. After dehydration in the presence of trehalose or glycerol, C. elegans stocks can be frozen and thawed multiple times while maintaining viability. Both stress-resistant dauer and non-dauer larvae survive desiccation and freezing. Survival after desiccation is dependent on the forkhead box transcription factor homologue DAF-16. Our technique is useful for storing stocks in a manner robust to freezer failures, and potentially for shipping strains between laboratories.

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Link to bioRxiv paper:

http://biorxiv.org/cgi/content/short/2020.07.29.225037v1?rss=1

Authors: Waneka, G., Vasquez, Y. M., Bennett, G. M., Sloan, D. B.

Abstract:

Compared to free-living bacteria, endosymbionts of sap-feeding insects have tiny and rapidly evolving genomes. Increased genetic drift, high mutation rates, and relaxed selection associated with host control of key cellular functions all likely contribute to genome decay. Phylogenetic comparisons have revealed massive variation in endosymbiont evolutionary rate, but such methods make it difficult to partition the effects of mutation vs. selection. For example, the ancestor of auchenorrhynchan insects contained two obligate endosymbionts, Sulcia and a betaproteobacterium (BetaSymb; called Nasuia in leafhoppers) that exhibit divergent rates of sequence evolution and different propensities for loss and replacement in the ensuing ~300 Ma. Here, we use the auchenorrhynchan leafhopper Macrosteles sp. nr. severini, which retains both of the ancestral endosymbionts, to test the hypothesis that differences in evolutionary rate are driven by differential mutagenesis. We used a high-fidelity technique known as duplex sequencing to measure and compare low-frequency variants in each endosymbiont. Our direct detection of de novo mutations reveals that the rapidly evolving endosymbiont (Nasuia) has a much higher frequency of single-nucleotide variants than the more stable endosymbiont (Sulcia) and a mutation spectrum that is even more AT-biased than implied by the 83.1% AT content of its genome. We show that indels are common in both endosymbionts but differ substantially in length and distribution around repetitive regions. Our results suggest that differences in long-term rates of sequence evolution in Sulcia vs. BetaSymb, and perhaps the contrasting degrees of stability of their relationships with the host, are driven by differences in mutagenesis.

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Link to bioRxiv paper:

http://biorxiv.org/cgi/content/short/2020.07.28.224998v1?rss=1

Authors: Froehlich, J., Uyar, B., Herzog, M., Theil, K., Glazar, P., Akalin, A., Rajewsky, N.

Abstract:

Understanding how regulatory sequences control gene expression is fundamental to explain how phenotypes arise in health and disease. Traditional reporter assays inform about function of individual regulatory elements, typically in isolation. However, regulatory elements must ultimately be understood by perturbing them within their genomic environment and developmental- or tissue-specific contexts. This is technically challenging; therefore, few regulatory elements have been characterized in vivo. Here, we used inducible Cas9 and multiplexed guide RNAs to create hundreds of mutations in enhancers/promoters and 3' UTRs of 16 genes in C. elegans. To quantify the consequences of mutations on expression, we developed a targeted RNA sequencing strategy across hundreds of mutant animals. We were also able to systematically and quantitatively assign fitness cost to mutations. Finally, we identified and characterized sequence elements that strongly regulate phenotypic traits. Our approach enables highly parallelized, functional analysis of regulatory sequences in vivo.

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Link to bioRxiv paper:

http://biorxiv.org/cgi/content/short/2020.07.29.222828v1?rss=1

Authors: Vona, B., Mazaheri, N., Lin, S.-J., Dunbar, L. A., Maroofian, R., Azaiez, H., Booth, K. T., Vitry, S., Rad, A., Varshney, P., Fowler, B., Alagramam, K. N., Murphy, D., Shariati, G., Sedaghat, A., Houlden, H., VijayKumar, S., Smith, R. J. H., Haaf, T., El-Amraoui, A., Bowl, M. R., Varshney, G. K., Galehdari, H.

Abstract:

Deafness, the most frequent sensory deficit in humans, is extremely heterogenous with hundreds of genes probably involved. Clinical and genetic analyses of an extended consanguineous family with pre-lingual, moderate-to-profound autosomal recessive sensorineural hearing loss, allowed us to identify CLRN2, encoding a tetraspan protein as a new deafness gene. Homozygosity mapping followed by exome sequencing identified a 15.2 Mb locus on chromosome 4p15.32p15.1 containing a missense pathogenic variant in CLRN2 (c.494C>A, NM_001079827.2) segregating with the disease. Using in vitro RNA splicing analysis, we show that the CLRN2 c.494C>A mutation leads to two events: 1) the substitution of a highly conserved threonine (uncharged amino acid) to lysine (charged amino acid) at position 165, p.(Thr165Lys), and 2) aberrant splicing, with the retention of intron 2 resulting in a stop codon after 26 additional amino acids, p.(Gly146Lysfs*26). Expression studies and phenotyping of newly produced zebrafish and mouse models deficient for clarin 2 further confirm that clarin 2, expressed in the inner ear hair cells, is essential for normal organization and maintenance of the auditory hair bundles, and for hearing function. Together, our findings identify CLRN2 as a new deafness gene, which will impact future diagnosis and treatment for deaf patients.

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Link to bioRxiv paper:

http://biorxiv.org/cgi/content/short/2020.07.28.225433v1?rss=1

Authors: Scarfo, M., Sciandra, C., Santovito, A.

Abstract:

Aging and longevity are complex processes controlled at different levels, including genetic level. We evaluated the association of seven drug and DNA-repair gene polymorphisms with longevity in an Italian cohort. A sample of 756 subjects aged 18-98 was genotyped for CYP1A1 exon 7 A>G, GSTT1 null, GSTM1 null, GSTP A>G, XRCC1 exon 6 C>T, XRCC1 exon 9 A> G and XPC exon 15 A>C gene polymorphisms. The association between the analyzed gene polymorphisms and longevity was evaluated by dividing the sample into three age groups: 10-50, 51-85, and 86-98. We observed a significant decrease in the frequency of the GSTT1 null, GSTP G and XPC C alleles in the oldest group with respect to the youngest one and with respect to 51-85 age group. We obtained the same results also subdividing the sample into 1-85 and 86-98 age groups. The general linear model analyses confirmed a significant decreasing trend of the above mentioned alleles with age. We hypothesized that these minor alleles, being important in the sensitivity against the development of different types of cancer, may reflect a reduced life-expectancy in carrier subjects and may explain their significantly lower frequency observed among subjects belonging to oldest age group.

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Link to bioRxiv paper:

http://biorxiv.org/cgi/content/short/2020.07.27.223552v1?rss=1

Authors: Diaz G., S., Leblanc, D. P., Gagne, R., Behan, N. A., Wong, A., Marchetti, F., MacFarlane, A. J.

Abstract:

Cancer incidence varies by tissue due to differences in environmental risk factor exposure, gene variant inheritance, and lifetime number of stem cell divisions in a tissue. Folate deficiency is associated with increased risk for colorectal cancer (CRC) and acute lymphocytic leukemia. Conversely, high folic acid (FA) intake has been associated with higher CRC risk. However, the mutagenic potential of FA intake in different tissues has not been characterized. Here we quantified mutations in folate-susceptible somatic tissues, namely bone marrow and colon, from the same MutaMouse mice and determined the FA-induced mutation profiles of both tissues using next generation sequencing. FA-induced mutagenesis was tissue- and dose-specific: FA deficiency increased mutant frequency (MF) in bone marrow while FA supplementation increased MF in colon. Analyses of mutation profiles suggested that FA interacted with mutagenic mechanisms that are unique to each tissue. These data illuminate potential mechanisms underpinning differences in susceptibility to FA-related cancers.

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Link to bioRxiv paper:

http://biorxiv.org/cgi/content/short/2020.07.27.222893v1?rss=1

Authors: Higgs, M. J., Hill, M. J., John, R. M., Isles, A. J.

Abstract:

Over 200 murine imprinted genes have currently been identified, many are expressed in the brain, and a subset have been associated with neurological and behavioural phenotypes. These studies have led to the suggestion that imprinted genes show enriched expression in certain key brain functions; however, this has never been formally tested. Here we use fifteen available single-cell RNA sequencing datasets to investigate imprinted gene over-representation in the brain, with the aim of identifying key hotspots of expression and, by extension, function. We establish that imprinted genes are indeed over-represented in the adult brain compared to other adult tissues, and in neurons specifically compared to other cell-types. Brain wide datasets allowed us to demonstrate enrichment of imprinted genes in the hypothalamus, ventral midbrain, pons and medulla. Finally, using scRNA-seq datasets focusing on these regions of enrichment, we were able to identify hypothalamic neuroendocrine populations and the monoaminergic hindbrain neurons as specific hotspots of imprinted gene expression. These analyses provide a robust assessment of the neural systems on which imprinted genes converge and in turn suggest the neuronal regulation of motivated behaviours such as feeding, parental behaviour and sleep as functional hotspots for imprinting.

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Link to bioRxiv paper:

http://biorxiv.org/cgi/content/short/2020.07.28.225730v1?rss=1

Authors: Marderstein, A. R., Davenport, E., Kulm, S., Van Hout, C. V., Elemento, O., Clark, A. G.

Abstract:

While thousands of loci have been associated with human phenotypes, the role of gene-environment (GxE) interactions in determining individual risk of human diseases remains unclear. This is partly due to the severe erosion of statistical power resulting from the massive number of statistical tests required to detect such interactions. Here, we focus on improving the power of GxE tests by developing a statistical framework for assessing quantitative trait loci (QTLs) associated with the trait means and/or trait variances. When applying this framework to body mass index (BMI), we find that GxE discovery and replication rates are significantly higher when prioritizing genetic variants associated with the variance of the phenotype (vQTLs) compared to assessing all genetic variants. Moreover, we find that vQTLs are enriched for associations with other non-BMI phenotypes having strong environmental influences, such as diabetes or ulcerative colitis. We show that GxE effects first identified in quantitative traits such as BMI can be used for GxE discovery in disease phenotypes such as diabetes. A clear conclusion is that strong GxE interactions mediate the genetic contribution to body weight and diabetes risk.

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Link to bioRxiv paper:

http://biorxiv.org/cgi/content/short/2020.07.27.223008v1?rss=1

Authors: Wang, Y., Hsu, A. Y., Walton, E. M., Syahirah, R., Wang, T., Zhou, W., Ding, C., Lemke, A. P., Tobin, D. M., Deng, Q.

Abstract:

Tissue-specific knockout techniques are widely applied in biological studies to probe the tissue-specific roles of specific genes in physiology, development, and disease. CRISPR/Cas9 is a widely used technology to perform fast and efficient genome editing in vitro and in vivo. Here, we report a robust CRISPR-based gateway system for tissue-specific gene inactivation in zebrafish. A transgenic fish line expressing Cas9 under the control of a neutrophil-restricted promoter was constructed. As proof of principle, we transiently disrupted rac2 or cdk2 in neutrophils using plasmids driving the expression of sgRNAs from U6 promoters. Loss of the rac2 or cdk2 gene in neutrophils resulted in significantly decreased cell motility, which could be restored by re-expressing Rac2 or Cdk2 in neutrophils in the corresponding knockout background. The subcellular location of Rac activation and actin structure and stress in the context of neutrophil migration was determined in both the wild-type and rac2 knockout neutrophils in vivo. In addition, we evaluated an alternative approach where the Cas9 protein is ubiquitously expressed while the sgRNA is processed by ribozymes and expressed in a neutrophil-restricted manner. Cell motility was also reduced upon rac2 sgRNA expression. Together, our work provides a potent tool that can be used to advance the utility of zebrafish in identification and characterization of gene functions in neutrophils.

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Link to bioRxiv paper:

http://biorxiv.org/cgi/content/short/2020.07.27.222364v1?rss=1

Authors: Li, Q., Cao, C., Perera, D., He, J., Chen, X., Azeem, F., Howe, A., Au, B., Yan, J., Long, Q.

Abstract:

Biological interactions are prevalent in the functioning organisms. Correspondingly, statistical geneticists developed various models to identify genetic interactions through genotype-phenotype association mapping. The current standard protocols in practice test single variants or single regions (that contain multiple local variants) sequentially along the genome, followed by functional annotations that involve various aspects including interactions. The testing of genetic interactions upfront is rare in practice due to the burden of testing a huge number of combinations, which lead to the multiple-test problem and the risk of overfitting. In this work, we developed interaction-integrated linear mixed model (ILMM), a novel model that integrates a priori knowledge into linear mixed models. ILMM enables statistical integration of genetic interactions upfront and overcomes the problems associated with combination searching.

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