Biology subjects

Postlethwait, J.

Publications and source records attributed to Postlethwait, J..

12 recordsLinked to original sources

bioRxiv2023

VPS13B is localized at the cis-trans Golgi complex interface and is a functional partner of FAM177A1

Mutations in VPS13B, a member of a protein family implicated in bulk lipid transport between adjacent membranes, cause Cohen syndrome. VPS13B is known to be concentrated in the Golgi complex, but its precise location within this organelle and thus the site(s) where it achieves lipid transport remains unclear. Here we show that VPS13B is localized at the interface between cis and trans Golgi sub-compartments and that Golgi complex re-formation after Brefeldin A (BFA) induced disruption is delayed in VPS13B KO cells. This delay is phenocopied by loss of FAM177A1, a Golgi complex protein of unknown function reported to be a VPS13B interactor and whose mutations also result in a developmental disorder. In zebrafish, the vps13b orthologue, not previously annotated in this organism, genetically interacts with fam177a1. Collectively, these findings raise the possibility that bulk lipid transport by VPS13B may play a role in expanding Golgi membranes and that VPS13B may be assisted in this function by FAM177A1.

cell biology↗

bioRxiv2023

Multi-genome comparisons reveal gain-and-loss evolution of the anti-Mullerian hormone receptor type 2 gene, an old master sex determining gene, in Percidae

The Percidae family comprises many fish species of major importance for aquaculture and fisheries. Based on three new chromosome-scale assemblies in Perca fluviatilis, Perca schrenkii and Sander vitreus along with additional percid fish reference genomes, we provide an evolutionary and comparative genomic analysis of their sex-determination systems. We explored the fate of a duplicated anti-Mullerian hormone receptor type-2 gene (amhr2bY), previously suggested to be the master sex determining (MSD) gene in P. flavescens. Phylogenetically related and structurally similar amhr2 duplications (amhr2b) were found in P. schrenkii and Sander lucioperca, potentially dating this duplication event to their last common ancestor around 19-27 Mya. In P. fluviatilis and S. vitreus, this amhr2b duplicate has been lost while it was subject to amplification in S. lucioperca. Analyses of the amhr2b locus in P. schrenkii suggest that this duplication could be also male-specific as it is in P. flavescens. In P. fluviatilis, a relatively small (100 kb) non-recombinant sex-determining region (SDR) was characterized on chromosome-18 using population-genomics approaches. This SDR is characterized by many male-specific single-nucleotide variants (SNVs) and no large duplication/insertion event, suggesting that P. fluviatilis has a male heterogametic sex determination system (XX/XY), generated by allelic diversification. This SDR contains six annotated genes, including three (c18h1orf198, hsdl1, tbc1d32) with higher expression in testis than ovary. Together, our results provide a new example of the highly dynamic sex chromosome turnover in teleosts and provide new genomic resources for Percidae, including sex-genotyping tools for all three known Perca species.

genomics↗

bioRxiv2023

Differential allelic representation (DAR) identifies candidate eQTLs and improves transcriptome analysis

In comparisons between mutant and wild-type genotypes, transcriptome analysis can reveal the direct impacts of a mutation, together with the homeostatic responses of the biological system. Recent studies have highlighted that, when homozygous mutations are studied in non-isogenic backgrounds, genes from the same chromosome as a mutation often appear over-represented among differentially expressed (DE) genes. One hypothesis suggests that DE genes chromosomally linked to a mutation may not reflect true biological responses to the mutation but, instead, result from differences in representation of expression quantitative trait loci (eQTLs) between sample groups selected on the basis of mutant or wild-type genotype. This is problematic when inclusion of spurious DE genes in a functional enrichment study results in incorrect inferences of mutation effect. Here we show that chromosomally co-located differentially expressed genes (CC-DEGs) can also be observed in analyses of dominant mutations in heterozygotes. We define a method and a metric to quantify, in RNA-sequencing data, localised differential allelic representation (DAR) between groups of samples subject to differential expression analysis. We show how the DAR metric can predict regions prone to eQTL-driven differential expression, and how it can improve functional enrichment analyses through gene exclusion or weighting of gene-level rankings. Advantageously, this improved ability to identify probable eQTLs also reveals examples of CC-DEGs that are likely to be functionally related to a mutant phenotype. This supports a long-standing prediction that selection for advantageous linkage disequilibrium influences chromosome evolution. By comparing the genomes of zebrafish (Danio rerio) and medaka (Oryzias latipes), a teleost with a conserved ancestral karyotype, we find possible examples of chromosomal aggregation of CC-DEGs during evolution of the zebrafish lineage. The DAR metric provides a solid foundation for addressing the eQTL issue in new and existing datasets because it relies solely on RNA-sequencing data.

genetics↗

bioRxiv2021

Evolution of the nitric oxide synthase family in vertebrates and novel insights in gill development

Nitric oxide (NO) is an ancestral key signaling molecule essential for life and has enormous versatility in biological systems, including cardiovascular homeostasis, neurotransmission, and immunity. Although our knowledge of nitric oxide synthases (Nos), the enzymes that synthesize NO in vivo, is substantial, the origin of a large and diversified repertoire of nos gene orthologs in fish with respect to tetrapods remains a puzzle. The recent identification of nos3 in the ray-finned fish spotted gar, which was considered lost in the ray-finned fish lineage, changed this perspective. This prompted us to explore nos gene evolution and expression in depth, surveying vertebrate species representing key evolutionary nodes. This study provides noteworthy findings: first, nos2 experienced several lineage-specific gene duplications and losses. Second, nos3 was found to be lost independently in two different teleost lineages, Elopomorpha and Clupeocephala. Third, the expression of at least one nos paralog in the gills of developing shark, bichir, sturgeon, and gar but not in arctic lamprey, suggest that nos expression in this organ likely arose in the last common ancestor of gnathostomes. These results provide a framework for continuing research on nos genes roles, highlighting subfunctionalization and reciprocal loss of function that occurred in different lineages during vertebrate genome duplications.

evolutionary biology↗

bioRxiv2021

Circulating miRNA diversity, origin and response to changing metabolic and reproductive states, new insights from the rainbow trout

Circulating miRNAs (c-miRNAs) are found in most, if not all, biological fluids and are becoming well established biomarkers of many human pathologies. The aim of the present study was to investigate the potential of c-miRNAs as biomarkers of reproductive and metabolic states in fish, a question that has received little attention. Plasma was collected throughout the reproductive cycle from rainbow trout females subjected to two different feeding levels to trigger contrasting metabolic states; ovarian fluid was sample at ovulation. Fluid samples were subjected to small RNA-seq analysis followed by quantitative PCR validation for a subset of promising c-miRNA biomarkers. A comprehensive miRNA repertoire, which was lacking in trout, was first established to allow subsequent analysis. We first showed that biological fluids miRNAomes are complex and encompass a high proportion of the overall species miRNAome. While sharing a high proportion of common miRNAs, plasma and ovarian fluid miRNAomes exhibited strong fluid-specific signatures. We further showed that the plasma miRNAome exhibited major significant changes depending on metabolic and reproductive state. We subsequently identified three (miR-1-1/2-3p, miR-133-a-1/2-3p and miR-206-3p) evolutionarily conserved muscle-specific miRNA that accumulate in the plasma in response to high feeding rates, making these myomiRs strong candidate biomarkers of active myogenesis. We also identified miR-202-5p as a candidate biomarker for reproductive success that could be used to predict ovulation and/or egg quality. These highly promising results reveal the high potential of c-miRNAs as physiologically relevant biomarkers and pave the way for the use of c-miRNAs for non-invasive phenotyping in various fish species.

genomics↗

bioRxiv2021

A complex genetic architecture in zebrafish relatives Danio quagga and D. kyathit underlies development of stripes and spots

Vertebrate pigmentation is a fundamentally important, multifaceted phenotype. Zebrafish, Danio rerio, has been a valuable model for understanding genetics and development of pigment pattern formation due to its genetic and experimental tractability, advantages that are shared across several Danio species having a striking array of pigment patterns. Here, we use the sister species D. quagga and D. kyathit, with stripes and spots, respectively, to understand how natural genetic variation impacts phenotypes at cellular and organismal levels. We first show that D. quagga and D. kyathit phenotypes resemble those of wild-type D. rerio and several single locus mutants of D. rerio, respectively, in a morphospace defined by pattern variation along dorsoventral and anteroposterior axes. We then identify differences in patterning at the cellular level between D. quagga and D. kyathit by repeated daily imaging during pattern development and quantitative comparisons of adult phenotypes, revealing that patterns are similar initially but diverge ontogenetically. To assess the genetic architecture of these differences, we employ reduced-representation sequencing of second-generation hybrids. Despite the similarity of D. quagga to D. rerio, and D. kyathit to some D. rerio mutants, our analyses reveal a complex genetic basis for differences between D. quagga and D. kyathit, with several quantitative trait loci contributing to variation in overall pattern and cellular phenotypes, epistatic interactions between loci, and abundant segregating variation within species. Our findings provide a window into the evolutionary genetics of pattern-forming mechanisms in Danio and highlight the complexity of differences that can arise even between sister species. Further studies of natural genetic diversity underlying pattern variation in D. quagga and D. kyathit should provide insights complementary to those from zebrafish mutant phenotypes and more distant species comparisons. Author SummaryPigment patterns of fishes are diverse and function in a wide range of behaviors. Common pattern themes include stripes and spots, exemplified by the closely related minnows Danio quagga and D. kyathit, respectively. We show that these patterns arise late in development owing to alterations in the development and arrangements of pigment cells. In the closely related model organism zebrafish (D. rerio) single genes can switch the pattern from stripes to spots. Yet, we show that pattern differences between D. quagga and D. kyathit have a more complex genetic basis, depending on multiple genes and interactions between these genes. Our findings illustrate the importance of characterizing naturally occuring genetic variants, in addition to laboratory induced mutations, for a more complete understanding of pigment pattern development and evolution.

developmental biology↗

bioRxiv2020

An intestinal cell type in zebrafish is the nexus for the SARS-CoV-2 receptor and the Renin-Angiotensin-Aldosterone System that contributes to COVID-19 comorbidities

People with underlying conditions, including hypertension, obesity, and diabetes, are especially susceptible to negative outcomes after infection with the coronavirus SARS-CoV-2. These COVID-19 comorbidities are exacerbated by the Renin-Angiotensin-Aldosterone System (RAAS), which normally protects from rapidly dropping blood pressure or dehydration via the peptide Angiotensin II (Ang II) produced by the enzyme Ace. The Ace paralog Ace2 degrades Ang II, thus counteracting its chronic effects. Ace2 is also the SARS-CoV-2 receptor. Ace, the coronavirus, and COVID-19 comorbidities all regulate Ace2, but we dont yet understand how. To exploit zebrafish (Danio rerio) as a disease model to understand mechanisms regulating the RAAS and its relationship to COVID-19 comorbidities, we must first identify zebrafish orthologs and co-orthologs of human RAAS genes, and second, understand where and when these genes are expressed in specific cells in zebrafish development. To achieve these goals, we conducted genomic analyses and investigated single cell transcriptomes. Results showed that most human RAAS genes have an ortholog in zebrafish and some have two or more co-orthologs. Results further identified a specific intestinal cell type in zebrafish larvae as the site of expression for key RAAS components, including Ace, Ace2, the coronavirus co-receptor Slc6a19, and the Angiotensin-related peptide cleaving enzymes Anpep and Enpep. Results also identified specific vascular cell subtypes as expressing Ang II receptors, apelin, and apelin receptor genes. These results identify specific genes and cell types to exploit zebrafish as a disease model for understanding the mechanisms leading to COVID-19 comorbidities. SUMMARY STATEMENTGenomic analyses identify zebrafish orthologs of the Renin-Angiotensin-Aldosterone System that contribute to COVID-19 comorbidities and single-cell transcriptomics show that they act in a specialized intestinal cell type.

developmental biology↗

bioRxiv2020

The rise and fall of the ancient northern pike master sex determining gene

Sexual reproduction is a ubiquitous basic feature of life and genetic sex determination is thus widespread, at least among eukaryotes. Understanding the remarkable diversity of sex determination mechanisms, however, is limited by the paucity of empirical studies. Here, we traced back the evolution of sex determination in an entire clade of vertebrates and uncovered that the northern pike (Esox lucius) master sex-determining gene initiated from a 65 to 90 million-year-old gene duplication and remained sex-linked on undifferentiated sex chromosomes for at least 56 million years. Contrasting with its ancient origin, we identified several independent species- or population-specific transitions of sex determination mechanisms in this lineage, including an unexpected complete and recent Y-chromosome loss in some North American northern pike populations. These findings highlight the diversity of the evolutionary fates of master sex-determining genes and raise the importance of careful considerations of population demographic history in sex determination studies. Our study also puts forward the hypothesis that occasional sex reversals and genetic bottlenecks provide a non-adaptive explanation for sex determination transitions.

evolutionary biology↗

bioRxiv2019

Sex chromosome and sex locus characterization in the goldfish, Carassius auratus

BackgroundGoldfish is an important model for various areas of research, including neural development and behavior and a species of significant importance in aquaculture, especially as an ornamental species. It has a male heterogametic (XX/XY) sex determination system that relies on both genetic and environmental factors, with high temperatures being able to produce female-to-male sex reversal. Little, however, is currently known on the molecular basis of genetic sex determination in this important cyprinid model. We used sequencing approaches to better characterize sex determination and sex-chromosomes in goldfish. ResultsOur results confirmed that sex determination in goldfish is a mix of environmental and genetic factors and that its sex determination system is male heterogametic (XX/XY). Using reduced representation (RAD-seq) and whole genome (pool-seq) approaches, we characterized sex-linked polymorphisms and developed male specific genetic markers. These male specific markers were used to distinguish sex-reversed XX neomales from XY males and to demonstrate that XX female-to-male sex reversal could even occur at a relatively low rearing temperature (18{degrees}C), for which sex reversal has been previously shown to be close to zero. We also characterized a relatively large non-recombining region ([~]11.7 Mb) on goldfish linkage group 22 (LG22) that contained a high-density of male-biased genetic polymorphisms. This large LG22 region harbors 373 genes, including a single candidate as a potential master sex gene, i.e., the anti-Mullerian hormone gene (amh). However, no sex-linked polymorphisms were detected in the goldfish amh gene or its 5 kb proximal promoter sequence. ConclusionsThese results show that goldfish have a relatively large sex locus on LG22, which is likely the goldfish Y chromosome. The presence of a few XX males even at low temperature also suggests that other environmental factors in addition to temperature could trigger female-to-male sex reversal. Finally, we also developed sex-linked genetic markers in goldfish, which will be important for future research on sex determination and aquaculture applications in this species.

genomics↗

bioRxiv2019

The roles of Amh in zebrafish gonad development and sex determination

Fetal mammalian testes secrete Amh (Anti-Mullerian hormone), which inhibits female reproductive tract (Mullerian duct) development. Amh also derives from mature mammalian ovarian follicles, which marks oocyte reserve and characterizes PCOS (polycystic ovarian syndrome). Zebrafish (Danio rerio) lacks Mullerian ducts and the Amh receptor gene amhr2 but, curiously, retains amh. To discover the roles of Amh in the absence of Mullerian ducts and the ancestral receptor gene, we made amh null alleles in zebrafish. Results showed that normal amh prevents female-biased sex ratios. Adult male amh mutants had enormous testes, half of which contained immature oocytes, demonstrating that Amh regulates male germ cell accumulation and inhibits oocyte development or survival. Mutant males formed sperm ducts and some produced a few offspring. Young female mutants laid a few fertile eggs, so they also had functional sex ducts. Older amh mutants accumulated non-vitellogenic follicles in exceedingly large but sterile ovaries, showing that Amh helps control ovarian follicle maturation and proliferation. RNA-seq data partitioned 21-day post-fertilization (dpf) juveniles into two groups that each contained mutant and wild type fish. Group21-1 up-regulated ovary genes compared to Group21-2, which were likely developing as males. By 35dpf, transcriptomes distinguished males from females and, within each sex, mutants from wild types. In adult mutants, ovaries greatly under-expressed granulosa and theca genes and testes under-expressed Leydig cell genes. These results show that ancestral Amh functions included development of the gonadal soma in ovaries and testes and regulation of gamete proliferation and maturation. A major gap in our understanding is the identity of the gene encoding a zebrafish Amh receptor; we show here that the loss of amhr2 is associated with the breakpoint of a chromosome rearrangement shared among cyprinid fishes.\n\nArticle SummaryAnti-Mullerian hormone (Amh) inhibits female reproductive duct development, signals oocyte reserve, and marks polycystic ovarian syndrome. Zebrafish lacks Mullerian ducts and the typical Amh receptor, questioning evolving roles of Amh. We made knockout mutations in zebrafish amh. Most mutants were female and the few males often had oocytes in their testes, showing that Amh promotes male development. Mutant reproductive ducts functioned, but testes were enormous and ovaries accumulated immature oocytes, showing that Amh regulates germ cell proliferation and maturation. Transcriptomics revealed that Amh controls development of steroid-producing gonad cells. Amh in zebrafish preserved ancestral roles despite losing Mullerian ducts and the Amh receptor.

genetics↗

bioRxiv2019

Identification of the master sex determining gene in Northern pike (Esox lucius) reveals restricted sex chromosome differentiation

Teleost fishes, thanks to their rapid evolution of sex determination mechanisms, provide remarkable opportunities to study the formation of sex chromosomes and the mechanisms driving the birth of new master sex determining (MSD) genes. However, the evolutionary interplay between the sex chromosomes and the MSD genes they harbor is rather unexplored. We characterized a male-specific duplicate of the anti-Mullerian hormone (amh) as the MSD gene in Northern Pike (Esox lucius), using genomic and expression evidences as well as by loss-of-function and gain-of-function experiments. Using RAD-Sequencing from a family panel, we identified Linkage Group (LG) 24 as the sex chromosome and positioned the sex locus in its sub-telomeric region. Furthermore, we demonstrated that this MSD originated from an ancient duplication of the autosomal amh gene, which was subsequently translocated to LG24. Using sex-specific pooled genome sequencing and a new male genome sequence assembled using Nanopore long reads, we also characterized the differentiation of the X and Y chromosomes, revealing a small male-specific insertion containing the MSD gene and a limited region with reduced recombination. Our study depicts an unexpected level of limited differentiation within a pair of sex chromosomes harboring an old MSD gene in a wild population of teleost fish, highlights the pivotal role of genes from the amh pathway in sex determination, as well as the importance of gene duplication as a mechanism driving the turnover of sex chromosomes in this clade. Author SummaryIn stark contrast to mammals and birds, teleosts have predominantly homomorphic sex chromosomes and display a high diversity of sex determining genes. Yet, population level knowledge of both the sex chromosome and the master sex determining gene is only available for the Japanese medaka, a model species. Here we identified and provided functional proofs of an old duplicate of anti-Mullerian hormone (Amh), a member of the Tgf-{beta} family, as the male master sex determining gene in the Northern pike, Esox lucius. We found that this duplicate, named amhby (Y-chromosome-specific anti-Mullerian hormone paralog b), was translocated to the sub-telomeric region of the new sex chromosome, and now amhby shows strong sequence divergence as well as substantial expression pattern differences from its autosomal paralog, amha. We assembled a male genome sequence using Nanopore long reads and identified a restricted region of differentiation within the sex chromosome pair in a wild population. Our results provide insight on the conserved players in sex determination pathways, the mechanisms of sex chromosome turnover, and the diversity of levels of differentiation between homomorphic sex chromosomes in teleosts.

evolutionary biology↗

bioRxiv2017

Evolution of gene expression after whole-genome duplication: new insights from the spotted gar genome

Whole genome duplications (WGD) are important evolutionary events. Our understanding of underlying mechanisms, including the evolution of duplicated genes after WGD, however remains incomplete. Teleost fish experienced a common WGD (teleost-specific genome duplication, or TGD) followed by a dramatic adaptive radiation leading to more than half of all vertebrate species. The analysis of gene expression patterns following TGD at the genome level has been limited by the lack of suitable genomic resources. The recent concomitant release of the genome sequence of spotted gar (a representative of holosteans, the closest lineage of teleosts that lacks the TGD) and the tissue-specific gene expression repertoires of over 20 holostean and teleostean fish species, including spotted gar, zebrafish and medaka (the PhyloFish project), offered a unique opportunity to study the evolution of gene expression following TGD in teleosts. We show that most TGD duplicates gained their current status (loss of one duplicate gene or retention of both duplicates) relatively rapidly after TGD (i.e. prior to the divergence of medaka and zebrafish lineages). The loss of one duplicate is the most common fate after TGD with a probability of approximately 80%. In addition, the fate of duplicate genes after TGD, including subfunctionalization, neofunctionalization, or retention of two similar copies occurred not only before, but also after the radiation of species tested, in consistency with a role of the TGD in speciation and/or evolution of gene function. Finally, we report novel cases of TGD ohnolog subfunctionalization and neofunctionalization that further illustrate the importance of these processes.

evolutionary biology↗