Systematic Analysis of Pleiotropy in C. elegans Early Embryogenesis

Pleiotropy refers to the phenomenon in which a single gene controls several distinct, and seemingly unrelated, phenotypic effects. We use C. elegans early embryogenesis as a model to conduct systematic studies of pleiotropy. We analyze high-throughput RNA interference (RNAi) data from C. elegans and identify “phenotypic signatures”, which are sets of cellular defects indicative of certain biological functions. By matching phenotypic profiles to our identified signatures, we assign genes with complex phenotypic profiles to multiple functional classes. Overall, we observe that pleiotropy occurs extensively among genes involved in early embryogenesis, and a small proportion of these genes are highly pleiotropic. We hypothesize that genes involved in early embryogenesis are organized into partially overlapping functional modules, and that pleiotropic genes represent “connectors” between these modules. In support of this hypothesis, we find that highly pleiotropic genes tend to reside in central positions in protein-protein interaction networks, suggesting that pleiotropic genes act as connecting points between different protein complexes or pathways

Systematic Analysis of Pleiotropy in C. elegans Early Embryogenesis

Pleiotropy refers to the phenomenon in which a single gene controls several distinct, and seemingly unrelated, phenotypic effects. We use C. elegans early embryogenesis as a model to conduct systematic studies of pleiotropy. We analyze high-throughput RNA interference (RNAi) data from C. elegans and identify “phenotypic signatures”, which are sets of cellular defects indicative of certain biological functions. By matching phenotypic profiles to our identified signatures, we assign genes with complex phenotypic profiles to multiple functional classes. Overall, we observe that pleiotropy occurs extensively among genes involved in early embryogenesis, and a small proportion of these genes are highly pleiotropic. We hypothesize that genes involved in early embryogenesis are organized into partially overlapping functional modules, and that pleiotropic genes represent “connectors” between these modules. In support of this hypothesis, we find that highly pleiotropic genes tend to reside in central positions in protein-protein interaction networks, suggesting that pleiotropic genes act as connecting points between different protein complexes or pathways

De Novo Inference of Systems-Level Mechanistic Models of Development from Live-Imaging-Based Phenotype Analysis

SummaryElucidation of complex phenotypes for mechanistic insights presents a significant challenge in systems biology. We report a strategy to automatically infer mechanistic models of cell fate differentiation based on live-imaging data. We use cell lineage tracing and combinations of tissue-specific marker expression to assay progenitor cell fate and detect fate changes upon genetic perturbation. Based on the cellular phenotypes, we further construct a model for how fate differentiation progresses in progenitor cells and predict cell-specific gene modules and cell-to-cell signaling events that regulate the series of fate choices. We validate our approach in C. elegans embryogenesis by perturbing 20 genes in over 300 embryos. The result not only recapitulates current knowledge but also provides insights into gene function and regulated fate choice, including an unexpected self-renewal. Our study provides a powerful approach for automated and quantitative interpretation of complex in vivo information

Ontogeny and phylogeny: molecular signatures of selection, constraint, and temporal pleiotropy in the development of Drosophila

<p>Abstract</p> <p>Background</p> <p>Karl Ernst Von Baer noted that species tend to show greater morphological divergence in later stages of development when compared to earlier stages. Darwin originally interpreted these observations via a selectionist framework, suggesting that divergence should be greatest during ontogenic stages in which organisms experienced varying 'conditions of existence' and opportunity for differential selection. Modern hypotheses have focused on the notion that genes and structures involved in early development will be under stronger purifying selection due to the deleterious pleiotropic effects of mutations propagating over the course of ontogeny, also known as the developmental constraint hypothesis.</p> <p>Results</p> <p>Using developmental stage-specific expressed sequence tag (EST) libraries, we tested the 2 hypotheses by comparing the rates of evolution of 7,180 genes obtained from 6 species of the <it>Drosophila melanogaster </it>group with respect to ontogeny, and sex and reproduction-related functions in gonadal tissues. Supporting morphological observations, we found evidence of a pattern of increasing mean evolutionary rate in genes that are expressed in subsequent stages of development. Furthermore, supporting expectations that early expressed genes are constrained in divergence, we found that embryo stage genes are involved in a higher mean number of interactions as compared to later stages. We noted that the accelerated divergence of genes in the adult stage is explained by those expressed specifically in the male gonads, whose divergence is driven by positive selection. In addition, accelerated gonadal gene divergence occurs only in the adult stage, suggesting that the effects of selection are observed primarily at the stages during which they are expected occur. Finally, we also found a significant correlation between temporal specificity of gene expression and evolutionary rate, supporting expectations that genes with ubiquitous expression are under stronger constraint.</p> <p>Conclusion</p> <p>Taken together, these results support both the developmental constraint hypothesis limiting the divergence of early expressed developmentally important genes, leading to a gradient of divergence rates over ontogeny (embryonic < larval/pupal < adult), as well as Darwin's 'selection opportunity' hypothesis leading to increased divergence in adults, particularly in the case of reproductive tissues. We suggest that a constraint early/opportunity late model best explains divergence over ontogeny.</p

Developmental constraints, innovations and robustness

During my PhD, I have been working on Evo-Devo patterns (especially the debate around the hourglass model) in transcriptomes, with an emphasis on adaptation. I have characterized patterns in model organisms in terms of constraints and especially in terms of positive selection. I found that the phylotypic stage (a stage in mid-embryonic development) is an evolutionary lockdown, with stronger purifying selection and less positive selection than other stages in terms of the evolution of protein sequences and of regulatory elements. To study the adaptive evolution of gene regulation during development, I have developed a machine leaning based in silico mutagenesis approach to detect positive selection on regulatory elements. In addition to transcriptome evolution, I have been working on the tension between precision and stochasticity of gene expression during development. More precisely, I have shown that expression noise follows an hourglass pattern, with lower noise at the phylotypic stage. This pattern can be explained by stronger histone modification mediated noise control at this stage. In addition, I propose that histone modifications contribute to mutational robustness in regulatory elements, and thus to conserved expression levels. These results provide insight into the role of robustness in the phenotypic and genetic patterns of evolutionary conservation in animal developmen

The hyperfunction theory: An emerging paradigm for the biology of aging

The process of senescence (aging) is predominantly determined by the action of wild-type genes. For most organisms, this does not reflect any adaptive function that senescence serves, but rather evolutionary effects of declining selection against genes with deleterious effects later in life. To understand aging requires an account of how evolutionary mechanisms give rise to pathogenic gene action and late-life disease, that integrates evolutionary (ultimate) and mechanistic (proximate) causes into a single explanation. A well-supported evolutionary explanation by G.C. Williams argues that senescence can evolve due to pleiotropic effects of alleles with antagonistic effects on fitness and late-life health (antagonistic pleiotropy, AP). What has remained unclear is how gene action gives rise to late-life disease pathophysiology. One ultimate-proximate account is T.B.L. Kirkwood's disposable soma theory. Based on the hypothesis that stochastic molecular damage causes senescence, this reasons that aging is coupled to reproductive fitness due to preferential investment of resources into reproduction, rather than somatic maintenance. An alternative and more recent ultimate-proximate theory argues that aging is largely caused by programmatic, developmental-type mechanisms. Here ideas about AP and programmatic aging are reviewed, particularly those of M.V. Blagosklonny (the hyperfunction theory) and J.P. de Magalhães (the developmental theory), and their capacity to make sense of diverse experimental findings is assessed

Landscape of pleiotropic proteins causing human disease: structural and system biology insights

Pleiotropy is the phenomenon by which the same gene can result in multiple phenotypes. Pleiotropic proteins are emerging as important contributors to rare and common disorders. Nevertheless , little is known on the mechanisms underlying pleiotropy and the characteris tic of pleiotropic proteins. We analysed disease - causing proteins reported in Uni P rot and observed that 12% are pleiotropic ( variants in the same protein cause more than one disease). Pleiotropic proteins were enriched in deleterious and rare variants , bu t not in common variants . Pleiotropic proteins were more likely to be involved in the pathogenesis of n eoplasms, neurological and circulatory diseases, and congenital malformations, whereas non - pleiotropic proteins in endocrine and metabolic disorders . Pleiotropic proteins were more essential and ha d a higher number of interacting partners compared to non -pleiotropic proteins. S ignificantly more pleiotropic than non - pleiotropic proteins contained at least one intrinsically long disordered region (p<0.001 ). Deleterious variants occurring in structurally disordered regions were more commonly found in pleiotropic, rather than non - pleiotropic proteins. 14 In conclusion, pleiotropic proteins are an important contributor to human disease. They represent a biologi cally different class of proteins compared to non - pleiotropic proteins and a better understanding of their characteristics and genetic variants, can greatly aid in the interpretation of genetic studies and drug design

Regeneration and the need for simpler model organisms

Journal ArticleThe problem of regeneration is fundamentally a problem of tissue homeostasis involving the replacement of cells lost to normal 18 wear and tear 19 (cell turnover), and/or injury. This attribute is of particular significance to organisms possessing relatively long lifespans, as maintenance of all body parts and their func- functional tional integration is essential for their survival. Because tissue replacement is broadly distributed among multicellular life-forms, and the molecules and mechanisms controlling cellular differentiation are considered ancient evolutionary inventions, it should be possible to gain key molecular insights about regenerative processes through the study of simpler animals. We have chosen to study and develop the freshwater planarian Schmidtea mediterranea as a model system because it is one of the simplest metazoans possessing tissue homeostasis and regeneration, and because it has become relatively easy to molecularly manipulate this organism. The developmental plasticity and longevity of S. mediterranea is in marked contrast to its better-characterized invertebrate cohorts: the fruitfly Drosophila melanogaster and the roundworm Caeno- Caenorhabditis rhabditis elegans elegans, both of which have short lifespans and are poor at regenerating tissues. Therefore, plan- planarians arians present us with new, experimentally accessible contexts in which to study the molecular actions guiding cell fate restriction, differentiation and patterning, each of which is crucial not only for regeneration to occur, but also for the survival and perpetuation of all multicellular organism

Genomic Regulation Of Abiotic Stress Response In The Soil Nematode Oscheius Tipulae

Oscheius tipulae is a species of free-living soil nematode that can be found in ecosystems worldwide. Because of this, individuals must be able to respond to heat, freezing, and desiccation stresses in order to survive. They do this by producing a suite of cellular responses, some of which are necessary to survive multiple stresses, and some are stress-specific. While these cellular responses are well known, the ways in which they are regulated in a genome-wide context are not. In this project, multiple high throughput sequencing and bioinformatics analyses were utilized to answer this question. First, the O. tipulae genome was sequenced via Illumina HiSeq, assembled, and annotated. An RNA-Seq experiment was performed to determine transcription patterns within stress responses. Pooled nematode samples were subjected to heat, freezing, or desiccation stress prior to RNA sequencing and read mapping. Results showed that shared cellular responses were controlled by the upregulation of both shared and stress-specific genes. This suggests that the genome remains efficient by utilizing overlapping response genes and reinforcing them with stress-specific genes. Whole genome bisulfite sequencing and MethylCap-Seq analyses were performed to assess DNA cytosine methylation presence in O. tipulae and the model organism Caenorhabditis elegans and to determine its role in the abiotic stress response process in O. tipulae. Methylated cytosines were found in both O. tipulae and C. elegans, contradicting the historical belief that cytosine methylation is absent in nematodes. Changes in DNA methylation were not associated with the abiotic stress response as very few methylation cites were found within upregulated genes. This project utilized new sequencing technologies and various bioinformatics programs to provide an in-depth look into the genome-wide responses to abiotic stress in O. tipulae

FUNCTIONAL AND MOLECULAR EVOLUTION OF THE PUF FAMILY

The modification of transcriptional regulation is a well-documented evolutionary mechanism in both plants and animals, but post-transcriptional controls have received less attention. The derived hermaphrodite of C. elegans has regulated spermatogenesis in an otherwise female body. PUF family RNA-binding proteins FBF-1 and FBF-2 limit XX spermatogenesis by repressing the male-promoting proteins FEM-3 and GLD-1. For my dissertation research, I examine the function of PUF homologs from other Caenorhabditis species, with emphasis on C. briggsae, which evolved selfing convergently. C. briggsae lacks a bona fide fbf-1/2 ortholog, but two members of the related PUF-2 subfamily, Cbr-puf-2 and Cbr-puf-1.2, do have a redundant germline sex determination role. Surprisingly, this is to promote, rather than limit, hermaphrodite spermatogenesis. I provide genetic, molecular, and biochemical evidence that Cbr-puf-2 and Cbr-puf-1.2 repress Cbr-gld-1 by a conserved mechanism. However, Cbr-gld-1 acts to limit, rather than promote, XX spermatogenesis. As with gld-1, no sex determination function for fbf or puf-2 orthologs is observed in gonochoristic Caenorhabditis. These results indicate that PUF family genes were coopted for sex determination in each hermaphrodite via their long-standing association with gld-1, and that their precise sex-determining roles depend on the species-specific context in which they act. Finally, I document non-redundant roles for Cbr-puf-2 in several aspects of somatic development. I show Cbr-puf-2 is required for reliable embryonic development, and that it is essential for vulval development and normal progression from early larval stage. I provide evidence suggesting that this latter role is related to pharyngeal muscle physiology. Thus, recently duplicated PUF paralogs, while redundant for some roles, can also rapidly acquire distinct non-redundant functions. This is consistent with theoretical models for the preservation of gene duplicates