Biased and unbiased approaches to identify candidate genes involved in Hydra ageing

Diplôme Master Biased and unbiased approaches to identify candidate genes involved in Hydra ageing eng 2022-08-19 author Simons Sasha Alex William 856438 director Galliot Brigitte 0000-0001-7596-8284 29979 Département de génétique et évolution (Genev) (BIANI) director Suknovic Nenad Slavko 813769 155 The Hydra cnidarian is a freshwater polyp consisting of two layers of cells derived from three distinct populations of adult stem cells that constantly self-renew and give these animals an unprecedented capacity for regeneration and negligible senescence. Hydra oligactis (Ho) shows no signs of ageing when maintained at 18°C in an asexual state. However, when transferred to 10°C, animals become sexual, lose the ability to regenerate and begin to age. Previous research in our laboratory has identified two distinct strains of Ho that show a differential response to cold-induced gametogenesis. One strain, known as cold-sensitive (Ho_CS), undergoes rapid sexual differentiation after cold transfer and shows physiological changes synonymous with aging: progressive loss of fitness and increased mortality over time. By contrast, the second strain, known as cold-resistant (Ho_CR), also undergoes gametogenesis induced by cold at 10°C, but is not subject to the same age-related degeneration, and its physical condition remains preserved. Previous studies have shown that autophagy is deficient in the epithelial stem cells of aging Ho_CS Hydra, but not in the cells of Ho_CR animals. This discovery highlights an essential metabolic difference between the two strains that explains their differential aging. This paves the way for further research to understand the differences between aging and non-aging animals, in the hope of identifying new regulators of the genetic pathways that might explain this differential response to aging. In this study, we use a series of bioinformatic approaches designed to identify differentially expressed genes (DEG) between Ho_CR and Ho_CS and validate some of them as candidate regulators of aging in Hydra. In a first approach, we used proteomic and transcriptomic data to select differentially expressed candidates overlapping between both data sets. We next analysed their expression patterns by in situ hybridisation (ISH) and could confirm the differential expression between Ho_CR and Ho_CS after transfer at 10°C. It was our intention to test their function and their impact on non-ageing animals, we would knock-down the expression of some candidate genes by RNA interference and despite a limited success, we identified CPB1 as a bona fide tool for monitoring aging in Hydra. In a second approach, we used only transcriptomic data and applied a targeted approach to select among DEGs those that are both conserved between humans and Hydra, and previously identified as being involved in aging in humans. In this way, we confirmed the differential expression of two genes GSR and NRF3 and showed a specific loss of fitness in siNRF-treated animals. We also performed an in-depth literature review to find their function(s) and infer their role in ageing. We propose that NRF3 plays a role in lipogenesis and thus be identified as part of the nutrient sensing pathway. In a third approach, unbiased this time, we performed a complete in silico analysis of the transcriptomic data to establish the repertoire of DEGs using pipelines such as: BLAST, Mapping, k-mean clustering and ingenuity pathway analysis. This was done in order to identify the key signalling cascades potentially involved in the phenotypic differences observed between aging and non-aging strains. This third approach allowed us to identify as a robustly supported candidate, the signalling cascades MYC as the centre for one of the top up-regulated pathways. Ageing or aging Autophagy Evolution Bioinformatics Differentially expressed genes (DEG) Proteomic and transcriptomic data Cnidarian Hydra oligactis Hydra cold-sensitive and cold-resistant strains RNA interference Ingenuity pathway analysis MYC as top-candidate regulator of ageing NRF3-like as candidate regulator Dewey 590 Master thesis in Biology specific option Genetics, Development & Evolution 44 Département de génétique et évolution 162 false 821 Groupe Galliot https://hydratlas.unige.ch/ ce814703-fa2a-4372-ad48-1832fc164f97 Master thesis Restricted 2022_SIMONS_Sasha_Master-Thesis_AO.pdf 14650908 application/pdf 73ddac4262432248532d5dddf56d1b2d6f9480b971f51e4e6010624202eec5f9 restrictedAccess Faculté des sciences / Section de biologie / Département de génétique et évolution

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