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Cell type-specific genome stability in the non-ageing animal Hydractinia symbiolongicarpus

Horkan, Helen R.
Citation
Abstract
The marine cnidarian Hydractinia is highly regenerative, does not age, develops no spontaneous neoplasia, and is highly resistant to ionizing irradiation (IR). These features are thought to depend on a population of adult pluripotent stem cells, called i-cells. I hypothesized that these features are supported by a highly stable genome in some or all Hydractinia cell types. Studying the mechanisms that underlie genome stability in this animal was the main aim of my PhD. As a first step, I show that Hydractinia possesses no unique protection against IR-induced DNA double-strand breaks (DSBs). Furthermore, DSB repair occurred within 24, in line with other organisms. However, in contrast to other animals, Hydractinia stem cells (i-cells) are not more sensitive to IR when compared to differentiated cells. Following irradiation, i-cells exited the cell cycle for an extended period but remained alive as demonstrated by flow cytometry. Eventually, cell cycle re-entry was observed, followed by the animals complete resumption to clonal growth and sexual reproduction. To explore genome stability at single cell level, high quality omics resources are required. I participated in the chromosome-level genome assembly and analysis for male clone 291-10. An additional chromosome-level assembly was generated for the female clone 295-8. A male cell type atlas of 200,000 cells was generated by mapping barcoded RNA reads from single-cell ACME SPLiT seq to the reference genome. All known cell types and some novel ones were identified, including a stolon enriched type. An additional female single cell dataset was generated and merged with the male atlas, identifying an oogenesis-specific cluster with novel marker genes. To assess IR response, I generated a single cell dataset from animals at 1- and 9-days post exposure to 50 Gy. I merged the post IR libraries with the reference atlas and found that all major cell types were captured in the IR dataset. I demonstrate, on a single cell level, that the mutational load decreases significantly between 1- and 9-days post IR, and is generally lower in i-cells, indicating the presence of a post DSB repair mutation elimination (PRiME) mechanism in some or all cell types. Such a mechanism has never been recorded previously in any organism. I identified Hydractinia homologues for 163 genes implicated in DNA damage response in humans, including all genes required for canonical homologous recombination and non-homologous end joining. I searched for genes involved in PRiME by assessing differential expression of genes between specific cell types during recovery from high dose IR, providing candidate genes for future functional studies. Understanding the mechanisms involved in PRiME could provide insight into cnidarians' overall resilience to age-related degeneration and cancer, and have future applications in human disease research.
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University of Galway
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Attribution-NonCommercial-NoDerivatives 4.0 International