ENDOGENOUS ROLES OF IMPORT OF DOUBLE-STRANDED RNA DURING DEVELOPMENT IN C. ELEGANS

Abstract

The information contained in a typical animal cell originates with the fusion of gametes but is transformed during development and influenced by the environment the organism has lived through. Such epigenetic changes can outlast the original perturbation and affect subsequent generations. Understanding how such heritable epigenetic changes are transmitted is therefore central to understanding both development and heredity. For epigenetic changes to be transmitted to the next generation, they need to alter the germ line. RNA-mediated gene regulation is one such mechanism. The import of double-stranded RNA (dsRNA) into the germ line can have consequences that last for many generations. However, the importance of such transgenerational regulation by extracellular dsRNA for an organism and its descendants is unclear. In the tractable animal Caenorhabditis elegans, the entry of exogenously introduced dsRNA into the cytosol requires the transmembrane protein SID-1 but not much is known about the endogenous roles of such dsRNA import, if any. Transient loss of SID-1 can cause changes in gene expression that can persist for hundreds of generations. This dissertation builds on these findings to answer what are the endogenous roles of imported extracellular dsRNAs and which developmental processes are susceptible to such heritable. Here we expand the number of SID-1-dependent genes (SDGs) that show transgenerational changes in gene expression and provide evidence for the regulation of reproduction by two such genes that are expressed in the germ line: sdg-1 and sdg-2. The SDG-1 protein shows dynamic subcellular localization that includes conditional enrichment within nuclei, colocalization with perinuclear germ granules, and colocalization with microtubules. Animals with sdg-1 mRNA overexpression show germline defects that include a delay in germline development, resulting in reduced numbers of early progeny. The SDG-2 protein is in the cytoplasm and nucleus and is predicted to interact with the Gli-type transcription factor TRA-1, which promotes spermatogenesis. Some lineages of animals lacking SDG-2 have fewer total progeny despite having normal numbers of sperm and they begin egg laying early. Therefore, changes in these two SDG proteins can alter the numbers of early progeny, which is a key variable for fitness in resource-limited settings. Analysis of additional SID-1-dependent genes could similarly reveal processes that have evolved to rely on SID-1, and potentially the regulation of genes by extracellular dsRNA, to buffer against the lingering impacts of ancestral epigenetic changes. Understanding which developmental processes are regulated by endogenous dsRNA import and how such regulation shapes heredity will have implications both for fundamental biology and for predicting the transgenerational consequences of dsRNA-based therapeutics.