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GENETIC AND MOLECULAR ANALYSIS OF GERMLINE SEX DETERMINATION IN CAENORHABDITIS BRIGGSAE, A MODEL FOR THE CONVERGENT EVOLUTION OF HERMAPHRODITISM
Doty, Alana V.
Haag, Eric S.
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Though sex determination and differentiation are critical biological processes, genetic mechanisms that specify sex have undergone profound and rapid evolutionary change across taxa. We may be able to infer processes that generate sex determination diversity by examining closely related species. Within the nematode genus Caenorhabditis, two species, C. elegans and C. briggsae, are androdioecious, producing self-fertile hermaphrodites and males; other Caenorhabditis species generate males/females. Interestingly, phylogenies reveal that C. elegans and C. briggsae independently acquired hermaphroditism, a relatively rare adaptation among animals. In this work, I describe differences in germline sex determination between C. elegans and C. briggsae that may help reveal the molecular basis of their convergent evolution of hermaphroditism. I first describe mutations in the pleiotropic, STAR family RNA-binding protein Cbr-GLD-1 that affect germline sex in C. briggsae. I find that C. briggsae gld-1 mutant hermaphrodites have a sex determination phenotype opposite to that of C. elegans: masculinized versus feminized germlines. I demonstrate that Cbr-GLD-1 coding-plus-regulatory sequences can rescue Ce-gld-1 null animals, arguing that this change in sex determination is not due to changes in GLD-1 function or expression. I further show that gld-1's role in regulating oogenesis is conserved across the Elegans group of Caenorhabditis, demonstrating that the oogenesis function of gld-1 is likely ancient, whereas its sperm-repressing role in C. briggsae has evolved recently. To identify mRNA targets of Cbr-GLD-1 that might be responsible for its sex determination function in C. briggsae, I use an in vivo genome-wide approach to isolate mRNAs associated with Cbr-GLD-1, including potential sex determination targets. I identify 800 putative mRNA targets and confirm specificity of this gene set via qRT-PCR and RNAi. Next, to reveal the roles of GLD-1 in evolutionary context, I create a phylogeny of STAR proteins across metazoans. Finally, I characterize a single feminizing allele recovered through forward screens in C. briggsae for germline sex determination mutants. This work thus begins to dissect the molecular and genetic basis of hermaphroditism in C. briggsae and contributes to a growing body of research on the evolution of germline sex determination differences between C. elegans and C. briggsae.