Self-fertile hermaphroditism arose independently multiple times in Caenorhabditis through the evolution of post-transcriptional gene regulation in the germline. Previous studies suggest that RNA binding proteins (RBPs) orchestrate the evolution of germline development by gaining and/or losing target transcripts. STAR domain RBPs are found across eukaryotes, and Caenorhabditis has a large nematode-specific family of them that includes the germline-specific GLD-1. GLD-1 has conserved, pleiotropic roles in meiotic commitment and oogenesis [1, 2] but has acquired a role in hermaphrodite sex determination in two androdioecious species, C. elegans and C. briggsae [2-4]. Its main known function with respect to sex determination in C. elegans is to bind and repress tra-2 mRNA in hermaphrodites [5, 6] allowing the production of sperm in an otherwise female soma. In contrast, Cbr-GLD-1 promotes oogenesis and does not detectably bind tra-2 in vivo. Despite GLD-1's opposing roles in the two species, Cbr-GLD-1 can rescue a Ce-gld-1 null mutant, suggesting that mRNA targets are the changing entity [2]. C. briggsae and C. elegans GLD-1 have both shared and species-specific GLD-1 targets [7], and some C. briggsae GLD-1 binding sites appear to lack GLD-1 binding motifs. To better understand the dynamic evolution of GLD-1 targets, we are developing PAR-CLIP (photoactivatable-ribonucleoside-enhanced crosslinking and immunoprecipitation) to obtain the exact site of GLD-1 binding on C. briggsae target mRNAs. PAR-CLIP causes a recognizable sequence change reducing the possible binding site to a 20-40 bp region. Having a clear picture of GLD-1 binding sites in two independently derived hermaphrodites is essential to our understanding of this evolutionary mechanism. In addition, to provide further evidence that GLD-1 has recently gained the tra-2 regulator in C. elegans, we are performing a simpler mRNA-immunoprecipitation-qRT-PCR assays for GLD-1-tra-2 mRNA association in C. remanei, C. brenneri and C. japonica (outcrossing). We expect that GLD-1 will only associate with tra-2 in C. elegans. 1. Francis, R., et al. Genetics, 1995. 139: p. 579-606. 2. Beadell, A.V., et al. Proc Natl Acad Sci U S A, 2011. 108(49): p. 19672-7. 3. Francis, R., E. Maine, and T. Schedl, Genetics, 1995. 139: p. 607-30. 4. Nayak, S., J. Goree, and T. Schedl, PLoS Biology, 2005. 3: p. e6. 5. Goodwin, E.B., et al. Cell, 1993. 75: p. 329-339. 6. Jan, E., et al. EMBO J., 1999. 18(1): p. 258-69. 7. Beadell, A.V. and E.S. Haag, Genome Biol Evol, 2015. 7(1): p. 314-35.

Affiliations:
- Department of Biology, University of Maryland, College Park, MD
- Program of Behavior, Evolution, Ecology, and Systematics, University of Maryland, College Park, MD