Investigating the Role of wnt9b in Zebrafish Sex Determination and the Role of gjc4b in Early Zebrafish Gonad Development

The gonad is vital for successful sexual reproduction and species’ survival. Gonad development is wellstudied in mammals, but far less is known about its development and subsequent sex determination in teleosts. In mammals, early gonad development begins with the specification of primordial germ cells (PGCs) that migrate to the genital ridge. After the coalescence of PGCs and somatic gonad cell precursors at the genital ridge, the primordial gonad undergoes significant proliferation and eventual sex differentiation. However, in zebrafish, early gonad development and sex determination remain to be understood. There is growing evidence that gap junction proteins have an integral role in supporting early germ cell development in mammals. Gap junctions serve to transfer large molecules between cells for communication as well as small metabolites. In the ovary, there are two known gap junction proteins, Gja1 and Gja4, that connect the oocyte to the somatic granulosa cells in order to facilitate metabolic support. We recently found that the gap junction protein gjc4b is expressed in zebrafish oocytes through single-cell RNAseq. We hypothesize that gjc4b may serve a similar role as Gja4 in connecting the somatic follicle cells to the developing oocytes. During our investigation, I discovered that gjc4b mutants have normal PGC migration and apparent survival but have deformed or absent germ cells past 20 days post fertilization (dpf) that retain a PGC-like state instead of transitioning to a gonocyte. Additionally, there is significant evidence in mammals that Wnt4 is the female sex determination factor. We have previously shown that the zebrafish ortholog, wnt4, is also required for primary female sex determination: mutants develop with a significant male bias and are largely incapable of initiating any oocyte development that would be indicative of the potential for female development. We have recently identified an additional Wnt ligand expressed in the juvenile ovary through single-cell RNAseq analysis called wnt9b. I investigated its role in sex determination and found through wnt9b mutant analysis that wnt9b plays a role in female sex maintenance