The basic helix-loop-helix transcription factor TCF4 recruits the Mediator Complex to activate gonadal genes and drive ovarian development.

The bipotential gonad is the precursor organ to both the ovary and testis and develops as part of the embryonic urogenital system. In mice, gonadogenesis initiates around embryonic day 9.5 (E9.5), when coelomic epithelial (CE) cells overlaying the mesonephric ducts proliferate and acquire the competence to differentiate into the two main cell types of the embryonic gonad, the pre-supporting cells and interstitial cell lineages. While some transcription factors that drive gonadal cell fate are known, HLH factors have not been investigated in this capacity. In the present study, we found that HLH binding sites are highly represented upstream of gonadal genes. We investigated the HLH factor Transcription Factor 4 (TCF4) which is expressed in the CE and GATA4+ somatic cells in both sexes prior to sex determination. TCF4 is maintained in ovarian pre-supporting cells and interstitial cells of both sexes but is silenced specifically in male pre-supporting cells. To characterize TCF4's role in gonad differentiation we acquired a mutant mouse model that lacks the TCF4 DNA-binding domain and assessed morphology of the gonads at E15.5. While mutants develop gonads, we observed sex-specific effects on the gonads. Relative to wildtype littermates, SOX9 expression was higher in the Sertoli cells of XY mutant testes, while FOXL2 and NR2F2 were reduced in the supporting and interstitial cell lineages of XX mutant ovaries, respectively. Furthermore, the supporting: interstitial cell ratio was altered in XX ovaries. These effects may occur downstream of changes to epigenetic programming or gene expression in somatic gonadal cells in mutant mice, as TCF4 binds the Mediator complex, RNA polymerase holoenzyme, and chromatin remodelers in early somatic cells. We hypothesize that TCF4 drives a gonadal program that advances female fate but is specifically silenced in male supporting cells as these pathways diverge.