The mammalian Y chromosome acts as a dominant male determinant as a result of the action of a single gene, Sry, whose role in sex determination is to initiate testis rather than ovary development from early bipotential gonads1,2,3. It does so by triggering the differentiation of Sertoli cells from supporting cell precursors, which would otherwise give follicle cells. The related autosomal gene Sox9 is also known from loss-of-function mutations in mice and humans to be essential for Sertoli cell differentiation4,5; moreover, its abnormal expression in an XX gonad can lead to male development in the absence of Sry6,7. These genetic data, together with the finding that Sox9 is upregulated in Sertoli cell precursors just after SRY expression begins8,9, has led to the proposal that Sox9 could be directly regulated by SRY. However, the mechanism by which SRY action might affect Sox9 expression was not understood. Here we show that SRY binds to multiple elements within a Sox9 gonad-specific enhancer in mice, and that it does so along with steroidogenic factor 1 (SF1, encoded by the gene Nr5a1 (Sf1)), an orphan nuclear receptor. Mutation, co-transfection and sex-reversal studies all point to a feedforward, self-reinforcing pathway in which SF1 and SRY cooperatively upregulate Sox9 and then, together with SF1, SOX9 also binds to the enhancer to help maintain its own expression after that of SRY has ceased. Our results open up the field, permitting further characterization of the molecular mechanisms regulating sex determination and how they have evolved, as well as how they fail in cases of sex reversal.
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We thank J. Carvajal and T. Kondo for teaching us BAC engineering and ChIP assays, respectively; D. Bell and S. Wood for help in generating transgenic mice; M. Cheung for analysing neural crest cells in transgenic embryos; K. Parker for providing SF1 complementary DNA and SF1 null mutant mice; A. Schedl for providing conditional Sox9 mutant mice; K. Morohashi for providing the Ad4BP/SF1 antibody; H. Sasaki for providing the Asshsp68lacZ vector; Y. Kamachi for providing the pCMV/SV2-Sox9 and pδ51LucII vectors; and V. Harley and the members of the Division of Developmental Genetics in the National Institute for Medical Research, especially C. Wise, F. W. Buaas, J. Turner and P. Burgoyne, for suggestions and discussions. This work was supported by the Medical Research Council and the Louis Jeantet Foundation. R.S. was a recipient of long-term fellowships from the European Molecular Biology Organization and the Human Frontier Science Program.
Author Contributions R.S. was responsible for most of the experiments, and R.L.B. for a few experiments as well as for directing the laboratory. Both planned the project and wrote the manuscript.
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Sekido, R., Lovell-Badge, R. Sex determination involves synergistic action of SRY and SF1 on a specific Sox9 enhancer. Nature 453, 930–934 (2008). https://doi.org/10.1038/nature06944
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