Precise control of gene expression is required for growth, development and metabolism. Abnormalities in transcription have been linked to a variety of human diseases from birth defects to neoplasia. The GATA-binding proteins are a family of zinc finger transcription factors that regulate gene expression, differentiation and cell proliferation in a variety of tissues by binding to the consensus DNA sequence (A/T)GATA(A/G). By analogy to the well studied hematopoietic GATA factors, the other vertebrate GATA-binding proteins are postulated to have critical roles in other cell types. We have previously shown that the expression of GATA-4 and GATA-6 transcripts in the ovary of normal mice responses to gonadotropin stimulation. Given the emerging role of GATA-4 and GATA-6 in gonadal cells, we have now studied the role of these transcription factors in mouse cell lines and their mRNA and protein expression in developing testis. In situ hybridization demonstrated GATA-4 expression in early mouse gonadal ridge (E11.5), in fetal (E16), neonatal(D4), juvenile (D14) and adult testis. Leydig cells were strongly positive, and GATA-4 mRNA was also present in a subset of Sertoli cells, spermatogonia and spermatocytes. RNase protection assay confirmed these findings. By IF, we showed GATA-4 protein in adult Leydig cells. Considerable GATA-6 mRNA was detected in embryonal (E16) Sertoli cells and less abundantly in neonatal through adult Sertoli cells. The in vivo finding on the expression of gonadal GATA transcripts was confirmed in studies on established Sertoli and Leydig cell lines. Based on the established role of GATA-binding proteins in gene expression and lineage commitment as well as the findings presented we propose that GATA-4 and -6 play critical roles in differentiation and function of the male gonad. We are in the process of dissecting the regulation of these GATA factors in testis, their target genes and their significance in disease using animal models and human testicular samples from pediatric and adult patients.