In the mammalian testis, the binding of FSH to Sertoli cells activates the cAMP-dependent protein kinase A signaling pathway, resulting in the phosphorylation of the cAMP response element binding protein (CREB). Previous studies have also shown that CREB gene expression is activated by cAMP in Sertoli cells and that 2 cAMP response elements (CREs) that bind CREB and a neighboring Sp1 binding site are required for basal and cAMP-inducible CREB promoter activity. In contrast, CREB expression has been less well characterized in testis germ cells. We demonstrated that CREB and Sp1 are expressed in early germ cells only through the midpachytene stage of spermatogenesis. Furthermore, CREB promoter activity was induced over 70-fold by transient overexpression of Sp1 in SL2 cells, suggesting that Sp1 is an important regulator of CREB expression. Further studies of the CREB promoter revealed an additional regulatory element in the −130 region between the Sp1 and CREB transcription factor binding sites that is necessary for full promoter activity. Proteins expressed in Sertoli cells and germ cells bind specifically to the newly identified regulatory region. These studies suggest that proteins binding to Sp1 motifs and the −130 region are required to activate the CREB promoter.