AP-2 family members regulate basal and cAMP-induced expression of human chorionic gonadotropin

The AP-2 family of transcriptional regulator proteins has three members, α, β and γ. AP-2α and γ are expressed in placenta and in the human trophoblast cell line JEG-3. AP-2 has been shown to regulate expression of the placental human chorionic gonadotropin (hCG) α- and β-subunit genes, however, previous work did not distinguish between the family members. Tryptic peptides of the AP-2 protein complexes purified from JEG-3 cells by oligo-affinity chromatography using the hCGα AP-2 site match the amino acid sequence of AP-2γ. The fact that AP-2γ is present at significant levels and binds the hCGα trophoblast-specific element suggests that AP-2γ is at least part of the binding complex in vivo and plays a role in regulating hCG expression. We show that mutation of each of four AP-2 binding sites within the hCGβ promoter decreases expression in transfection assays, demonstrating that all four sites are required for maximal expression in JEG-3 cells. Furthermore, we find differences in regulation of the family members: AP-2α mRNA levels increase in response to cAMP while AP-2γ mRNA levels do not. The demonstrated importance of the AP-2 sites in controlling hCGα and β expression and the likely involvement of more than one family member suggest that a balance in AP-2 proteins is involved in coordinate regulation of these genes. Moreover, many placenta-restricted genes are regulated by AP-2 proteins, thus members of this family may play an important overall role in placenta-specific expression.