Identification of a zinc finger homoeodomain enhancer protein after AT(2) receptor stimulation by differential mRNA display.

Using differential mRNA display, we isolated differentially expressed genes after stimulation of angiotensin II (Ang II) type 2 (AT(2)) receptors in PC12w cells. Among the polymerase chain reaction products isolated, we identified Zfhep, a zinc finger homoeodomain enhancer-binding protein and the angiotensin AT(2) receptor itself, both implicated in developmental processes. In the presence of epidermal growth factor (EGF) a concordant expression pattern of Zfhep and AT(2) mRNA was observed with marked downregulation 6 hours after stimulation with EGF+Ang II. In quiescent PC12w cells, Zfhep mRNA was time-dependently induced by Ang II (10(-7) mol/L) up to 3 hours, an effect that was blocked by the selective AT(2) antagonist PD123177 (10(-6) mol/L). We extended our studies on Zfhep mRNA expression to cells of vascular origin (coronary endothelial cells [CECs]), that express AT(1) and AT(2) receptors, and that respond differently to Ang II dependent on which receptor subtype is stimulated. In CECs, Zfhep mRNA expression was induced up to 3 hours after AT(2) stimulation. Interestingly, this Zfhep induction was observed only when the AT(1) receptor was blocked by using the selective AT(1) receptor antagonist, losartan (10(-5) mol/L), suggesting a negative-regulatory influence of AT(1) receptors on AT(2)-induced Zfhep mRNA induction. In conclusion, Zfhep not only represents a suitable marker for AT(2) receptor activation, but it may also link AT(2) signaling and downstream events involved in the proposed function of the AT(2) receptor in development and regeneration.