In vivo expression profile of a H+-K+-ATPase alpha2-subunit promoter-reporter transgene.

Abstract

Because little is known about the molecular basis of transcriptional regulation of the murine H(+)-K(+)-ATPase alpha(2) (HKalpha(2)) gene or other genes whose expression is restricted in part to the collecting duct, especially in vivo, we developed transgenic mice carrying an insertional HKalpha(2) promoter-reporter gene construct. In these mice, the region -7,264/+253 of the HKalpha(2) 5'-flanking region controls expression of the reporter gene enhanced green fluorescent protein (EGFP). Patterns of HKalpha(2)/EGFP transgene expression were examined by fluorescence microscopy and immunoblotting. Of 10 major organs examined, EGFP immunoreactivity was detected abundantly in the kidney, and to a far lesser extent, in the brain and lung. Within the kidney, EGFP fluorescence was detected exclusively in the collecting ducts of transgenic mice and colocalized with the cellular distribution of both endogenous HKalpha(2) and aquaporin-2, consistent with the known expression pattern of endogenous HKalpha(2) in principal cells. Surprisingly, no transgene expression was evident by immunoblotting or fluorescence microscopy in the distal colon, the site of the highest endogenous HKalpha(2) expression. Although previous studies of steady-state mRNA levels suggested differences in HKalpha(2) gene regulation in the kidney and colon, our results provide the first direct evidence of differential transcriptional control of the HKalpha(2) gene in these organs and suggest that regions outside the 5'-flanking region or other regulatory factors play a role in HKalpha(2) expression in the distal colon.