In vivo expression profile of a H+-K+-ATPase α2-subunit promoter-reporter transgene

Abstract

Because little is known about the molecular basis of transcriptional regulation of the murine H⁺-K⁺-ATPase α₂(HKα₂) gene or other genes whose expression is restricted in part to the collecting duct, especially in vivo, we developed transgenic mice carrying an insertional HKα₂promoter-reporter gene construct. In these mice, the region −7,264/+253 of the HKα₂5′-flanking region controls expression of the reporter gene enhanced green fluorescent protein (EGFP). Patterns of HKα₂/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 HKα₂and aquaporin-2, consistent with the known expression pattern of endogenous HKα₂in principal cells. Surprisingly, no transgene expression was evident by immunoblotting or fluorescence microscopy in the distal colon, the site of the highest endogenous HKα₂expression. Although previous studies of steady-state mRNA levels suggested differences in HKα₂gene regulation in the kidney and colon, our results provide the first direct evidence of differential transcriptional control of the HKα₂gene in these organs and suggest that regions outside the 5′-flanking region or other regulatory factors play a role in HKα₂expression in the distal colon.