Angiotensin II Differentially Regulates Morg1 Expression in Kidney Cells

Abstract

Background: The mitogen-activated protein kinase organizer 1 (Morg1) belongs to the WD-40 repeat protein family and is a scaffold molecule for the extracellular regulated kinase signaling pathway. Morg1 also binds to prolyl-hydroxylase 3 (PHD3) and regulates the hypoxia-inducible factor-1α (HIF-1α) expression via PHD3 stabilization. Morg1 has been detected in the kidney as well as in other cell tissues but its expression in renal cells has not been well investigated. It has been widely shown that angiotensin II (ANG II) mediates renal damage. We have previously shown that ANG II downregulates the expression of PHD3 in PC12 cells. The aim of this study was to analyze whether ANG II regulates Morg1 expression in mouse mesangial cells (MMC), mouse proximal tubular cells (MTC) and in differentiated podocytes. The correlation between the expression of Morg1 and PHD3 activity was also addressed. Methods: Effect of ANG II on the Morg1 mRNA expression level was assessed by real-time PCR. Morg1 and HIF-1α cellular localization was analyzed by immunohistochemistry. HIF-1α promoter activity was investigated using a reporter gene system. PHD3 hydroxylase activity test was measured with a hydroxylation-coupled decarboxylation assay. Results: ANG II differentially regulates Morg1 expression in MMC, MTC and differentiated podocytes. We detected a biphasic effect of ANG II on Morg1 mRNA expression which was time dependent. While 9-hour ANG II treatment downregulated Morg1 expression in MMC, it induced Morg1 expression in MTC. Conversely, 24-hour ANG II stimulation upregulated the expression of Morg1 mRNA in MMC, but showed an opposite effect in MTC and differentiated podocytes. In addition, we found that ANG II signals mostly through the AT₁ receptor subtype in MMC and via the AT₂ subtype in MTC. PHD3 activity correlated to Morg1 expression patterns. Our data also demonstrate that HIF-1α transcriptional activity in MTC contrasted to PHD3 activity at 9 and 24 h, whereas in the MMC and in podocytes we did not find any correlation between PHD3 HIF-1α hydroxylation ability and HIF-1α transcriptional activation, suggesting a different mechanism of regulation in these cell types. Interestingly, the reduced expression of Morg1 in mesangial cells isolated from Morg1 (+/–) heterozygous mice correlated with a reduced PHD3 enzymatic activity and an increased HIF-1α transcriptional activity compared with mesangial cells originated from wild-type (Morg1 +/+) mice. Conclusions: We show for the first time in various renal cells that ANG II modulates Morg1 expression and HIF-1α transcriptional activity via cell type-specific mechanisms, demonstrating a novel mechanism by which ANG II may contribute to renal disease.