Metalloprotease-mediated cleavage secretion of pulmonary ACE by vascular endothelial and kidney epithelial cells.

Abstract

The pulmonary isozyme of angiotensin-converting enzyme (ACEP) is present in the body both as a cell-associated protein in endothelial, epithelial, and monocytic cells and as a soluble protein in various body fluids including serum. The mechanism by which soluble ACEP is produced in vivo is unknown. Using in vitro transfected cell culture systems, we previously demonstrated that the rabbit testicular isozyme of ACE (ACET), which shares extensive homology with ACEP, is first synthesized as a plasma membrane-anchored ectoprotein and then secreted to the culture medium by cleavage removal of its COOH-terminal membrane-anchored tail. Here, using in vitro cultures of arterial endothelial cells and acutely isolated renal epithelial cells, we demonstrate that ACEP is also cleavage secreted from their natural producer cells. Biochemical and immunological characterization of the in vitro secreted ACEP protein revealed that it is missing the COOH-terminal membrane-anchored region of the cell-associated ACEP. Similar analysis of ACEP proteins present in rabbit serum, lung, and kidney established that ACEP secretion in vivo is also caused by the cleavage removal of the COOH-terminal region of the cell-associated protein. To characterize the proteolytic enzyme responsible for ACEP secretion, we employed rabbit renal proximal tubular epithelial cells and demonstrated significant inhibition of secretion by compound 3, a hydroxamic acid-based inhibitor of specific metalloproteases. In contrast, the inhibitors of chymotrypsin, trypsin, serine, aspartate, and cysteine proteases were ineffective. These results indicate that soluble ACEP production by vascular endothelial and renal epithelial cells, both in vitro and in vivo, is achieved by cleavage removal of its membrane-anchoring COOH-terminal tail by a metalloprotease.