TIN-ag-RP, a novel catalytically inactive cathepsin B-related protein withEGF domains, is predominantly expressed in vascular smooth muscle cells

A human cDNA of 2166 bp encoding a novel cathepsin B-related protein was is olated and characterized. The amino acid sequence of the predicted protein of 467 aa was 46% identical with that of human tubulointerstitial nephritis antigen (TIN-ag), and therefore, the protein was tentatively designated as the TIN-ag-related protein (TIN-ag-RP). The amino acid sequence of TIN-ag- RP is composed of a 21 aa long signal sequence, a 181 aa long N-terminal do main containing two epidermal growth factor-like domains, a follistatin mot if, and a 265 aa long cathepsin B-like domain. Interestingly, a serine resi due has replaced the active site cysteine residue in the cathepsin B-like d omain, resulting in a proteolytically inactive protein. Evolutionary analys is revealed that a distinct family of "TIN-ag-like" proteins had evolved in vertebrates. Northern blot analysis revealed a single TIN-ag-RP transcript of 2.4 kb in various tissues with the highest transcript levels detected i n aorta, heart, placenta, skeletal muscle, kidney, and a colorectal adenoca rcinoma cell line. Using a polyclonal anti-TIN-ag-RP antibody, TIN-ag-RP ex pression was predominantly seen in vascular smooth muscle (VSM) cells, but also in cardiac and skeletal muscle cells as well as in kidney. Interesting ly, uterine smooth muscle cells completely lacked TIN-ag-RP expression, imp lying a regulated gene expression. Localization studies in HeLa cells stabl y transfected with TIN-ag-RP cDNA showed that TIN-ag-RP is glycosylated and actively secreted, a finding in line with its proposed function as a struc tural or regulatory protein similar to TIN-ag.