Intracellular Localization of Cthrc1 Characterizes Differentiated Smooth Muscle

Abstract

Objective— We recently reported expression of collagen triple helix repeat containing-1 (Cthrc1) in injured arteries and proteolytic cleavage of Cthrc1 in smooth muscle cells in vitro. The present study characterizes Cthrc1 processing and determines its biological significance. Methods and Results— Domain-specific antibodies localized full-length Cthrc1 in the cytoplasm of vascular, gastrointestinal, and uterine smooth muscle as well as in some neurons. Unlike smooth muscle α-actin, Cthrc1 was not expressed in the embryonic myocardium. Intracellular localization of full-length Cthrc1 was sharply reduced in dedifferentiated smooth muscle of the developing neointima despite the previously shown increase in mRNA levels with accompanying extracellular Cthrc1 immunoreactivity. Immunoblotting suggested an apparent covalent association of monomeric full-length Cthrc1 with a cytoplasmic protein present in differentiated smooth muscle. Plasmin was identified as a protease that cleaved a putative propeptide generating an N-terminally truncated form of Cthrc1 with increased inhibitory activity of procollagen synthesis. Conclusions— Our data show that the differentiated smooth muscle cell phenotype is associated with the intracellular localization of noncleaved Cthrc1 despite the presence of a signal peptide. On arterial injury, increased Cthrc1 expression with apparent extracellular localization of N-terminally truncated Cthrc1 occurs. Removal of the propeptide correlated with increased activity of the molecule.