The RGD motif and the C-terminal segment of proprotein convertase 1 are critical for its cellular trafficking but not for its intracellular binding to integrin alpha(5)beta(1)

Cellular trafficking of subtilisin/kexin-like precursor convertases (PCs) m ay be regulated by a number of motifs, some of which are present within the P-domain and in the C-terminal sequence. Six of the seven known PCs contai n a conserved RGD sequence within the P domain. In order to investigate the functional importance of this motif, we generated mutants of PC1 that cont ain a Myc tag epitope inserted between the prosegment and the catalytic sub unit. Cellular expression of vaccinia virus recombinants revealed that this tag did not seem to influence the autocatalytic conversion of proPC1 into PC1 or its bioactivity, The two PC1 variants produced possess either the wi ld type RGD sequence or its RGE mutant. Stable transfectants of these varia nts in AtT20 cells revealed that similar to the wild type enzyme, PC1-RGD-M yc is sorted to secretory granules. In contrast, PC1-RGE-Mye exits the cell via the constitutive secretory pathway. In vitro, a 14-mer peptide spannin g the RGD sequence of PC1, but not its RGE mutant, binds to cell surface vi tronectin-binding integrins of Chinese hamster ovary cells. However, within the endoplasmic reticulum and in an RGD-independent fashion, integrin alph a(5)beta(1), associates primarily with the zymogens proPC1, proPC1-Delta C (missing the C-terminal 137 residues), as well as proPC(2). Thus, the obser ved discrimination between the secretion routes of PC1-RGD and PC1-RGE does not implicate integrins such as alpha(5)beta(1).