ACTIVATED RELEASE OF MEMBRANE-ANCHORED TGF-ALPHA IN THE ABSENCE OF CYTOSOL

The ectodomain of proTGF-alpha, a membrane-anchored growth factor, is converted into soluble TGF-alpha by a regulated cellular proteolytic system that recognizes proTGF-alpha via the C-terminal valine of its cytoplasmic tail. In order to define the biochemical components involved in proTGF-alpha cleavage, we have used cells permeabilized with streptolysin O (SLO) that have been extensively washed to remove cytosol. PMA, acting through a Ca2+-independent protein kinase C, activates cleavage as efficiently in permeabilized cells as it does in intact cells. ProTGF-alpha cleavage is also stimulated by GTPgammaS through a mechanism whose pharmacological properties suggest the involvement of a heterotrimeric G protein acting upstream of the PMA-sensitive Ca2+-independent protein kinase C. Activated proTGF-alpha cleavage is dependent on ATP hydrolysis, appears not to require vesicular traffic, and acts specifically on proTGF-alpha that has reached the cell surface. These results indicate that proTGF-alpha is cleaved from the cell surface by a regulated system whose signaling, recognition, and proteolytic components are retained in cells devoid of cytosol.