Calpain cleavage of integrin beta cytoplasmic domains

We showed previously that the calcium-dependent protease, calpain, cleaves the cytoplasmic domain of the integrin beta 3 subunit, To investigate wheth er susceptibility to calpain is a common feature of all integrin beta subun its, and to map calpain cleavage sites in different integrin beta tails, we treated recombinant cytoplasmic domains of integrin beta 1A, beta 1D, beta 2, beta 3 and beta 7 subunits with purified calpain in vitro. We found tha t the cytoplasmic domains of all these integrin chains were cleaved by calp ain, HPLC followed by mass spectrometry was used to identify calpain cleava ge sites. These sites were clustered in the C-terminal half of the integrin beta cytoplasmic domains in regions flanking the two NXXY motifs, suggesti ng the possibility that the structural framework provided by these motifs i s recognized by calpain, We used the knowledge of these cleavage sites to d evelop cleavage site-specific antibodies and to demonstrate cleavage of the beta 1A cytoplasmic domain in intact platelets stimulated with calcium ion ophore or thrombin, Thus susceptibility to calpain cleavage is common to in tegrin beta subunits, can be induced in intact cells, and appears to favor regions surrounding two conserved NXXY motifs, (C) 1999 Federation of Europ ean Biochemical Societies.