IN-VITRO PHOSPHORYLATION OF CAVEOLIN-RICH MEMBRANE DOMAINS - IDENTIFICATION OF AN ASSOCIATED SERINE KINASE-ACTIVITY AS A CASEIN KINASE II-LIKE ENZYME

Citation
M. Sargiacomo et al., IN-VITRO PHOSPHORYLATION OF CAVEOLIN-RICH MEMBRANE DOMAINS - IDENTIFICATION OF AN ASSOCIATED SERINE KINASE-ACTIVITY AS A CASEIN KINASE II-LIKE ENZYME, Oncogene, 9(9), 1994, pp. 2589-2595
Citations number
36
Language
INGLESE
art.tipo
Article
Categorie Soggetti
Genetics & Heredity",Oncology
Journal title
ISSN journal
0950-9232
Volume
9
Issue
9
Year of publication
1994
Pages
2589 - 2595
Database
ISI
SICI code
0950-9232(1994)9:9<2589:IPOCMD>2.0.ZU;2-Z
Abstract
Caveolae are flask-shaped micro-invaginations associated with the plas ma membrane of a wide variety of cell types. Caveolin, an integral mem brane component of caveolae, was first identified as the major phospho protein whose phosphorylation was elevated in v-Src transformed cells. As both v-Src transformation and elevated caveolin phosphorylation we re dependent on membrane attachment of v-Src, it has been suggested th at caveolin is a critical target in v-Src transformation. Although an increase in tyrosine phosphorylation of caveolin was evident, the incr ease in caveolin phosphorylation was predominantly on serine residues. In accordance with these in vivo observations, isolated caveolin-rich membrane domains undergo phosphorylation in vitro predominantly on se rine and contain an unidentified serine kinase activity. Here, we have identified this serine kinase activity as a casein kinase II-like enz yme, since the phosphorylation of caveolin-rich membrane domains is st imulated and inhibited by known effecters of casein kinase II (poly-L- lysine, endogenous polyamines, and a casein kinase II inhibitor peptid e), but is unaffected by modulators of other known kinases. In support of these observations, caveolin contains a consensus sequence for cas ein kinase II phosphorylation in its cytoplasmic N-terminal domain (Se r-88). A peptide containing this sequence inhibits the in vitro phosph orylation of caveolin-rich membrane domains, while many other peptides derived from the N-terminal domain of caveolin do not affect phosphor ylation. Caveolin-rich membrane domains were also a substrate for exog enously added purified casein kinase II, but not casein kinase I. Fina lly, immunoblotting of these domains with an antibody directed against the alpha and alpha' subunits of casein kinase II reveals two bands w ith apparent molecular weights consistent with the known molecular wei ghts of the alpha and alpha' subunits of casein kinase II. As casein k inase II appears to play a role in mitogenic signalling events and cas ein kinase II activators (endogenous polyamines) are required for v-Sr c transformation, our results may have implications for understanding the mechanism of v-Src oncogenesis.