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Structural characterization of the PtdIns(4,5)P 2 ‐PKCα‐C2 domain interaction
Author(s) -
CorbalanGarcia Senena,
GuerreroValero Marta,
FerrerOrta Cristina,
QuerolAudi Jordi,
MarinVicente Consuelo,
GomezFernandez Juan C,
Fita Ignacio,
Verdaguer Nuria
Publication year - 2009
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.23.1_supplement.520.5
Subject(s) - diacylglycerol kinase , c2 domain , microbiology and biotechnology , chemistry , vesicle , phosphatidylserine , protein kinase c , endoplasmic reticulum , phosphotyrosine binding domain , mutant , biochemistry , biophysics , membrane , biology , phosphorylation , phospholipid , sh2 domain , gene , receptor tyrosine kinase
Classical PKCs contain two structural modules, the C1 and the C2 domains, in their regulatory region. These domains, in turn, dictate the cofactor‐dependence of the isoenzymes, being the C1 domain the diacylglycerol sensor and the C2 domain the Ca 2+ and acidic phospholipids sensor. Here we show that a polybasic region located in the β3‐β4 strands of the C2 domain of classical PKCs, interacts specifically with PtdIns(4,5)P 2 rather than with other polyphosphoinositides or phosphatidylserine. We have used X‐ray crystallography to analyze the structure of the C2 domain of PKCα in complex with PtdIns(4,5)P 2 , which was found at the conserved polybasic cluster. Site‐directed mutagenesis, on the residues from this cluster that interact directly with the phosphoinositide, revealed an important decrease in the ability of PKCα to interact with the plasma membrane of living cells. Furthermore, the mutant proteins were not able to translocate to the plasma membrane when the cells were stimulated with ionomycin and diacylglycerol. Instead, the protein localized in vesicles and Golgi apparatus, suggesting that the residues mutated are involved in recognizing the proper target of the C2 domain in the plasma membrane. Together, these data reveal the importance of the PtdIns(4,5)P 2 ‐C2 domain interaction for the plasma membrane localization of PKCα.