Open Accessβ2-Chimaerin is a novel target for diacylglycerol: Binding properties and changes in subcellular localization mediated by ligand binding to its C1 domain
Author(s) -
Marı́a J. Caloca,
Laura GarcíaBermejo,
Peter M. Blumberg,
Nancy E. Lewin,
Elisabeth Kremmer,
Harald Mischak,
Shaomeng Wang,
Kassoum Nacro,
Bruno Bienfait,
Víctor E. Márquez,
Marcelo G. Kazanietz
Publication year - 1999
Publication title -
proceedings of the national academy of sciences of the united states of america
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 5.011
H-Index - 771
eISSN - 1091-6490
pISSN - 0027-8424
DOI - 10.1073/pnas.96.21.11854
Subject(s) - diacylglycerol kinase , protein kinase c , biochemistry , biology , receptor , binding domain , fusion protein , c2 domain , phorbol , binding site , signal transduction , microbiology and biotechnology , chemistry , gene , recombinant dna , membrane
The members of the chimaerin family of Rac-GTPase-activating proteins possess a single C1 domain with high homology to those present in protein kinase C (PKC) isozymes. This domain in PKCs is involved in phorbol ester and diacylglycerol (DAG) binding. We previously have demonstrated that one of the chimaerin isoforms, β2-chimaerin, binds phorbol esters with high affinity. In this study we analyzed the properties of β2-chimaerin as a DAG receptor by using a series of conformationally constrained cyclic DAG analogues (DAG lactones) as probes. We identified analogs that bind to β2-chimaerin with more than 100-fold higher affinity than 1-oleoyl-2-acetylglycerol. The potencies of these analogs approach those of the potent phorbol ester tumor promoters. The different DAG lactones show some selectivity for this novel receptor compared with PKCα. Cellular studies revealed that these DAG analogs induce translocation of β2-chimaerin from cytosolic (soluble) to particulate fractions. Using green fluorescent protein-fusion proteins for β2-chimaerin we determined that this novel receptor translocates to the perinuclear region after treatment with DAG lactones. Binding and translocation were prevented by mutation of the conserved Cys-246 in the C1 domain. The structural homology between the C1 domain of β2-chimaerin and the C1b domain of PKCδ also was confirmed by modeling analysis. Our results demonstrate that β2-chimaerin is a high affinity receptor for DAG through binding to its C1 domain and supports the emerging concept that multiple pathways transduce signaling through DAG and the phorbol esters.