title: Immunophilins control T lymphocyte adhesion and migration by regulating CrkII binding to C3G.
abstract: Crk adaptor proteins are key players in signal transduction from a variety of cell surface receptors. CrkI and CrkII, the two alternative spliced forms of CRK, possess an N-terminal Src homology 2 domain, followed by a Src homology 3 (SH3) domain, whereas CrkII possesses in addition a C-terminal linker region plus a SH3 domain, which operate as regulatory moieties. In this study, we investigated the ability of immunophilins, which function as peptidyl-prolyl isomerases, to regulate Crk proteins in human T lymphocytes. We found that endogenous CrkII, but not CrkI, associates with the immunophilins, cyclophilin A, and 12-kDa FK506-binding protein, in resting human Jurkat T cells. In addition, cyclophilin A increased Crk SH3 domain-binding guanine-nucleotide releasing factor (C3G) binding to CrkII, whereas inhibitors of immunophilins, such as cyclosporine A (CsA) and FK506, inhibited CrkII, but not CrkI association with C3G. Expression in Jurkat T cells of phosphorylation indicator of Crk chimeric unit plasmid, a plasmid encoding the human CrkII1-236 sandwiched between cyan fluorescent protein and yellow fluorescent protein, demonstrated a basal level of fluorescence resonance energy transfer, which increased in response to cell treatment with CsA and FK506, reflecting increased trans-to-cis conversion of CrkII. Crk-C3G complexes are known to play an important role in integrin-mediated cell adhesion and migration. We found that overexpression of CrkI or CrkII increased adhesion and migration of Jurkat T cells. However, immunophilin inhibitors suppressed the ability of CrkII- but not CrkI-overexpressing cells to adhere to fibronectin-coated surfaces and migrate toward the stromal cell-derived factor 1α chemokine. The present data demonstrate that immunophilins regulate CrkII, but not CrkI activity in T cells and suggest that CsA and FK506 inhibit selected effector T cell functions via a CrkII-dependent mechanism.
authors.affiliation: Shraga Segal Department of Microbiology, Immunology, and Genetics, Faculty of Health Sciences, Ben Gurion University of the Negev, Beer Sheva 84105, Israel.
authors.forename: Pulak Ranjan
authors.initials: PR
authors.lastname: Nath
authors.affiliation: Shraga Segal Department of Microbiology, Immunology, and Genetics, Faculty of Health Sciences, Ben Gurion University of the Negev, Beer Sheva 84105, Israel.
authors.forename: Guangyu
authors.initials: G
authors.lastname: Dong
authors.affiliation: Shraga Segal Department of Microbiology, Immunology, and Genetics, Faculty of Health Sciences, Ben Gurion University of the Negev, Beer Sheva 84105, Israel.
authors.forename: Alex
authors.initials: A
authors.lastname: Braiman
authors.affiliation: Shraga Segal Department of Microbiology, Immunology, and Genetics, Faculty of Health Sciences, Ben Gurion University of the Negev, Beer Sheva 84105, Israel noah@bgu.ac.il.
authors.forename: Noah
authors.initials: N
authors.lastname: Isakov
mesh_terms: D002448:Cell Adhesion / Q000276:immunology; D002465:Cell Movement / Q000276:immunology; D054377:Chemokine CXCL12 / Q000276:immunology; D021984:Cyclophilin A / Q000037:antagonists & inhibitors* / Q000378:metabolism; D016572:Cyclosporine / Q000494:pharmacology; D020735:Guanine Nucleotide-Releasing Factor 2 / Q000378:metabolism*; D006801:Humans; D019169:Jurkat Cells; D008213:Lymphocyte Activation / Q000187:drug effects / Q000276:immunology; D010957:Plasmids / Q000235:genetics; D011485:Protein Binding / Q000187:drug effects; D017434:Protein Structure, Tertiary; D050737:Proto-Oncogene Proteins c-crk / Q000378:metabolism*; D015398:Signal Transduction / Q000276:immunology; D013601:T-Lymphocytes / Q000276:immunology*; D016559:Tacrolimus / Q000494:pharmacology; D018909:src Homology Domains / Q000235:genetics
chemical_list: C485285:CRK protein, human; D054377:Chemokine CXCL12; D020735:Guanine Nucleotide-Releasing Factor 2; D050737:Proto-Oncogene Proteins c-crk; D016572:Cyclosporine; D021984:Cyclophilin A; D016559:Tacrolimus