DOCK2 associates with CrkL and regulates Rac1 in human leukemia cell lines.
The CDM (ced-5 of Caenorhabditis elegans, DOCK180 [downstream of Crk with molecular weight of 180 kDa] of humans, and myoblast city of Drosophila melanogaster) family of proteins has been shown to play a pivotal role in the integrin-mediated signaling pathway under the regulation of an adaptor molecule c-CT10-related kinase II (c-Crk-II) in adherent cells. Recently, hematopoietic cell-specific CDM protein DOCK2 has been shown to be indispensable for lymphocyte migration. However, the regulatory mechanism for DOCK2 is still unknown because DOCK2 lacks a c-Crk-II binding consensus motif. In this study, we demonstrated that DOCK2 bound to CrkL, which is present exclusively in hematopoietic cells both in vivo and in vitro, and we also found that 2 separate regions of DOCK2 contributed to its binding to Src homology 3 (SH3) domain of CrkL. Colocalization of DOCK2 with Crk-like (CrkL) and F-actin was shown by immunocytochemical analysis with the use of Jurkat cells. We also found that CrkL-induced activation of small guanine triphosphatase (GTPase) Rac1 was significantly inhibited by the DOCK2-dCS mutant in 293T cells. Furthermore, the association of DOCK2 and Vav, the guanine-nucleotide exchanging factor (GEF) for Rac1, was demonstrated in Jurkat cells. Finally, the stable expression of DOCK2-dCS mutant in Jurkat cells was shown to reduce cell attachment. These data suggest the presence of a novel protein complex of CrkL, DOCK2, and Vav to regulate Rac1 in leukemia cell lines.
Nishihara
Hiroshi
H
Laboratory of Molecular and Cellular Pathology, Hokkaido University School of Medicine, Sapporo, Japan.
Maeda
Masae
M
Oda
Atsushi
A
Tsuda
Masumi
M
Sawa
Hirofumi
H
Nagashima
Kazuo
K
Tanaka
Shinya
S
D000199:Actins / Q000378:metabolism; D048868:Adaptor Proteins, Signal Transducing; D002352:Carrier Proteins / Q000378:metabolism* / Q000502:physiology; D002448:Cell Adhesion; D020690:GTPase-Activating Proteins; D015972:Gene Expression Regulation, Neoplastic; D020662:Guanine Nucleotide Exchange Factors; D006801:Humans; D007938:Leukemia / Q000209:etiology / Q000378:metabolism / Q000473:pathology*; D014411:Neoplastic Stem Cells / Q000378:metabolism / Q000473:pathology; D009687:Nuclear Proteins / Q000378:metabolism; D015513:Oncogene Proteins / Q000378:metabolism; D011485:Protein Binding; D051567:Proto-Oncogene Proteins c-vav; D014407:Tumor Cells, Cultured; D020830:rac1 GTP-Binding Protein / Q000096:biosynthesis* / Q000235:genetics / Q000378:metabolism* / Q000502:physiology
D000199:Actins; D048868:Adaptor Proteins, Signal Transducing; C082869:CRKL protein; D002352:Carrier Proteins; C402110:DOCK2 protein, human; D020690:GTPase-Activating Proteins; D020662:Guanine Nucleotide Exchange Factors; D009687:Nuclear Proteins; D015513:Oncogene Proteins; D051567:Proto-Oncogene Proteins c-vav; C496618:VAV1 protein, human; D020830:rac1 GTP-Binding Protein