Membrane recruitment of DOCK180 by binding to PtdIns(3,4,5)P3.
DOCK180 was originally identified as one of two major proteins bound to the Crk oncogene product and became an archetype of the CDM family of proteins, including Ced-5 of Caenorhabditis elegans and Mbc of Drosophila melanogaster. Further study has suggested that DOCK180 is involved in the activation of Rac by the CrkII-p130(Cas) complex. With the use of deletion mutants of DOCK180, we found that the C-terminal region containing a cluster of basic amino acids was required for binding to and activation of Rac. This region showed high amino-acid sequence similarity to the consensus sequence of the phosphoinositide-binding site; this led us to examine whether this basic region binds to phosphoinositides. For this purpose we used PtdIns(3,4,5)P(3)-APB beads, as reported previously [Shirai, Tanaka, Terada, Sawada, Shirai, Hashimoto, Nagata, Iwamatsu, Okawa, Li et al. (1998) Biochim. Biophys. Acta 1402, 292-302]. By using various competitors, we demonstrated the specific binding of DOCK180 to PtdIns(3,4,5)P(3). The expression of active phosphoinositide 3-kinase (PI-3K) did not enhance a DOCK180-induced increase in GTP-Rac; however, the expression of PI-3K translocated DOCK180 to the plasma membrane. Thus DOCK180 contained a phosphoinositide-binding domain, as did the other guanine nucleotide exchange factors with a Dbl homology domain, and was translocated to the plasma membrane on the activation of PI-3K.
Kobayashi
S
S
Department of Pathology, Research Institute, International Medical Center of Japan, 1-21-1 Toyama, Shinjuku-ku, Tokyo 162-8655, Japan.
Shirai
T
T
Kiyokawa
E
E
Mochizuki
N
N
Matsuda
M
M
Fukui
Y
Y
D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D006801:Humans; D018129:Phosphatidylinositol Phosphates / Q000378:metabolism*; D011485:Protein Binding; D011506:Proteins / Q000378:metabolism*; D020744:rac GTP-Binding Proteins / Q000378:metabolism
C099818:DOCK1 protein, human; D018129:Phosphatidylinositol Phosphates; D011506:Proteins; C060974:phosphatidylinositol 3,4,5-triphosphate; D020744:rac GTP-Binding Proteins