p85 beta-PIX is required for cell motility through phosphorylations of focal adhesion kinase and p38 MAP kinase.
Lysophosphatidic acid (LPA) mediates diverse biological responses, including cell migration, through the activation of G-protein-coupled receptors. Recently, we have shown that LPA stimulates p21-activated kinase (PAK) that is critical for focal adhesion kinase (FAK) phosphorylation and cell motility. Here, we provide the direct evidence that p85 beta-PIX is required for cell motility of NIH-3T3 cells by LPA through FAK and p38 MAP kinase phosphorylations. LPA induced p85 beta-PIX binding to FAK in NIH-3T3 cells that was inhibited by pretreatment of the cells with phosphoinositide 3-kinase inhibitor, LY294002. Furthermore, the similar inhibition of the complex formation was also observed, when the cells were transfected with either p85 beta-PIX mutant that cannot bind GIT or dominant negative mutants of Rac1 (N17Rac1) and PAK (PAK-PID). Transfection of the cells with specific p85 beta-PIX siRNA led to drastic inhibition of LPA-induced FAK phosphorylation, peripheral redistribution of p85 beta-PIX with FAK and GIT1, and cell motility. p85 beta-PIX was also required for p38 MAP kinase phosphorylation induced by LPA. Finally, dominant negative mutant of Rho (N19Rho)-transfected cells did not affect PAK activation, while the cells stably transfected with p85 beta-PIX siRNA or N17Rac1 showed the reduction of LPA-induced PAK activation. Taken together, the present data suggest that p85 beta-PIX, located downstream of Rac1, is a key regulator for the activations of FAK or p38 MAP kinase and plays a pivotal role in focal complex formation and cell motility induced by LPA.
Lee
Jangsoon
J
Department of Pharmacology, College of Medicine, Konyang University, Nonsan, 320-711, Republic of Korea.
Jung
In Duk
ID
Chang
Won Keun
WK
Park
Chang Gyo
CG
Cho
Do Yeun
DY
Shin
Eun-Young
EY
Seo
Dong Wan
DW
Kim
Yong Kee
YK
Lee
Hyang Woo
HW
Han
Jeung-Whan
JW
Lee
Hoi Young
HY
D000818:Animals; D018797:Cell Cycle Proteins / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D002462:Cell Membrane / Q000378:metabolism; D002465:Cell Movement / Q000187:drug effects / Q000502:physiology*; D003598:Cytoskeletal Proteins / Q000378:metabolism; D051417:Focal Adhesion Kinase 1; D051416:Focal Adhesion Protein-Tyrosine Kinases; D020690:GTPase-Activating Proteins; D020662:Guanine Nucleotide Exchange Factors / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D007155:Immunologic Factors / Q000494:pharmacology; D008246:Lysophospholipids / Q000494:pharmacology; D051379:Mice; D009154:Mutation / Q000235:genetics; D041681:NIH 3T3 Cells; D009479:Neuropeptides / Q000378:metabolism; D051419:Paxillin; D010750:Phosphoproteins / Q000378:metabolism; D010766:Phosphorylation / Q000187:drug effects; D011485:Protein Binding / Q000187:drug effects; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism; D011505:Protein-Tyrosine Kinases / Q000378:metabolism*; D064067:Rho Guanine Nucleotide Exchange Factors; D014162:Transfection; D014443:Tyrosine / Q000378:metabolism; D054462:p21-Activated Kinases; D048051:p38 Mitogen-Activated Protein Kinases / Q000378:metabolism*; D020744:rac GTP-Binding Proteins / Q000378:metabolism; D020830:rac1 GTP-Binding Protein
D018797:Cell Cycle Proteins; D003598:Cytoskeletal Proteins; D020690:GTPase-Activating Proteins; C120766:Git1 protein, mouse; D020662:Guanine Nucleotide Exchange Factors; D007155:Immunologic Factors; D008246:Lysophospholipids; D009479:Neuropeptides; D051419:Paxillin; D010750:Phosphoproteins; D047428:Protein Kinase Inhibitors; C496062:Pxn protein, mouse; C484602:Rac1 protein, mouse; D064067:Rho Guanine Nucleotide Exchange Factors; D014443:Tyrosine; D011505:Protein-Tyrosine Kinases; D051417:Focal Adhesion Kinase 1; D051416:Focal Adhesion Protein-Tyrosine Kinases; C496051:Ptk2 protein, mouse; C515905:Pak1 protein, mouse; D017346:Protein Serine-Threonine Kinases; D054462:p21-Activated Kinases; D048051:p38 Mitogen-Activated Protein Kinases; D020744:rac GTP-Binding Proteins; D020830:rac1 GTP-Binding Protein; C032881:lysophosphatidic acid