The first CH domain of affixin activates Cdc42 and Rac1 through alphaPIX, a Cdc42/Rac1-specific guanine nucleotide exchanging factor.
Rho GTPases, Cdc42 and Rac1, play pivotal roles in cell migration by efficiently integrating cell-substrate adhesion and actin polymerization. Although it has been suggested that integrins stimulate these Rho GTPases via some of integrin binding proteins such as focal adhesion kinase (FAK) and paxillin, the precise molecular mechanism is largely unknown. In this study, we showed that the over-expression of RP1 corresponding to the first CH domain (CH1) of affixin, an integrin-linked kinase (ILK)-binding protein, induced a significant actin reorganization in MDCK cells by activating Cdc42/Rac1. Affixin full length and RP1 co-immunoprecipitated with alphaPIX, a Cdc42/Rac1-specific guanine nucleotide exchanging factor (GEF), and they co-localized at the tips of lamellipodia in motile cells. The involvement of alphaPIX in the RP1-induced Cdc42 activation was demonstrated by the significant dominant negative effect of a point mutant of alphaPIX, alphaPIX (L383R, L384S), lacking GEF activity. Our data strongly support that ILK and affixin provide a novel signalling pathway that links integrin signalling to Cdc42/Rac1 activation.
Department of Internal Medicine and Clinical Immunology, Yokohama City University Graduate School of Medicine, 3-9 Fukuura Kanazawa-ku, Yokohama 236-0004 Japan.
Wataru
W
Mishima
Atsushi
A
Suzuki
Satoshi
S
Yamaji
Ryusuke
R
Yoshimi
Atsuhisa
A
Ueda
Takeshi
T
Kaneko
Junko
J
Tanaka
Yoshihiro
Y
Miwa
Shigeo
S
Ohno
Yoshiaki
Y
Ishigatsubo
D000185:Actinin / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D000199:Actins / Q000378:metabolism; D000256:Adenoviridae / Q000235:genetics; D000818:Animals; D001665:Binding Sites; D018797:Cell Cycle Proteins / Q000378:metabolism*; D002460:Cell Line; D002465:Cell Movement / Q000502:physiology; D016764:Cell Polarity / Q000502:physiology; D005822:Genetic Vectors; D020662:Guanine Nucleotide Exchange Factors / Q000378:metabolism*; D006801:Humans; D007120:Immunochemistry; D008840:Microfilament Proteins; D011233:Precipitin Tests; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism; D017434:Protein Structure, Tertiary / Q000235:genetics; D064067:Rho Guanine Nucleotide Exchange Factors; D017384:Sequence Deletion / Q000235:genetics; D015398:Signal Transduction; D020798:Two-Hybrid System Techniques; D020764:cdc42 GTP-Binding Protein / Q000378:metabolism*; D020830:rac1 GTP-Binding Protein / Q000378:metabolism*; D000099200:Scaffold Protein ILK; D000096985:Calponins
D000185:Actinin; D000199:Actins; D018797:Cell Cycle Proteins; D020662:Guanine Nucleotide Exchange Factors; D008840:Microfilament Proteins; D017346:Protein Serine-Threonine Kinases; D064067:Rho Guanine Nucleotide Exchange Factors; D020764:cdc42 GTP-Binding Protein; D020830:rac1 GTP-Binding Protein; D000099200:Scaffold Protein ILK; C486590:PARVA protein, human; D000096985:Calponins; C486591:PARVB protein, human