Roles of Necl-5/poliovirus receptor and Rho-associated kinase (ROCK) in the regulation of transformation of integrin alpha(V)beta(3)-based focal complexes into focal adhesions.
Focal complexes are continuously formed and transformed into focal adhesions during cell movement. We previously demonstrated that Necl-5 co-localizes with integrin alpha(V)beta(3) at focal complexes, whereas Necl-5 does not localize at focal adhesions in moving NIH3T3 cells, suggesting that Necl-5 may be dissociated from integrin alpha(V)beta(3) during the transformation of focal complexes into focal adhesions, but the underlying mechanism remains unknown. Here, we explore the roles of Necl-5 and Rho-associated kinase (ROCK) in the regulation of the transformation of focal complexes into focal adhesions. We found that inhibition of Necl-5 expression and expression of a constitutively active mutant of ROCK1 enhanced, whereas treatment with a ROCK inhibitor Y-27632 inhibited the transformation of focal complexes into focal adhesions. In HEK293 cells ectopically expressing Necl-5 and integrin alpha(V)beta(3), treatment of cells with Y-27632 increased the binding of Necl-5 to clustered integrin alpha(V)beta(3). The experiments using inhibitors of myosin ATPase and actin polymerization revealed that actomyosin-driven contractility exerts a similar function as ROCK. The phosphorylation of integrin beta(3) at Tyr(747), which is known to be critical for the formation of focal adhesions, plays a pivotal role for the interaction between Necl-5 and integrin alpha(V)beta(3). These results indicate that the transformation of focal complexes into focal adhesions is negatively and positively regulated by Necl-5 and ROCK, respectively, and that ROCK-dependent actomyosin-driven contractility is a critical determinant for the regulation of the interaction between Necl-5 and integrin alpha(V)beta(3).
Nagamatsu
Yuichi
Y
Division of Molecular and Cellular Biology, Department of Biochemistry and Molecular Biology, Kobe University Graduate School of Medicine, Kobe, Japan.
Rikitake
Yoshiyuki
Y
Takahashi
Motonori
M
Deki
Yuko
Y
Ikeda
Wataru
W
Hirata
Ken-ichi
K
Takai
Yoshimi
Y
D000818:Animals; D002460:Cell Line; D022001:Focal Adhesions / Q000378:metabolism*; D006801:Humans; D039302:Integrin alphaVbeta3 / Q000235:genetics / Q000378:metabolism*; D051379:Mice; D019000:Phosphotyrosine / Q000378:metabolism; D011485:Protein Binding; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D011991:Receptors, Virus / Q000235:genetics / Q000378:metabolism*; D019076:Transgenes / Q000235:genetics; D054460:rho-Associated Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism*
D039302:Integrin alphaVbeta3; D047428:Protein Kinase Inhibitors; D011991:Receptors, Virus; C066100:poliovirus receptor; D019000:Phosphotyrosine; D054460:rho-Associated Kinases