Integrin-linked kinase associated with integrin activation.
Platelet integrin alphaIIbbeta3 activation is tightly controlled by intracellular signaling pathways, and several molecules, including talin, have been identified as critical for alphaIIbbeta3 activation. However, the whole pathway associated with alphaIIbbeta3 activation remains to be determined. To address this issue, we established a Chinese hamster ovary cell line (parental cells) that expresses constitutively activated chimeric integrin alphaIIbalpha6Bbeta3, and then obtained mutant cells expressing inactivated alphaIIbalpha6Bbeta3 by genome-wide mutagenesis. We have performed expression cloning to isolate signaling molecules responsible for integrin activation in the mutant cells. We show that integrin-linked kinase (ILK) complements defective integrin activation in the mutant cells. ILK mRNAs in the mutant cells contained 2 nonsense mutations, R317X and W383X, in a compound heterozygous state, resulting in a complete loss of ILK expression. Moreover, the mutant cells showed partially impaired activation of endogenous beta1 integrins. Knockdown of ILK in parental cells significantly suppressed the activated state of alphaIIbalpha6Bbeta3. However, ILK overexpression did not rescue the impaired integrin activation in talin knocked-down parental cells, whereas overexpression of talin-F3, a subdomain of the talin head domain, restored the function. Our present data suggest that ILK contributes to inside-out integrin activation.
Honda
Shigenori
S
National Cardiovascular Center Research Institute, Osaka, Japan. shige@ri.ncvc.go.jp
Shirotani-Ikejima
Hiroko
H
Tadokoro
Seiji
S
Maeda
Yusuke
Y
Kinoshita
Taroh
T
Tomiyama
Yoshiaki
Y
Miyata
Toshiyuki
T
D000818:Animals; D016466:CHO Cells; D018389:Codon, Nonsense; D006224:Cricetinae; D003412:Cricetulus; D019012:Integrin beta1 / Q000378:metabolism; D016023:Integrins / Q000378:metabolism*; D016296:Mutagenesis; D019039:Platelet Glycoprotein GPIIb-IIIa Complex / Q000235:genetics / Q000378:metabolism*; D017346:Protein Serine-Threonine Kinases / Q000235:genetics / Q000378:metabolism*; D012333:RNA, Messenger / Q000235:genetics; D015398:Signal Transduction; D016608:Talin; D000099200:Scaffold Protein ILK
D018389:Codon, Nonsense; D019012:Integrin beta1; D016023:Integrins; D019039:Platelet Glycoprotein GPIIb-IIIa Complex; D017346:Protein Serine-Threonine Kinases; D012333:RNA, Messenger; D016608:Talin; D000099200:Scaffold Protein ILK