Kotani K, Yonezawa K, Hara K, Ueda H, Kitamura Y, Sakaue H, Ando A, Chavanieu A, Calas B, Grigorescu F
Second Department of Internal Medicine, Kobe University School of Medicine, Japan.
EMBO J. 1994 May 15;13(10):2313-21. doi: 10.1002/j.1460-2075.1994.tb06515.x.
Insulin, IGF-1 or EGF induce membrane ruffling through their respective tyrosine kinase receptors. To elucidate the molecular link between receptor activation and membrane ruffling, we microinjected phosphorylated peptides containing YMXM motifs or a mutant 85 kDa subunit of phosphoinositide (PI) 3-kinase (delta p85) which lacks a binding site for the catalytic 110 kDa subunit of PI 3-kinase into the cytoplasm of human epidermoid carcinoma KB cells. Both inhibited the association of insulin receptor substrate-1 (IRS-1) with PI 3-kinase in a cell-free system and also inhibited insulin- or IGF-1-induced, but not EGF-induced, membrane ruffling in KB cells. Microinjection of nonphosphorylated analogues, phosphorylated peptides containing the EYYE motif or wild-type 85 kDa subunit (Wp85), all of which did not inhibit the association of IRS-1 with PI 3-kinase in a cell-free system, did not inhibit membrane ruffling in KB cells. In addition, wortmannin, an inhibitor of PI 3-kinase activity, inhibited insulin- or IGF-1-induced membrane ruffling. These results suggest that the association of IRS-1 with PI 3-kinase followed by the activation of PI 3-kinase are required for insulin- or IGF-1-induced, but not for EGF-induced, membrane ruffling.
胰岛素、胰岛素样生长因子-1（IGF-1）或表皮生长因子（EGF）通过各自的酪氨酸激酶受体诱导细胞膜褶皱。为了阐明受体激活与细胞膜褶皱之间的分子联系，我们将含有YMXM基序的磷酸化肽或缺乏与磷脂酰肌醇（PI）3激酶催化性110 kDa亚基结合位点的PI 3激酶85 kDa亚基突变体（δ p85）显微注射到人表皮样癌KB细胞的细胞质中。二者在无细胞体系中均抑制胰岛素受体底物-1（IRS-1）与PI 3激酶的结合，且在KB细胞中抑制胰岛素或IGF-1诱导的而非EGF诱导的细胞膜褶皱。显微注射非磷酸化类似物、含有EYYE基序的磷酸化肽或野生型85 kDa亚基（Wp85），在无细胞体系中它们均不抑制IRS-1与PI 3激酶的结合，在KB细胞中也不抑制细胞膜褶皱。此外，PI 3激酶活性抑制剂渥曼青霉素抑制胰岛素或IGF-1诱导的细胞膜褶皱。这些结果表明，胰岛素或IGF-1诱导而非EGF诱导的细胞膜褶皱需要IRS-1与PI 3激酶结合，随后激活PI 3激酶。
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