Kavanaugh W M, Pot D A, Chin S M, Deuter-Reinhard M, Jefferson A B, Norris F A, Masiarz F R, Cousens L S, Majerus P W, Williams L T
Chiron Corporation, Emeryville, California 94608, USA.
Curr Biol. 1996 Apr 1;6(4):438-45. doi: 10.1016/s0960-9822(02)00511-0.
Shc and Grb2 form a complex in cells in response to growth factor stimulation and link tyrosine kinases to Ras during the resulting signaling process. Shc and Grb2 each contain domains that mediate interactions with other unidentified intracellular proteins. For example, the Shc PTB domain binds to 130 kDa and 145 kDa tyrosine-phosphorylated proteins in response to stimulation of cells by growth factors, cytokines and crosslinking of antigen receptors. The Grb2 SH3 domains bind to an unidentified 116 kDa protein in T cells. We have identified three proteins, of 110 kDa, 130 kDa and 145 kDa, as a new family of molecules encoded by the same gene. In vivo studies show that these proteins form signal transduction complexes with Shc and with Grb2.
The 130 kDa and 145 kDa tyrosine-phosphorylated proteins that associate with the Shc PTB domain were purified by conventional chromatographic methods. Partial peptide and cDNA sequences corresponding to these proteins, termed SIP-145 and SIP-130 (SIP for signaling inositol polyphosphate 5-phosphatase), identified them as SH2 domain-containing products of a single gene and as members of the inositol polyphosphate 5-phosphatase family. The SIP-130 and SIP-145 proteins and inositol polyphosphate 5-phosphatase activity associated with Shc in vivo in response to B-cell activation. By using an independent approach, expression cloning, we found that the Grb2 SH3 domains bind specifically to SIP-110, a 110 kDa splice variant of SIP-145 and SIP-130, which lacks the SH2 domain. The SIP proteins hydrolyzed phosphatidylinositol (3,4,5)-trisphosphate (PtdIns (3,4,5)-P3) and Ins (1,3,4,5)-P4, but not PtdIns (4,5)-P2 or Ins (1,4,5)-P3.
These findings strongly implicate the inositol polyphosphate 5-phosphatases in Shc- and Grb2-mediated signal transduction. Furthermore, SIP-110, SIP-130 and SIP-145 prefer 3-phosphorylated substrates, suggesting a link to the phosphatidylinositol 3-kinase signaling pathway.
在细胞中，Shc和Grb2会因生长因子刺激而形成复合物，并在随后的信号传导过程中将酪氨酸激酶与Ras连接起来。Shc和Grb2各自包含介导与其他未鉴定的细胞内蛋白质相互作用的结构域。例如，Shc的PTB结构域在生长因子、细胞因子刺激细胞以及抗原受体交联后，会与130 kDa和145 kDa的酪氨酸磷酸化蛋白结合。Grb2的SH3结构域在T细胞中与一种未鉴定的116 kDa蛋白结合。我们已鉴定出三种蛋白，分子量分别为110 kDa、130 kDa和145 kDa，它们是由同一基因编码的一个新分子家族。体内研究表明，这些蛋白与Shc和Grb2形成信号转导复合物。
通过传统色谱方法纯化了与Shc的PTB结构域相关的130 kDa和145 kDa酪氨酸磷酸化蛋白。与这些蛋白对应的部分肽段和cDNA序列，分别称为SIP - 145和SIP - 130（SIP代表信号传导性肌醇多磷酸5 - 磷酸酶），将它们鉴定为一个单基因的含SH2结构域的产物，以及肌醇多磷酸5 - 磷酸酶家族的成员。SIP - 130和SIP - 145蛋白以及肌醇多磷酸5 - 磷酸酶活性在体内响应B细胞活化时与Shc相关联。通过一种独立的方法，即表达克隆，我们发现Grb2的SH3结构域特异性地与SIP - 110结合，SIP - 110是SIP - 145和SIP - 130的一种110 kDa剪接变体，它缺乏SH2结构域。SIP蛋白可水解磷脂酰肌醇(3,4,5) - 三磷酸（PtdIns(3,4,5) - P3）和Ins(1,3,4,5) - P4，但不能水解PtdIns(4,5) - P2或Ins(1,4,5) - P3。
这些发现有力地表明肌醇多磷酸5 - 磷酸酶参与了Shc和Grb2介导的信号转导。此外，SIP - 110、SIP - 130和SIP - 145更倾向于3 - 磷酸化底物，这表明它们与磷脂酰肌醇3 - 激酶信号通路存在联系。
