Martin Negin P, Mohney Robert P, Dunn Sara, Das Margaret, Scappini Erica, O'Bryan John P
Laboratory of Signal Transduction, National Institute of Environmental Health Sciences, National Institutes of Health, Research Triangle Park, North Carolina, USA.
Mol Pharmacol. 2006 Nov;70(5):1643-53. doi: 10.1124/mol.106.028274. Epub 2006 Aug 16.
Receptor tyrosine kinases (RTKs) are critical for normal cell growth, differentiation, and development, but they contribute to various pathological conditions when disrupted. Activation of RTKs stimulates a plethora of pathways, including the ubiquitylation and endocytosis of the receptor itself. Although endocytosis terminates RTK signaling, it has emerged as a requisite step in RTK activation of signaling pathways. We have discovered that the endocytic scaffolding protein intersectin (ITSN) cooperated with epidermal growth factor receptor (EGFR) in the regulation of cell growth and signaling. However, a biochemical link between ITSN and EGFR was not defined. In this study, we demonstrate that ITSN is a scaffold for the E3 ubiquitin ligase Cbl. ITSN forms a complex with Cbl in vivo mediated by the Src homology (SH) 3 domains binding to the Pro-rich COOH terminus of Cbl. This interaction stimulates the ubiquitylation and degradation of the activated EGFR. Furthermore, silencing ITSN by RNA interference attenuated EGFR internalization as well as activation of the extracellular signal-regulated kinasemitogen-activated protein kinase pathway, thereby demonstrating the importance of ITSN in EGFR function. Given the cooperativity between ITSN and additional RTKs, these results point to an important evolutionarily conserved, regulatory role for ITSN in RTK function that is necessary for both signaling from receptors as well as the ultimate termination of receptor signaling.
受体酪氨酸激酶（RTKs）对正常细胞生长、分化和发育至关重要，但当它们被破坏时会导致各种病理状况。RTKs的激活会刺激大量信号通路，包括受体自身的泛素化和内吞作用。虽然内吞作用会终止RTK信号传导，但它已成为RTK激活信号通路的一个必要步骤。我们发现内吞支架蛋白相交蛋白（ITSN）与表皮生长因子受体（EGFR）在细胞生长和信号传导的调节中相互协作。然而，ITSN与EGFR之间的生化联系尚未明确。在本研究中，我们证明ITSN是E3泛素连接酶Cbl的一个支架。ITSN在体内与Cbl形成复合物，该复合物由Src同源（SH）3结构域与Cbl富含脯氨酸的COOH末端结合介导。这种相互作用会刺激活化的EGFR的泛素化和降解。此外，通过RNA干扰使ITSN沉默会减弱EGFR的内化以及细胞外信号调节激酶/丝裂原活化蛋白激酶信号通路的激活，从而证明ITSN在EGFR功能中的重要性。鉴于ITSN与其他RTKs之间的协同作用，这些结果表明ITSN在RTK功能中具有重要的进化保守调节作用，这对于受体信号传导以及受体信号的最终终止都是必需的。
