Pratt J C, Weiss M, Sieff C A, Shoelson S E, Burakoff S J, Ravichandran K S
Division of Pediatric Oncology, Dana-Farber Cancer Institute, Boston, Massachusetts, USA.
J Biol Chem. 1996 May 24;271(21):12137-40. doi: 10.1074/jbc.271.21.12137.
Granulocyte-macrophage colony-stimulating factor (GM-CSF) regulates the growth and function of several myeloid cell types at different stages of maturation. The effects of GM-CSF are mediated through a high affinity receptor that is composed of two chains: a unique, ligand-specific alpha chain and a beta common chain (beta c) that is also a component of the receptors for interleukin 3 (IL-3) and IL-5. Beta c plays an essential role in the transduction of extra cellular signals to the nucleus through its recruitment of secondary messengers. Several downstream signaling events induced by GM-CSF stimulation have been described, including activation of tyrosine kinases and tyrosine phosphorylation of cellular proteins (including beta c) and activation of the Ras/mitogen-activated protein kinase and the JAK/STAT pathways. A region within the beta c cytoplasmic tail (amino acids 517-763) has been reported to be necessary for tyrosine phosphorylation of the adapter protein, Shc, and for the subsequent GM-CSF-induced activation of Ras. In this paper, we describe a physical association between the tyrosine phosphorylated GM-CSF receptor (GMR)-beta c chain and Shc in vivo. Using a series of cytoplasmic truncation mutants of beta c and various mutant Shc proteins, we demonstrate that the N-terminal phosphotyrosine-binding (PTB) domain of Shc binds to a short region of beta c (amino acids 549-656) that contains Tyr577. Addition of a specific phosphopeptide encoding amino acids surrounding this tyrosine inhibited the interaction between beta c and shc. Moreover, mutation of a key residue within the phosphotyrosine binding pocket of the Shc-PTB domain abrogated its association with beta c. These observations provide an explanation for the previously described requirement for Tyr577 of beta c for GM-CSF-induced tyrosine phosphorylation of Shc and have implications for Ras activation through the GM-CSF, IL-3, and IL-5 receptors.
粒细胞-巨噬细胞集落刺激因子（GM-CSF）在不同成熟阶段调节多种髓系细胞类型的生长和功能。GM-CSF的作用是通过一种高亲和力受体介导的，该受体由两条链组成：一条独特的、配体特异性的α链和一条β共同链（βc），βc也是白细胞介素3（IL-3）和IL-5受体的组成部分。βc通过募集二级信使在将细胞外信号转导至细胞核的过程中起重要作用。已经描述了GM-CSF刺激诱导的几种下游信号事件，包括酪氨酸激酶的激活、细胞蛋白（包括βc）的酪氨酸磷酸化以及Ras/丝裂原活化蛋白激酶和JAK/STAT途径的激活。据报道，βc细胞质尾部的一个区域（氨基酸517 - 763）对于衔接蛋白Shc的酪氨酸磷酸化以及随后GM-CSF诱导的Ras激活是必需的。在本文中，我们描述了体内酪氨酸磷酸化的GM-CSF受体（GMR）-βc链与Shc之间的物理关联。使用一系列βc的细胞质截短突变体和各种突变的Shc蛋白，我们证明Shc的N端磷酸酪氨酸结合（PTB）结构域与βc的一个短区域（氨基酸549 - 656）结合，该区域包含Tyr577。添加编码该酪氨酸周围氨基酸的特定磷酸肽可抑制βc与Shc之间的相互作用。此外，Shc-PTB结构域的磷酸酪氨酸结合口袋内一个关键残基的突变消除了其与βc的关联。这些观察结果为先前描述的βc的Tyr577对于GM-CSF诱导的Shc酪氨酸磷酸化所必需的现象提供了解释，并对通过GM-CSF、IL-3和IL-5受体激活Ras具有启示意义。
