A novel protein distinguishes between quiescent and activated forms of the type I transforming growth factor β receptor
A novel protein distinguishes between quiescent and activated forms of the type I transforming growth factor β receptor
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DOI：
10.1074/jbc.273.16.9365
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发表时间：
1998-04-17
影响因子：
4.8
通讯作者：
Schneider, MD
中科院分区：
文献类型：
--
作者：
Charng, MJ;Zhang, D;Schneider, MD
转化生长因子-β(TGF-β)信号转导是由两个受体丝氨酸/苏氨酸(Ser/Thr)受体串联作用介导的，即II型转化生长因子β受体(TβR-II)磷酸化I型转化生长因子β受体(TβR-I)。由于到目前为止，相互作用克隆的失败可能是由于使用非活性TβR-I作为诱饵，因此我们试图通过在酵母双杂交系统中使用三重突变L193A、P194A和T204D来识别与活性TβR-I特异相互作用的分子。Leu-Pro取代阻止了与FK506结合蛋白12(FKBP12)的相互作用，我们先前已经证明FKBP12在转化生长因子β信号转导中的作用是错误的；Thr(204)上的电荷取代结构性地激活TβR-I。与以前使用轻型型TβR-I的筛查不同，FKBP12占主导地位，产生的菌落都没有编码FKBP12。发现了一种新的蛋白质，TβR-I相关蛋白-1(TRAP-1)，它在酵母中与突变激活的TβR-I特异地相互作用，但不与野生型TβR-I、TβR-II或无关蛋白相互作用。在哺乳动物细胞中，TRAP-1只与突变激活的TβR-I和配体激活的TβR-I共沉淀，而野生型TβR-I在没有转化生长因子β的情况下不能共沉淀。部分TRAP-I蛋白特异性地结合这些突变和配体激活的TβR-I农场，可以抑制天然受体在转化生长因子β刺激后的信号传递，或通过依赖转化生长因子β的报告基因检测到的结构性激活的受体突变。因此，TRAP-1可以在缺乏转化生长因子β的情况下区分激活形式的受体和野生型受体，并可能在转化生长因子β信号转导中发挥作用。
Transforming growth factor beta (TGF beta) signal transduction is mediated by two receptor Ser/Thr kinases acting in series, type II TGF beta receptor (T beta R-II) phosphorylating type I TGF beta receptor (T beta R-I). Because the failure of interaction cloning, thus far, to identify bona fide T beta R-I substrates might reasonably have been due to the use of inactive T beta R-I as bait, we sought to identify molecules that interact specifically with active T beta R-I, employing the triple mutation L193A,P194A,T204D in a yeast two-hybrid system. The Leu-Pro substitutions prevent interaction with FK506-binding protein 12 (FKBP12), whose putative function in TGF beta signaling we have previously disproved; the charge substitution at Thr(204) constitutively activates T beta R-I. Unlike previous screens using mild-type T beta R-I, where FKBP12 predominated, none of the resulting colonies encoded FKBP12. A novel protein was identified, T beta R-I-associated protein-1 (TRAP-1), that interacts in yeast specifically with mutationally activated T beta R-I, but not wild-type T beta R-I, T beta R-II, or irrelevant proteins. In mammalian cells, TRAP-1 was co-precipitated only by mutationally activated T beta R-I and ligand-activated T beta R-I, but not wild-type T beta R-I in the absence of TGF beta. The partial TRAP-I protein that specifically binds these mutationally and ligand-activated farms of T beta R-I can inhibit signaling by the native receptor after stimulation with TGF beta or by the constitutively activated receptor mutation, as measured by a TGF beta-dependent reporter gene. Thus, TRAP-1 can distinguish activated forms of the receptor from wild-type receptor in the absence of TGF beta and may potentially have a functional role in TGF beta signaling.