MOB1 Mediated Phospho-recognition in the Core Mammalian Hippo Pathway.
MOB1 Mediated Phospho-recognition in the Core Mammalian Hippo Pathway.
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DOI：
10.1074/mcp.m116.065490
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发表时间：
2017-06
期刊：
影响因子：
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通讯作者：
Gingras AC
中科院分区：
文献类型：
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作者：
Couzens AL;Xiong S;Knight JDR;Mao DY;Guettler S;Picaud S;Kurinov I;Filippakopoulos P;Sicheri F;Gingras AC
Hippo肿瘤抑制通路根据不同的信号输入调节器官大小和组织平衡。该通路的核心由一个短激酶级联组成：MST1和MST2磷酸化并激活LATS1和LATS2，而LATS1和LATS2反过来磷酸化并灭活关键的转录共激活因子YAP1和TAZ（基因WWTR1）。MOB1适配蛋白首先通过与上游MST和下游LATS激酶同时结合实现反式磷酸化反应，其次通过变速激活LATS1和LATS2的催化功能直接刺激YAP和TAZ的磷酸化，从而调控两种磷酸化反应。对酵母Mob1和人类Mob1的研究表明，在它们的相互作用物Nud1和MST2中分别识别磷酸肽序列的能力，对于它们调节酵母有丝分裂退出网络和后生动物Hippo通路的作用至关重要。然而，人类MOB1识别磷酸肽的基本规律、Hippo通路信号的结合特异性以及磷酸肽与其他人类MOB家族成员的结合功能的普遍性仍然是未知的。通过蛋白质组学、肽阵列和生化分析，我们系统地检测了MOB1的磷酸化肽结合特异性，发现它与MST1和MST2的底物磷酸化特异性高度互补。我们证明了MST1和MST2在几个苏氨酸残基上的自磷酸化提供了多个具有不同结合亲和力的MOB1结合位点，这反过来又有助于细胞中MST1-MOB1蛋白相互作用的冗余。MOB1A与MST1中两个有利的磷酸肽位点的复合物的晶体结构允许对MOB1A磷酸肽结合的共识进行完整的描述。最后，我们发现MOB1A的磷酸肽结合特性在人类七种MOB家族成员中除一种外都是保守的，从而为揭示其难以捉摸的细胞功能提供了一个起点。
The Hippo tumor suppressor pathway regulates organ size and tissue homoeostasis in response to diverse signaling inputs. The core of the pathway consists of a short kinase cascade: MST1 and MST2 phosphorylate and activate LATS1 and LATS2, which in turn phosphorylate and inactivate key transcriptional coactivators, YAP1 and TAZ (gene WWTR1). The MOB1 adapter protein regulates both phosphorylation reactions firstly by concurrently binding to the upstream MST and downstream LATS kinases to enable the trans phosphorylation reaction, and secondly by allosterically activating the catalytic function of LATS1 and LATS2 to directly stimulate phosphorylation of YAP and TAZ. Studies of yeast Mob1 and human MOB1 revealed that the ability to recognize phosphopeptide sequences in their interactors, Nud1 and MST2 respectively, was critical to their roles in regulating the Mitotic Exit Network in yeast and the Hippo pathway in metazoans. However, the underlying rules of phosphopeptide recognition by human MOB1, the implications of binding specificity for Hippo pathway signaling, and the generality of phosphopeptide binding function to other human MOB family members remained elusive. Employing proteomics, peptide arrays and biochemical analyses, we systematically examine the phosphopeptide binding specificity of MOB1 and find it to be highly complementary to the substrate phosphorylation specificity of MST1 and MST2. We demonstrate that autophosphorylation of MST1 and MST2 on several threonine residues provides multiple MOB1 binding sites with varying binding affinities which in turn contribute to a redundancy of MST1-MOB1 protein interactions in cells. The crystal structures of MOB1A in complex with two favored phosphopeptide sites in MST1 allow for a full description of the MOB1A phosphopeptide-binding consensus. Lastly, we show that the phosphopeptide binding properties of MOB1A are conserved in all but one of the seven MOB family members in humans, thus providing a starting point for uncovering their elusive cellular functions.