Qadota Hiroshi, Luo Yating, Matsunaga Yohei, Park Angela S, Gernert Kim M, Benian Guy M
From the Department of Pathology, Emory University, Atlanta, Georgia 30322 and.
the Biomolecular Computing Resource, Emory University, Atlanta, Georgia 30322.
J Biol Chem. 2014 May 16;289(20):14252-62. doi: 10.1074/jbc.M114.556308. Epub 2014 Apr 1.
Caenorhabditis elegans striated muscle cells attach to basement membrane and transmit the force of muscle contraction through integrin adhesion complexes. The cytoplasmic tail of β-integrin (PAT-3) is associated with a conserved four-protein complex that includes UNC-112 (kindlin), PAT-4 (integrin-linked kinase), PAT-6 (α-parvin/actopaxin), and UNC-97 (PINCH). The proper localization of UNC-112 to muscle integrin adhesion sites requires PAT-4. A recent report (Qadota, H., Moerman, D. G., and Benian, G. M. (2012) A molecular mechanism for the requirement of PAT-4 (integrin-linked kinase (ILK)) for the localization of UNC-112 (kindlin) to integrin adhesion sites. J. Biol. Chem. 287, 28537-28551) suggests a possible molecular mechanism for this requirement: that UNC-112 exists in closed inactive and open active conformations, and conversion to the open active form is promoted by binding to PAT-4 (ILK). Previously, we also reported identification of a single missense mutation in UNC-112, D382V, which abolishes both binding to PAT-4 and normal localization to integrin adhesion sites in vivo. In this report, we describe isolation and characterization of PAT-4 missense mutations that permit binding with UNC-112 D382V and place nine affected residues on a homology model of PAT-4. These nine residues cluster in two regions on the surface of PAT-4, do not overlap the likely binding surface for PAT-6 (α-parvin), and therefore may reside along the interaction surface of PAT-4 for UNC-112 (kindlin). We also show that one of these PAT-4 mutations restores the ability of UNC-112 D382V to localize to integrin adhesions and participate in complex formation.
秀丽隐杆线虫的横纹肌细胞附着于基底膜,并通过整合素黏附复合体传递肌肉收缩力。β-整合素(PAT-3)的胞质尾部与一个保守的四蛋白复合体相关联,该复合体包括UNC-112(纽蛋白)、PAT-4(整合素连接激酶)、PAT-6(α-帕文/肌动蛋白斑蛋白)和UNC-97(PINCH)。UNC-112在肌肉整合素黏附位点的正确定位需要PAT-4。最近的一份报告(Qadota, H., Moerman, D. G., and Benian, G. M. (2012) A molecular mechanism for the requirement of PAT-4 (integrin-linked kinase (ILK)) for the localization of UNC-112 (kindlin) to integrin adhesion sites. J. Biol. Chem. 287, 28537-28551)提出了这一需求的一种可能分子机制:UNC-112以封闭的无活性构象和开放的活性构象存在,与PAT-4(整合素连接激酶)结合可促进其转变为开放的活性形式。此前,我们还报告了在UNC-112中鉴定出一个单错义突变D382V,该突变消除了其在体内与PAT-4的结合以及在整合素黏附位点的正常定位。在本报告中,我们描述了PAT-4错义突变的分离和特性,这些突变允许与UNC-112 D382V结合,并在PAT-4的同源模型上确定了九个受影响的残基。这九个残基聚集在PAT-4表面的两个区域,不与PAT-6(α-帕文)可能的结合表面重叠,因此可能位于PAT-4与UNC-112(纽蛋白)的相互作用表面上。我们还表明,这些PAT-4突变之一恢复了UNC-112 D382V定位于整合素黏附位点并参与复合体形成的能力。