Loss of mXinα, an intercalated disk protein, results in cardiac hypertrophy and cardiomyopathy with conduction defects
Loss of mXinα, an intercalated disk protein, results in cardiac hypertrophy and cardiomyopathy with conduction defects
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DOI：
10.1074/jbc.m707639200
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发表时间：
2007-12-07
影响因子：
4.8
通讯作者：
Lin, Jim J. -C.
中科院分区：
文献类型：
--
作者：
Choi, Sunju;Gustafson-Wagner, Elisabeth A.;Lin, Jim J. -C.
靶向mXin α缺失导致心肌肥厚和心肌病伴传导缺陷(Gustafson-Wagner， E., Sinn, h.w., Chen, y.l。王德志。，瑞特，r.s.，林，j.l.c。杨,B。威廉姆森,r, Chen J.N。林,我。，林，j.j.c。（2007）[j] .中国生物医学工程学报，2009,33(2):444 - 444。为了了解导致这种心脏缺陷的潜在机制，研究了mXin α及其相互作用蛋白的功能域。利用共免疫沉淀、拉下和酵母双杂交实验进行的相互作用研究表明，mXin α直接与β -连环蛋白相互作用。mXin α上的β -连环蛋白结合位点被定位在535 - 636氨基酸上，这与由Xin重复序列组成的已知肌动蛋白结合域重叠。这些结构域的重叠性质提供了对mXin α定位和功能的分子机制的深入了解。经共沉淀和电镜研究证实，纯化的重组谷胱甘肽s-转移酶或his标记的mXin α蛋白能够结合和捆绑肌动蛋白细丝。与肌动蛋白的结合在近似的9个肌动蛋白单体与1个mXin α的化学计量下饱和。与全长mXin α与肌动蛋白的相互作用相比，mXin α c端缺失与肌动蛋白的相互作用更强。此外，在培养细胞中，与力表达的GFP-mXin α相比，与mXin α c端缺失融合的绿色荧光蛋白的力表达显示出更大的应力纤维定位。这些结果提示了一种模型，即mXin α的C端可能阻止全长分子与肌动蛋白结合，直到β -连环蛋白结合区域被β -连环蛋白占据。mXin与-连环蛋白在粘附体连接处的结合会促进肌动蛋白的结合。为了支持这个模型，我们发现mXin α的肌动蛋白结合和捆绑活性在β -catenin的存在下增强。
Targeted deletion of mXin alpha results in cardiac hypertrophy and cardiomyopathy with conduction defects (Gustafson-Wagner, E., Sinn, H. W., Chen, Y.-L., Wang, D.-Z., Reiter, R. S., Lin, J.L.-C., Yang, B., Williamson, R. A., Chen, J.N., Lin, C.-I., and Lin, J.J.-C. ( 2007) Am. J. Physiol. 293, H2680 - H2692). To understand the underlying mechanisms leading to such cardiac defects, the functional domains of mXin alpha and its interacting proteins were investigated. Interaction studies using co-immunoprecipitation, pull-down, and yeast two-hybrid assays revealed that mXin alpha directly interacts with beta-catenin. The beta-catenin-binding site on mXin alpha was mapped to amino acids 535 - 636, which overlaps with the known actin-binding domains composed of the Xin repeats. The overlapping nature of these domains provides insight into the molecular mechanism for mXin alpha localization and function. Purified recombinant glutathione S-transferase- or His-tagged mXin alpha proteins are capable of binding and bundling actin filaments, as determined by co-sedimentation and electron microscopic studies. The binding to actin was saturated at an approximate stoichiometry of nine actin monomers to one mXin alpha. A stronger interaction was observed between mXin alpha C-terminal deletion and actin as compared with the interaction between full-length mXin alpha and actin. Furthermore, force expression of green fluorescent protein fused to an mXin alpha C-terminal deletion in cultured cells showed greater stress fiber localization compared with force-expressed GFP-mXin alpha. These results suggest a model whereby the C terminus of mXin alpha may prevent the full-length molecule from binding to actin, until the beta-catenin- binding domain is occupied by beta-catenin. The binding of mXin alpha to beta-catenin at the adherens junction would then facilitate actin binding. In support of this model, we found that the actin binding and bundling activity of mXin alpha was enhanced in the presence of beta-catenin.