Nuclear Import Mechanism for Myocardin Family Members and Their Correlation with Vascular Smooth Muscle Cell Phenotype
Nuclear Import Mechanism for Myocardin Family Members and Their Correlation with Vascular Smooth Muscle Cell Phenotype
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DOI：
10.1074/jbc.m110.180786
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发表时间：
2010-11-26
影响因子：
4.8
通讯作者：
Sobue, Kenji
中科院分区：
文献类型：
--
作者：
Nakamura, Seiji;Hayashi, Ken'ichiro;Sobue, Kenji
肌心蛋白（IMycd）是平滑肌细胞谱系分化所必需的，其组成型位于细胞核中，尽管其家族成员肌心蛋白相关转录因子A和B（MRTF-A/B）主要驻留在细胞质中并响应Rho信号转导而易位至细胞核。他们进口核武器的机制尚不清楚。在这里，我们研究了Mycd家族成员的核输入机制，并证明了这种机制与血管平滑肌细胞（IVSMC）表型之间的任何相关性。在培养的VSMC中，importin β 1的敲低抑制Mycd和MRTF-A/B的核输入。通过体外分析，其NH 2-末端碱性结构域被鉴定为输入素α/Chi 1的结合位点。然而，即使在不存在G-肌动蛋白的情况下，Mycd对输入蛋白α/β 1的亲和力也高于MRTF-A/B，并且Mycd对输入蛋白α 1/β 1的亲和力强于任何其他输入蛋白α/β 1异二聚体。Mycd与importin α/β 1的结合对G-肌动蛋白不敏感，而MRTF-A/B的结合受到G-肌动蛋白不同程度的抑制。在去分化的VSMCs中，随着Mycd、血清反应因子和平滑肌细胞标志物的下调，importins α 1和β 1的水平降低。相反，在分化的VSMCs中，它们的表达上调。因此，VSMC中Mycd家族成员的核输入依赖于输入蛋白α/β 1，并且它们对输入蛋白α/β 1异二聚体的相对亲和力决定了Mycd核输入。Mycd核输入机制的表达与VSMC表型依赖性平滑肌细胞标志物的表达水平相关。
Myocardin (IMycd), which is essential for the differentiation of the smooth muscle cell lineage, is constitutively located in the nucleus, although its family members, myocardin-related transcription factors A and B (MRTF-A/B), mostly reside in the cytoplasm and translocate to the nucleus in response to Rho signaling. The mechanism for their nuclear import is unclear. Here we investigated the mechanism for the nuclear import of Mycd family members and demonstrated any correlation between such mechanism and the phenotype of vascular smooth muscle cells (IVSMCs). In cultured VSMCs, the knockdown of importin beta 1 inhibited the nuclear import of Mycd and MRTF-A/B. Their NH2-terminal basic domain was identified as a binding site for importin alpha/Chi 1 by in vitro analyses. However, Mycd had a higher affinity for importin alpha/beta 1 than did MRTF-A/B, even in the absence of G-actin, and Mycd affinity for importin alpha 1/beta 1 was stronger than for any other importin alpha/beta 1 heterodimers. The binding of Mycd to importin alpha/beta 1 was insensitive to G-actin, whereas that of MRTF-A/B was differently inhibited by G-actin. In dedifferentiated VSMCs, the levels of importins alpha 1 and beta 1 were reduced concomitant with down-regulation of Mycd, serum response factor, and smooth muscle cell markers. By contrast, in differentiated VSMCs, their expressions were up-regulated. Thus, the nuclear import of Mycd family members in VSMCs depends on importin alpha/beta 1, and their relative affinities for importin alpha/beta 1 heterodimers determine Mycd nuclear import. The expression of Mycd nuclear import machineries is related to the expression levels of VSMC phenotype-dependent smooth muscle cell markers.