Department of Internal Medicine III, University of Heidelberg, Germany.
Circ Res. 2010 Nov 12;107(10):1253-64. doi: 10.1161/CIRCRESAHA.110.222372. Epub 2010 Sep 16.
The M-band represents a transverse structure in the center of the sarcomeric A-band and provides an anchor for the myosin-containing thick filaments. In contrast to other sarcomeric structures, eg, the Z-disc, only few M-band-specific proteins have been identified to date, and its exact molecular composition remains unclear.
Using a bioinformatic approach to identify novel heart- and muscle-specific genes, we found a leucine rich protein, myomasp (Myosin-interacting, M-band-associated stress-responsive protein)/LRRC39. RT-PCR and Northern and Western blot analyses confirmed a cardiac-enriched expression pattern, and immunolocalization of myomasp revealed a strong and specific signal at the sarcomeric M-band. Yeast 2-hybrid screens, as well as coimmunoprecipitation experiments, identified the C terminus of myosin heavy chain (MYH)7 as an interaction partner for myomasp. Knockdown of myomasp in neonatal rat ventricular myocytes (NRVCMs) led to a significant upregulation of the stretch-sensitive genes GDF-15 and BNP. Conversely, the expression of MYH7 and the M-band proteins myomesin-1 and -2 was found to be markedly reduced. Mechanistically, knockdown of myomasp in NRVCM led to a dose-dependent suppression of serum response factor-dependent gene expression, consistent with earlier observations linking the M-band to serum response factor-mediated signaling. Finally, downregulation of myomasp/LRRC39 in spontaneously beating engineered heart tissue constructs resulted in significantly lower force generation and reduced fractional shortening. Likewise, knockdown of the myomasp/LRRC39 ortholog in zebrafish resulted in severely impaired heart function and cardiomyopathy in vivo.
These findings reveal myomasp as a previously unrecognized component of an M-band-associated signaling pathway that regulates cardiomyocyte gene expression in response to biomechanical stress.
M 带是肌节 A 带中心的一种横向结构，为含有肌球蛋白的粗丝提供了一个附着点。与其他肌节结构（例如 Z 盘）不同，迄今为止仅鉴定出少数特定于 M 带的蛋白质，其确切的分子组成仍不清楚。
我们使用生物信息学方法来鉴定新的心脏和肌肉特异性基因，发现了一种富含亮氨酸的蛋白质，肌球蛋白相互作用蛋白（Myosin-interacting，M-band-associated stress-responsive protein）/LRRC39。RT-PCR、Northern 和 Western blot 分析证实了心脏丰富的表达模式，肌球蛋白相互作用蛋白的免疫定位显示肌节 M 带上有强烈而特异的信号。酵母 2 杂交筛选以及共免疫沉淀实验鉴定肌球蛋白重链（MYH）7 的 C 端为肌球蛋白相互作用蛋白的相互作用伙伴。在新生大鼠心室肌细胞（NRVCM）中敲低肌球蛋白相互作用蛋白导致伸展敏感基因 GDF-15 和 BNP 的显著上调。相反，MYH7 和 M 带蛋白肌联蛋白-1 和 -2 的表达明显降低。从机制上讲，NRVCM 中肌球蛋白相互作用蛋白的敲低导致血清反应因子依赖性基因表达呈剂量依赖性抑制，这与将 M 带与血清反应因子介导的信号联系起来的早期观察结果一致。最后，在自发搏动的工程心脏组织构建体中下调肌球蛋白相互作用蛋白/LRRC39 导致产生的力显著降低和分数缩短减少。同样，在斑马鱼中敲低肌球蛋白相互作用蛋白/LRRC39 的同源物导致体内心脏功能严重受损和心肌病。
这些发现揭示了肌球蛋白相互作用蛋白作为一种以前未被识别的 M 带相关信号通路的组成部分，该信号通路调节心肌细胞基因表达以响应生物力学应激。
