Graduate School of Medicine, Osaka University, Toyonaka City, Osaka, Japan.
J Appl Physiol (1985). 2012 Jan;112(1):176-86. doi: 10.1152/japplphysiol.00783.2011. Epub 2011 Oct 13.
The main purpose of the present study was to investigate the role(s) of 25-kDa heat shock protein (HSP25) in the regulation and integration of myofibrillar Z-disc structure during down- or upregulation of the size in rat soleus muscle fibers. Hindlimb unloading by tail suspension was performed in adult rats for 7 days, and reloading was allowed for 5 days after the termination of suspension. Interaction of HSP25 and Z-disc proteins, phosphorylation status, distribution, and complex formation of HSP25 were investigated. Non- and single-phosphorylated HSP25s were generally expressed in the cytoplasmic fraction of normal muscle. The level of total HSP25, as well as the phosphorylation ratio, did not change significantly in response to atrophy. Increased expressions of HSP25, phosphorylated at serine 15 (p-Ser15) and dual-phosphorylated form, were noted, when atrophied muscles were reloaded. Myofibrillar HSP25 was also noted in reloaded muscle. Histochemical analysis further indicated the localization of p-Ser15 in the regions with disorganization of Z-disc structure in reloaded muscle fibers. HSP25 formed a large molecular complex in the cytoplasmic fraction of normal muscle, whereas dissociation of free HSP25 with Ser15 phosphorylation was noted in reloaded muscle. The interaction of p-Ser15 with desmin and actinin was detected in Z-discs by proximity ligation assay. Strong interaction between p-Ser15 and desmin, but not actinin, was noted in the disorganized areas. These results indicated that HSP25 contributed to the desmin cytoskeletal organization following the phosphorylation at Ser15 during reloading and regrowing of soleus muscle.
本研究的主要目的是探讨 25kDa 热休克蛋白（HSP25）在大鼠比目鱼肌纤维大小下调或上调过程中调节和整合肌原纤维 Z 盘结构中的作用。通过尾部悬吊使成年大鼠后肢去负荷 7 天，并在悬吊结束后允许再负荷 5 天。研究了 HSP25 与 Z 盘蛋白的相互作用、磷酸化状态、分布以及 HSP25 的复合物形成。非磷酸化和单磷酸化的 HSP25 通常在正常肌肉的细胞质部分表达。总 HSP25 的水平以及磷酸化比率在对萎缩的反应中没有明显变化。当萎缩的肌肉再负荷时，观察到 HSP25 的表达增加，磷酸化在丝氨酸 15 （p-Ser15）和双磷酸化形式。在再负荷的肌肉中也观察到肌原纤维 HSP25。组织化学分析进一步表明，p-Ser15 在再负荷肌肉纤维中 Z 盘结构紊乱的区域定位。HSP25 在正常肌肉的细胞质部分形成一个大的分子复合物，而在再负荷的肌肉中则观察到游离 HSP25 与 Ser15 磷酸化的解离。通过邻近连接测定法在 Z 盘上检测到 p-Ser15 与结蛋白和肌动蛋白的相互作用。在紊乱区域观察到 p-Ser15 与结蛋白之间的强烈相互作用，但与肌动蛋白之间没有。这些结果表明，在比目鱼肌再负荷和再生过程中，HSP25 通过 Ser15 磷酸化对结蛋白细胞骨架组织起作用。
