Borisov Andrei B, Raeker Maide O, Russell Mark W
Division of Pediatric Cardiology, Department of Pediatrics and Communicable Diseases, University of Michigan Medical School, Ann Arbor, Michigan 48109-0204, USA.
J Cell Biochem. 2008 Apr 1;103(5):1621-35. doi: 10.1002/jcb.21551.
Obscurin and obscurin-associated kinase are two products of the obscurin transcriptional unit that encodes a recently identified giant muscle-specific protein obscurin. In this study, we characterized the developmental expression and cellular localization of obscurin and obscurin-associated kinase in cardiac muscle cells. We cloned murine obscurin-associated kinase and found that it is abundantly expressed in the heart as two isotypes encoded by 2.2 and 4.9 kb sequences. The 2.2 kb isotype of the kinase was more prominently expressed than the 4.9 kb isotype. Both obscurin and the kinase-like domains were progressively upregulated since the early stages of cardiac development. Obscurin-associated kinase was expressed at higher levels than obscurin at early stages of cardiomyogenesis. Increasing intensity of obscurin expression in the developing heart positively correlated with progressive cell differentiation and was higher in the ventricles compared to the atria. These data were supported by the results of experiments with primary cardiac cell cultures. Obscurin localization changed from a weakly immunopositive diffuse pattern in poorly differentiated cells to an intensely immunolabeled cross-striated distribution at the level of mid-A-bands and Z-disks during the assembly of the myofibrillar contractile apparatus. In dividing myocytes, unlike the interphase cells, obscurin translocated from disassembling myofibrils into a diffuse granulated pattern segregated separately from alpha-actinin-immunopositive aggregates. Obscurin-associated kinase was localized mainly to cell nuclei with increasing incorporation into the Z-disks during differentiation. Our results suggest that these two novel proteins are involved in the progression of cardiac myogenesis during the transition to advanced stages of heart development.
obscurin和obscurin相关激酶是obscurin转录单元的两个产物，该转录单元编码一种最近发现的巨大肌肉特异性蛋白obscurin。在本研究中，我们对obscurin和obscurin相关激酶在心肌细胞中的发育表达和细胞定位进行了表征。我们克隆了小鼠obscurin相关激酶，发现它在心脏中大量表达，有两种由2.2和4.9 kb序列编码的同种型。激酶的2.2 kb同种型比4.9 kb同种型表达更显著。自心脏发育早期以来，obscurin和激酶样结构域均逐渐上调。在心肌发生早期，obscurin相关激酶的表达水平高于obscurin。发育中的心脏中obscurin表达强度的增加与细胞的逐渐分化呈正相关，且心室中的表达高于心房。原代心脏细胞培养实验的结果支持了这些数据。在肌原纤维收缩装置组装过程中，obscurin的定位从分化不良细胞中的弱免疫阳性弥漫模式转变为A带中部和Z盘水平的强免疫标记横纹分布。在分裂的心肌细胞中，与间期细胞不同，obscurin从解体的肌原纤维转移到与α-肌动蛋白免疫阳性聚集体分开的弥漫颗粒模式中。obscurin相关激酶在分化过程中主要定位于细胞核，并逐渐整合到Z盘中。我们的结果表明，这两种新蛋白在心脏发育向晚期转变过程中参与了心肌发生的进程。
