Hall Thomas E, Cole Nicholas J, Johnston Ian A
Gatty Marine Laboratory, School of Biology, University of St Andrews, Fife, KY16 8LB, UK.
J Exp Biol. 2003 Sep;206(Pt 18):3187-200. doi: 10.1242/jeb.00535.
Seven cDNA clones coding for different muscle-specific proteins (MSPs) were isolated from the fast muscle tissue of Atlantic cod Gadus morhua L. In situ hybridization using cRNA probes was used to characterize the temporal and spatial patterns of gene expression with respect to somite stage in embryos incubated at 4 degrees C, 7 degrees C and 10 degrees C. MyoD transcripts were first observed in the presomitic mesoderm prior to somite formation, and in the lateral compartment of the forming somites. MyoD expression was not observed in the adaxial cells that give rise to the slow muscle layer, and expression was undetectable by in situ hybridization in the lateral somitic mesoderm after the 35-somite stage, during development of the final approximately 15 somites. RT-PCR analysis, however, confirmed the presence of low levels of the transcript during these later stages. A phylogenetic comparison of the deduced aminoacid sequences of the full-length MyoD cDNA clone and those from other teleosts, and inference from the in situ expression pattern suggested homology with a second paralogue (MyoD2) recently isolated from the gilthead seabream Sparus aurata. Following MyoD expression, alpha-actin was the first structural gene to be switched on at the 16-somite stage, followed by myosin heavy chain, troponin T, troponin I and muscle creatine kinase. The final mRNA in the series to be expressed was troponin C. All genes were switched on prior to myofibril assembly. The troponin C sequence was unusual in that it showed the greatest sequence identity with the rainbow trout Oncorhynchus mykiss cardiac/slow form, but was expressed in the fast myotomal muscle and not in the heart. In addition, the third TnC calcium binding site showed a lower level of sequence conservation than the rest of the sequence. No differences were seen in the timing of appearance or rate of posterior progression (relative to somite stage) of any MSP transcripts between embryos raised at the different temperatures. It was concluded that myofibrillar genes are activated asynchronously in a distinct temporal order prior to myofibrillar assembly and that this process was highly canalized over the temperature range studied.
从大西洋鳕鱼（Gadus morhua L.）的快肌组织中分离出了7个编码不同肌肉特异性蛋白（MSP）的cDNA克隆。使用cRNA探针进行原位杂交，以表征在4℃、7℃和10℃下孵育的胚胎中，基因表达相对于体节阶段的时间和空间模式。MyoD转录本首先在体节形成前的前体节中胚层以及正在形成的体节的外侧部分中被观察到。在产生慢肌层的近轴细胞中未观察到MyoD表达，并且在最终约15个体节发育过程中的35体节阶段之后，在体节外侧中胚层中通过原位杂交无法检测到表达。然而，RT-PCR分析证实了在这些后期阶段存在低水平的转录本。对全长MyoD cDNA克隆与其他硬骨鱼的推导氨基酸序列进行系统发育比较，并根据原位表达模式推断，表明与最近从金头鲷（Sparus aurata）中分离出的第二个旁系同源物（MyoD2）具有同源性。在MyoD表达之后，α-肌动蛋白是在16体节阶段第一个被开启的结构基因，随后是肌球蛋白重链、肌钙蛋白T、肌钙蛋白I和肌肉肌酸激酶。该系列中最后一个表达的mRNA是肌钙蛋白C。所有基因在肌原纤维组装之前被开启。肌钙蛋白C序列不同寻常，因为它与虹鳟（Oncorhynchus mykiss）心脏/慢肌形式的序列同一性最高，但在快肌节肌中表达，而不在心脏中表达。此外，第三个TnC钙结合位点的序列保守性低于序列的其余部分。在不同温度下饲养的胚胎之间，任何MSP转录本出现的时间或向后进展的速率（相对于体节阶段）均未观察到差异。得出的结论是，肌原纤维基因在肌原纤维组装之前以不同的时间顺序异步激活，并且该过程在所研究的温度范围内高度稳定。
