Hastings K E, Koppe R I, Marmor E, Bader D, Shimada Y, Toyota N
Montreal Neurological Institute, Department of Neurology/Neurosurgery, McGill University, Canada.
J Biol Chem. 1991 Oct 15;266(29):19659-65.
We isolated avian (chicken and quail) cardiac troponin I (TnIcardiac) cDNA clones for studies of Tn-Icardiac protein structure/evolution and developmental gene regulation. Comparison of the cDNA-predicted avian TnIcardiac amino acid sequences with known TnI sequences indicated 1) that the presence of an N-terminal extension sequence carrying a dual protein kinase A phosphorylation target site and an adjacent proline-rich segment is an ancient cardiac-specific feature of TnI which has been conserved since the bird/mammal divergence, 2) that features of the near-N-terminal troponin C (TnC)-binding site sequence suggest isoform-specific adaptation of TnI and TnC, and 3) that the avian TnIcardiac internal actin/TnC-binding, actomyosin-inhibitory, domain shows significant sequence divergence from mammalian TnIcardiac sequences, including the absence of a protein kinase C target site which is a cardiac-specific feature of TnI in mammals. Use of the cDNA clones to probe TnIcardiac mRNA expression during striated muscle development showed active expression in cardiac muscle from early developmental times (day 4 in ovo), but not in embryonic or adult skeletal muscle or in embryonic skeletal muscle cell cultures. Transcriptional run-on analysis showed that the heart-specific expression of TnIcardiac mRNA in embryonic striated muscle reflects transcriptional control of TnIcardiac gene expression. In many other contractile protein gene families, genes encoding cardiac isoforms are expressed early in skeletal muscle development and are later repressed. Thus, the restriction of active TnIcardiac gene expression to the cardiac muscle cell lineage is an unusual expression pattern for cardiac contractile protein genes and indicates that diverse gene regulatory mechanisms direct the differential expression of cardiac and skeletal muscle isoforms in different muscle gene families.
我们分离出了禽类（鸡和鹌鹑）心肌肌钙蛋白I（TnIcardiac）的cDNA克隆，用于研究TnIcardiac蛋白结构/进化以及发育过程中的基因调控。将cDNA预测的禽类TnIcardiac氨基酸序列与已知的TnI序列进行比较，结果表明：1）携带双蛋白激酶A磷酸化靶点位点的N端延伸序列以及相邻富含脯氨酸片段的存在是TnI的一个古老的心脏特异性特征，自鸟类/哺乳动物分化以来一直保守；2）近N端肌钙蛋白C（TnC）结合位点序列的特征表明TnI和TnC存在亚型特异性适应性；3）禽类TnIcardiac内部肌动蛋白/TnC结合、肌动球蛋白抑制结构域与哺乳动物TnIcardiac序列存在显著的序列差异，包括缺少蛋白激酶C靶点位点，而该位点是哺乳动物TnI的心脏特异性特征。利用cDNA克隆探测横纹肌发育过程中TnIcardiac mRNA的表达，结果显示从发育早期（胚胎期第4天）开始，TnIcardiac就在心肌中活跃表达，但在胚胎或成年骨骼肌以及胚胎骨骼肌细胞培养物中不表达。转录延伸分析表明，胚胎横纹肌中TnIcardiac mRNA的心脏特异性表达反映了TnIcardiac基因表达的转录调控。在许多其他收缩蛋白基因家族中，编码心脏亚型的基因在骨骼肌发育早期表达，随后被抑制。因此，将活跃的TnIcardiac基因表达限制在心肌细胞谱系是心脏收缩蛋白基因一种不寻常的表达模式，这表明不同的基因调控机制指导着不同肌肉基因家族中心脏和骨骼肌亚型的差异表达。
