Regulatory mechanism in gene expression of regulatory proteins in cardiac and vascular smooth muscles

钙钙蛋白是我们在平滑肌细胞中发现的肌动蛋白，原肌球蛋白结合蛋白。本课题研究了钙钙蛋白在平滑肌细胞中的定位及钙钙蛋白的生理功能。(1)以7∶0.9∶0.6∶0.7的摩尔比获得了含有肌动蛋白、原肌球蛋白、钙调素和钙钙蛋白的细纤维。在大鼠主动脉平滑肌细胞的应激纤维上发现肌动蛋白和钙钙蛋白的共定位。(2)体外研究钙钙蛋白的功能。钙钙蛋白以剂量依赖性的方式抑制肌球蛋白MgATPase活性。这种抑制作用与钙无关。肌球蛋白酶活性的降低与钙钙蛋白与肌动蛋白-促球蛋白丝的结合有关。Caldesmon进一步抑制了钙钙蛋白诱导的硫代磷酸化肌球蛋白的MgATPase活性的抑制。钙调蛋白的加入逆转了钙调蛋白对肌球蛋白MgATPase的抑制作用。这些结果表明钙调蛋白作为平滑肌细丝的抑制成分。(3)克隆鸡胗平滑肌和大鼠主动脉平滑肌中SM22的cDNA。从鸡胗平滑肌中克隆的SM22编码cDNA全长1214bp，包含一个开放阅读框，编码200个氨基酸，计算分子量为22214。预测的氨基酸序列与发现该蛋白的Pearlstone等人确定的序列完全一致，除了在c端有两个额外的残基异亮氨酸和丝氨酸。来自大鼠主动脉的cDNA为1186bp。编码201个氨基酸（Mr 22602）。预测的氨基酸序列与鸡胗SM22的同源性为85%。

Calponin is actin-, tropomyosin binding protein which we found in the smooth muscle cells. In this project we studied the localization of calponin in smooth muscle cell and the physiological function of calponin.(1) We obtained thin filaments which contained actin, tropomyosin, caldesmon and calponin in molar rations of 7 : 0.9 : 0.6 : 0.7. Co-localization of actin and calponin was shown along stress fibers of smooth muscle cells from rat aorta.(2) We studied the function of calponin in vitro. Calponin inhibited the actinactivated myosin MgATPase activity in a dose-dependent manner. This inhibition was Ca-independent. The decrease in enzymatic activity of myosin was correlated with binding of calponin to actin-tropgmypsin filaments. Caldesmon showed a further inhibition of the calponin-induced inhibition of MgATPase activity of the thiophosphorylated myosin. Calponin-induced inhibition of the myosin MgATPase was reversed by the addition of calmodulin in the presence of Ca. These results suggest that calponin acts as an inhibitory component of smooth muscle thin filaments.(3) We performed cDNA cloning of SM22 from the chicken gizzard smooth muscle and rat aorta smooth muscle. Cloned cDNA encoding SM22 from the chicken gizzard smooth muscle had a total length of 1214bp and contained a single open reading frame which encodes 200 amino acids with a calculated molecular weight 22214. The predicted amino acid sequence was in complete agreement with the sequence determined by Pearlstone et al. who discovered the protein, except for two additional residues, isoleucine and serine at the C-terminus. The cDNA from rat aorta was 1186bp. in length and encoded 201 amino acids (Mr 22602). The predicted amino acid sequence showed an 85% Tdentity with the chicken gizzard SM22.