Mak A S, Watson M H, Litwin C M, Wang J H
Department of Biochemistry, Queen's University, Kingston, Ontario, Canada.
J Biol Chem. 1991 Apr 15;266(11):6678-81.
A recent report that mitosis-specific phosphorylation causes the nonmuscle caldesmon to dissociate from microfilaments (Yamashiro, S., Yamakita, Y., Ishikawa, R., and Matsumura, F. (1990) Nature 344, 675-678) suggests that this process may contribute to the major structural reorganization of the eukaryotic cell at mitosis. In this study we have demonstrated that smooth muscle caldesmon is phosphorylated in vitro by cdc2 kinase from mitotic phase HeLa cells to 1.2 mol of phosphate/mol of caldesmon. Tryptic maps showed three major phosphorylated spots and approximately equal amounts of phosphorylated Ser and Thr were identified. F-actin or calmodulin in the presence of Ca2+ blocks the phosphorylation of caldesmon. Phosphorylation of caldesmon greatly reduced its binding to F-actin. The phosphorylation sites were located in a 10,000-Da CnBr fragment at the COOH-terminal end of the caldesmon molecule known to house the binding sites for actin and calmodulin (Bartegi A., Fattoum, A., Derancourt, J., and Kassab, R. (1990) J. Biol. Chem. 265, 15231-15238). Our finding supports the model that phosphorylation of caldesmon by cdc2 kinase at mitosis may contribute to the disassembly of the microfilament bundles during prophase.
最近有报道称，有丝分裂特异性磷酸化导致非肌肉钙调蛋白从微丝上解离（Yamashiro, S., Yamakita, Y., Ishikawa, R., and Matsumura, F. (1990) Nature 344, 675 - 678），这表明该过程可能有助于真核细胞在有丝分裂时进行主要的结构重组。在本研究中，我们已证明，来自有丝分裂期HeLa细胞的cdc2激酶可在体外将平滑肌钙调蛋白磷酸化，磷酸化程度为每摩尔钙调蛋白1.2摩尔磷酸。胰蛋白酶图谱显示有三个主要的磷酸化位点，并且已鉴定出磷酸化的丝氨酸（Ser）和苏氨酸（Thr）的量大致相等。在Ca2+存在的情况下，F - 肌动蛋白或钙调蛋白可阻断钙调蛋白的磷酸化。钙调蛋白的磷酸化大大降低了其与F - 肌动蛋白的结合。磷酸化位点位于钙调蛋白分子COOH末端一个10,000道尔顿的溴化氰（CnBr）片段中，已知该片段包含肌动蛋白和钙调蛋白的结合位点（Bartegi A., Fattoum, A., Derancourt, J., and Kassab, R. (1990) J. Biol. Chem. 265, 15231 - 15238）。我们的发现支持这样一个模型，即有丝分裂时cdc2激酶对钙调蛋白的磷酸化可能有助于前期微丝束的解体。
