Liu G, Newell P C
Department of Biochemistry, University of Oxford, UK.
J Cell Sci. 1991 Apr;98 ( Pt 4):483-90. doi: 10.1242/jcs.98.4.483.
Previous studies have implicated cyclic GMP in the regulation of myosin II heavy chain (MHC) association with the cytoskeleton in Dictyostelium discoideum. Here we provide evidence that cyclic GMP may regulate MHC association with the cytoskeleton through MHC phosphorylation. Comparative data are presented of MHC phosphorylation in the wild-type strain NC4, the parental strain XP55 and streamer mutants NP368 and NP377. Using an anti-MHC monoclonal antibody to immunoprecipitate MHC from [32P]phosphate-labelled developing cells, we found that cyclic AMP stimulation of the wild-type strain NC4 and parental strain XP55 induced MHC phosphorylation in vivo. A peak of phosphorylation was observed at 30-40 s, followed by a gradual decrease to basal level at 160 s. In contrast, in both of the streamer mutants NP368 and NP377 (which have prolonged cyclic GMP accumulation and prolonged MHC association with the cytoskeleton), the phosphorylation of MHC was delayed and did not form a peak until 60-80 s after cyclic AMP stimulation. We also found that cytoskeletal MHC showed only minor phosphorylation, the majority of the phosphorylated MHC being found in the cytosol. We present a model to account for these results in which cyclic GMP regulates MHC association with the cytoskeleton by regulating the phosphorylation/dephosphorylation cycle of MHC in these cells.
先前的研究表明，环鸟苷酸（cGMP）参与调控盘基网柄菌中肌球蛋白II重链（MHC）与细胞骨架的结合。在此，我们提供证据表明，环鸟苷酸可能通过MHC磷酸化来调控MHC与细胞骨架的结合。文中给出了野生型菌株NC4、亲本菌株XP55以及拖尾突变体NP368和NP377中MHC磷酸化的比较数据。我们使用抗MHC单克隆抗体从[32P]磷酸盐标记的发育细胞中免疫沉淀MHC，发现环磷酸腺苷（cAMP）刺激野生型菌株NC4和亲本菌株XP55可在体内诱导MHC磷酸化。在30 - 40秒时观察到磷酸化峰值，随后在160秒时逐渐降至基础水平。相比之下，在拖尾突变体NP368和NP377中（这两个突变体具有延长的环鸟苷酸积累以及延长的MHC与细胞骨架的结合），MHC的磷酸化延迟，直到环磷酸腺苷刺激后60 - 80秒才形成峰值。我们还发现细胞骨架中的MHC仅显示轻微磷酸化，大部分磷酸化的MHC存在于细胞质中。我们提出了一个模型来解释这些结果，即环鸟苷酸通过调节这些细胞中MHC的磷酸化/去磷酸化循环来调控MHC与细胞骨架的结合。
