Sorci G, Agneletti A L, Donato R
Section of Anatomy, Department of Experimental Medicine and Biochemical Sciences, University of Perugia, Via del Giochetto, C.P. 81 Succ. 3, 06122, Perugia, Italy.
Neuroscience. 2000;99(4):773-83. doi: 10.1016/s0306-4522(00)00238-4.
S100A1 and S100B are members of a multigenic family of Ca(2+)-binding proteins of the EF-hand type highly abundant in astrocyte and striated muscle cells that have been implicated in the Ca(2+)-dependent regulation of several intracellular activities including the assembly and disassembly of microtubules and type III intermediate filaments. In the present work we tested S100A1 and S100B for their ability to cause microtubule and/or intermediate filament disassembly in situ using triton-cytoskeletons obtained from U251 glioma cells and rat L6 myoblasts. Our results indicate that: (i) both proteins cause a Ca(2+)-dependent disassembly of cytoplasmic microtubules in a dose-dependent manner; (ii) the S100A1- and S100B-inhibitory peptide, TRTK-12, blocks the S100A1 and S100B effects on microtubules; (iii) S100A1Delta88-93, an S100A1 mutant lacking the C-terminal extension, does not affect microtubule stability; and (iv) no obvious S100A1- or S100B-dependent intermediate filament disassembly could be observed under the experimental conditions used in the present study, but S100A1- and S100B-dependent microtubule disassembly results in a tendency of vimentin intermediate filaments to aggregate into bundles and/or to condense. Together, these results suggest that S100A1 and S100B probably cause microtubule disassembly by interacting with the microtubule wall, and that the two proteins do not affect intermediate filament stability via interaction with preformed intermediate filaments, in agreement with previous biochemical investigation. Our present data lend support to the possibility that S100A1 and S100B might have a role in the in vivo regulation of the state of assembly of microtubules in a Ca(2+)-regulated manner and, potentially, on microtubule-based activities in astrocytes and myoblasts. Also, these data suggest that the both S100 proteins use their C-terminal extension for interacting with microtubules.
S100A1和S100B是EF手型钙结合蛋白多基因家族的成员，在星形胶质细胞和横纹肌细胞中高度丰富，参与了包括微管和III型中间丝的组装与拆卸在内的多种细胞内活动的钙依赖性调节。在本研究中，我们使用从U251胶质瘤细胞和大鼠L6成肌细胞获得的Triton细胞骨架，测试了S100A1和S100B原位引起微管和/或中间丝拆卸的能力。我们的结果表明：（i）两种蛋白质均以剂量依赖性方式引起细胞质微管的钙依赖性拆卸；（ii）S100A1和S100B抑制肽TRTK-12可阻断S100A1和S100B对微管的作用；（iii）缺少C末端延伸的S100A1突变体S100A1Delta88-93不影响微管稳定性；（iv）在本研究使用的实验条件下，未观察到明显的S100A1或S100B依赖性中间丝拆卸，但S100A1和S100B依赖性微管拆卸导致波形蛋白中间丝聚集形成束状和/或浓缩的趋势。总之，这些结果表明S100A1和S100B可能通过与微管壁相互作用引起微管拆卸，并且这两种蛋白质不会通过与预先形成的中间丝相互作用来影响中间丝稳定性，这与先前的生化研究一致。我们目前的数据支持S100A1和S100B可能以钙调节的方式在体内调节微管组装状态以及潜在地对星形胶质细胞和成肌细胞中基于微管的活动起作用的可能性。此外，这些数据表明这两种S100蛋白都利用其C末端延伸与微管相互作用。
