Birkenfeld Jörg, Betz Heinrich, Roth Dagmar
Department of Neurochemistry, Max-Planck-Institute for Brain Research, Deutschordenstrasse 46, 60528 Frankfurt, Germany.
Biochem J. 2003 Jan 1;369(Pt 1):45-54. doi: 10.1042/BJ20021152.
Proteins of the 14-3-3 family have been implicated in various physiological processes, and are thought to function as adaptors in various signal transduction pathways. In addition, 14-3-3 proteins may contribute to the reorganization of the actin cytoskeleton by interacting with as yet unidentified actin-binding proteins. Here we show that the 14-3-3 zeta isoform interacts with both the actin-depolymerizing factor cofilin and its regulatory kinase, LIM (Lin-11/Isl-1/Mec-3)-domain-containing protein kinase 1 (LIMK1). In both yeast two-hybrid assays and glutathione S-transferase pull-down experiments, these proteins bound efficiently to 14-3-3 zeta. Deletion analysis revealed consensus 14-3-3 binding sites on both cofilin and LIMK1. Furthermore, the C-terminal region of 14-3-3 zeta inhibited the binding of cofilin to actin in co-sedimentation experiments. Upon co-transfection into COS-7 cells, 14-3-3 zeta-specific immunoreactivity was redistributed into characteristic LIMK1-induced actin aggregations. Our data are consistent with 14-3-3-protein-induced changes to the actin cytoskeleton resulting from interactions with cofilin and/or LIMK1.
14-3-3家族蛋白参与了多种生理过程，被认为在各种信号转导途径中作为衔接蛋白发挥作用。此外，14-3-3蛋白可能通过与尚未确定的肌动蛋白结合蛋白相互作用，促进肌动蛋白细胞骨架的重组。在此，我们发现14-3-3ζ亚型与肌动蛋白解聚因子cofilin及其调节激酶含LIM（Lin-11/Isl-1/Mec-3）结构域的蛋白激酶1（LIMK1）相互作用。在酵母双杂交试验和谷胱甘肽S-转移酶下拉实验中，这些蛋白都能有效地与14-3-3ζ结合。缺失分析揭示了cofilin和LIMK1上的14-3-3共有结合位点。此外，在共沉降实验中，14-3-3ζ的C末端区域抑制了cofilin与肌动蛋白的结合。共转染到COS-7细胞后，14-3-3ζ特异性免疫反应重新分布到由LIMK1诱导的特征性肌动蛋白聚集体中。我们的数据与14-3-3蛋白通过与cofilin和/或LIMK1相互作用引起肌动蛋白细胞骨架变化的观点一致。
