Identification of a new mechanism for targeting myosin II heavy chain phosphorylation by Dictyostelium myosin heavy chain kinase B.
Identification of a new mechanism for targeting myosin II heavy chain phosphorylation by Dictyostelium myosin heavy chain kinase B.
复制标题
鉴定网网柄菌肌球蛋白重链激酶 B 靶向肌球蛋白 II 重链磷酸化的新机制。
DOI：
10.1186/1756-0500-3-56
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发表时间：
2010-03-03
影响因子：
1.8
通讯作者：
Steimle, Paul A
中科院分区：
文献类型：
--
作者：
Underwood, Julie;Greene, Jonathan;Steimle, Paul A
重链磷酸化在调节网骨藻和高等真核非肌肉细胞中肌球蛋白II双极丝组装中起着重要作用。我们先前的工作已经证明，与MHCK-A中的对应物不同，网骨藻肌球蛋白II重链激酶B（MHCK-B）的WD重复结构域对于靶向激酶磷酸化MHC不是绝对必需的。因此，我们测试了这样的假设，即MHCK-B特有的富含天冬酰胺且结构无序的区域本身可以在底物靶向中发挥作用。比较全长MHCK-B、仅缺乏WD重复结构域的截短物（B-Δ-WD）和缺乏富N区和WD重复结构域的截短物（B-Δ-N-WD）的活性的生物化学测定揭示富N区靶向MHCK-B以导致双极丝解体的方式磷酸化MHC。这种靶向是生理学相关的，因为B-Δ-WD截短而非B-Δ-N-WD截短的细胞过表达导致肌球蛋白II细丝组装水平显著降低以及胞质分裂和多细胞发育中的相关缺陷。这里呈现的结果表明，MHCK-B的固有非结构化和富含天冬酰胺的区域可以介导激酶特异性靶向磷酸化肌球蛋白II重链。这种靶向涉及与肌球蛋白II细丝的直接结合相互作用。在调节肌球蛋白双极丝组装方面，我们的研究结果表明，影响MHCK-B这一独特区域活性的因素可以允许以不同于网骨藻中其他MHCKs的方式调节MHCK-B。
Heavy chain phosphorylation plays a central role in regulating myosin II bipolar filament assembly in Dictyostelium, as well as in higher eukaryotic nonmuscle cells. Our previous work has demonstrated that the WD-repeat domain of Dictyostelium myosin II heavy chain kinase B (MHCK-B), unlike its counterpart in MHCK-A, is not absolutely required for targeting of the kinase to phosphorylate MHC. Thus, we tested the hypothesis that an asparagine-rich and structurally disordered region that is unique to MHCK-B can by itself function in substrate targeting. Biochemical assays comparing the activities of full-length MHCK-B, a truncation lacking only the WD-repeat domain (B-Δ-WD), and a truncation lacking both the N-rich region and the WD-repeat domain (B-Δ-N-WD) revealed that the N-rich region targets MHCK-B to phosphorylate MHC in a manner that leads to bipolar filament disassembly. This targeting is physiologically relevant since cellular over-expression of the B-Δ-WD truncation, but not the B-Δ-N-WD truncation, leads to dramatically reduced levels of myosin II filament assembly and associated defects in cytokinesis and multicellular development. The results presented here demonstrate that an intrinsically unstructured, and asparagine-rich, region of a MHCK-B can mediate specific targeting of the kinase to phosphorylate myosin II heavy chain. This targeting involves a direct binding interaction with myosin II filaments. In terms of regulating myosin bipolar filament assembly, our results suggest that factors affecting the activity of this unique region of MHCK-B could allow for regulation of MHCK-B in a manner that is distinct from the other MHCKs in Dictyostelium.