MRCKβ links Dasm1 to actin rearrangements to promote dendrite development.
MRCKβ links Dasm1 to actin rearrangements to promote dendrite development.
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DOI：
10.1016/j.jbc.2021.100730
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发表时间：
2021-01
期刊：
影响因子：
--
通讯作者：
Chen S
中科院分区：
文献类型：
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作者：
Wang XX;Zhang S;Dong PP;Li YH;Zhang L;Shi SH;Yu ZQ;Chen S
正确的树突形态发生和突触形成对神经元的发育和功能至关重要。Dasm1是免疫球蛋白超家族的一员，已知在体外促进树突生长和兴奋性突触成熟。然而，Dasm1在神经元发育中的体内功能及其潜在机制尚不清楚。为了了解更多信息，我们构建了Dasm1敲除小鼠，并利用它们来证实Dasm1在体内调节树突乔木化和脊柱形成。我们使用Dasm1进行酵母双杂交筛选，发现MRCKβ是假定的伴侣；其他证据证实了这种相互作用，并鉴定了Dasm1和MRCKβ自激活激酶结构域（CC1, 410-744 aa）的细胞质脯氨酸丰富区（823-947 aa）是结合的必要和充分条件。通过共免疫沉淀实验、自磷酸化实验和BS3交联实验，我们发现Dasm1结合触发MRCKβ构象的变化和随后的二聚化，导致自磷酸化和激活。激活的MRCKβ反过来磷酸化2类调节肌球蛋白轻链，导致肌动蛋白重排增强，导致前面观察到的树突生长和脊柱形成。在小鼠中去除Dasm1会导致行为异常。总之，这些结果揭示了一个重要的分子途径介导细胞表面和细胞内信号通信，以调节肌动蛋白动力学和哺乳动物大脑中的神经元发育。
Proper dendrite morphogenesis and synapse formation are essential for neuronal development and function. Dasm1, a member of the immunoglobulin superfamily, is known to promote dendrite outgrowth and excitatory synapse maturation in vitro. However, the in vivo function of Dasm1 in neuronal development and the underlying mechanisms are not well understood. To learn more, Dasm1 knockout mice were constructed and employed to confirm that Dasm1 regulates dendrite arborization and spine formation in vivo. We performed a yeast two-hybrid screen using Dasm1, revealing MRCKβ as a putative partner; additional lines of evidence confirmed this interaction and identified cytoplasmic proline-rich region (823–947 aa) of Dasm1 and MRCKβ self-activated kinase domain (CC1, 410–744 aa) as necessary and sufficient for binding. Using co-immunoprecipitation assay, autophosphorylation assay, and BS3 cross-linking assay, we show that Dasm1 binding triggers a change in MRCKβ’s conformation and subsequent dimerization, resulting in autophosphorylation and activation. Activated MRCKβ in turn phosphorylates a class 2 regulatory myosin light chain, which leads to enhanced actin rearrangement, causing the dendrite outgrowth and spine formation observed before. Removal of Dasm1 in mice leads to behavioral abnormalities. Together, these results reveal a crucial molecular pathway mediating cell surface and intracellular signaling communication to regulate actin dynamics and neuronal development in the mammalian brain.