Casein Kinase 1 Delta in Wnt Signaling and Beyond

(A)我们建立了尤文肿瘤细胞对WNT-3a的反应形成神经突起，并已开始确定解释这一效应的机制。敲除非典型的PKCiota也阻止了Wnt-3a依赖的神经突起的生长。WNT-3a可刺激PKCi的磷酸化。Dvl2与PKCi共沉淀，当Dvl2中的CK1磷酸化位点被丙氨酸残基取代时，这种相互作用不发生。这些结果提示，Dvl2与PKCi的结合需要CK1对Dvl2的磷酸化，这可能是轴突生长所必需的。这项工作意义重大，不仅是因为它提供了对神经突起形成机制的洞察。许多参与轴突生长的因素在其他环境中对细胞极性有贡献，例如对细胞迁移至关重要的细胞延伸的形成。(B)抑制酪蛋白激酶1增量(CK1d)，但不抑制CK1e，可阻断hTERT-RPE和mIMCD3细胞的原发纤毛发生。小鼠胚胎成纤维细胞(MEF)和Csnk1d(CK1d)缺失小鼠的视网膜细胞也表现出纤毛生成缺陷。在CK1d缺失的MEF中，纤毛的形成需要CK1d催化活性和中心体定位信号(CLS)。此外，在对照MEF中，含有CLS的截断衍生物的表达使全长CK1d从中心体移位并减少了纤毛长度，表明中心体CK1d在纤毛发生中起作用。抑制CK1d还改变了参与纤毛运输的几种蛋白质，包括Rab11a，Rab8a，CEP290，PCM1和Polycystin-2，以及其结合伙伴AKAP450的高尔基分布。根据对AKAP450的报道，CK1d是高尔基体微管成核和维持高尔基体完整性所必需的。含有CK1d结合位点的AKAP450片段的过表达抑制了高尔基体来源的微管成核、IFT20的高尔基体分布和纤毛发生。我们的结果表明，CK1d通过多种机制介导原发纤毛发生，一种涉及其中心体功能，另一种依赖于它与高尔基体AKAP450的相互作用，在高尔基体中，它对维持高尔基体组织和多种介导纤毛运输的因子的极化运输具有重要作用。(C)目前的研究集中在观察到抑制CK1d的表达或催化活性会扰乱DNA损伤修复和细胞周期调节。

(A) We established that cells from Ewing tumors form neurites in response to Wnt-3a and have begun to define the mechanisms that account for this effect. Knockdown of the atypical PKCiota also blocked Wnt-3a-dependent neurite outgrowth. Wnt-3a stimulated the phosphorylation of PKCi. Dvl2 co-immunoprecipitated with PKCi, and this interaction did not occur when CK1 phosphorylation sites in Dvl2 were replaced with alanine residues. These results suggested that Dvl2 phosphorylation by CK1 was required for Dvl2-PKCi binding, which in turn might be necessary for neurite outgrowth. This work is significant not only because it provides insights about mechanisms involved in the formation of neurites. Many of the factors that participate in neurite outgrowth contribute to cell polarity in other contexts such as the formation of cellular extensions critical for cell migration. (B) Inhibition of casein kinase 1 delta (CK1d), but not CK1e, blocked primary ciliogenesis in hTERT-RPE and mIMCD3 cells. Mouse embryonic fibroblasts (MEFs) and retinal cells from Csnk1d (CK1d) null mice also exhibited ciliogenesis defects. CK1d catalytic activity and centrosomal localization signal (CLS) were required to rescue cilia formation in CK1d null MEFs. Furthermore, expression of a truncated derivative containing the CLS displaced full-length CK1d from the centrosome and decreased ciliary length in control MEFs, suggesting that centrosomal CK1d has a role in ciliogenesis. CK1d inhibition also altered the pericentrosomal or ciliary distribution of several proteins involved in ciliary transport, including Rab11a, Rab8a, CEP290, PCM1 and polycystin-2, as well as the Golgi distribution of its binding partner, AKAP450. As reported for AKAP450, CK1d was required for microtubule nucleation at the Golgi and maintenance of Golgi integrity. Overexpression of an AKAP450 fragment containing the CK1d binding site inhibited Golgi-derived microtubule nucleation, Golgi distribution of IFT20 and ciliogenesis. Our results suggest that CK1d mediates primary ciliogenesis by multiple mechanisms, one involving its centrosomal function and another dependent on its interaction with AKAP450 at the Golgi where it is important for maintaining Golgi organization and polarized trafficking of multiple factors that mediate ciliary transport. (C) Current research is focused on observations that inhibition of CK1d expression or catalytic activity disrupts DNA damage repair and cell cycle regulation.