STIL binding to Polo-box 3 of PLK4 regulates centriole duplication.
STIL binding to Polo-box 3 of PLK4 regulates centriole duplication.
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DOI：
10.7554/elife.07888
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发表时间：
2015-07-18
期刊：
影响因子：
7.7
通讯作者：
Maier T
中科院分区：
文献类型：
--
作者：
Arquint C;Gabryjonczyk AM;Imseng S;Böhm R;Sauer E;Hiller S;Nigg EA;Maier T
Polo样激酶（PLK）是真核细胞周期进程、有丝分裂和胞质分裂的调节因子; PLK 4是中心粒复制的主调节因子。在这里，我们证明了SCL/TAL 1中断位点（STIL）蛋白通过其卷曲螺旋区（STIL-CC）与PLK 4在体内相互作用。STIL-CC是第一个鉴定的PLK 4的Polo-box 3（PB 3）的相互作用伴侣，并且还使用PLK 4 L1区域中的次级相互作用位点。通过NMR和晶体学对游离PLK 4-PB 3及其STIL-CC复合物的结构测定揭示了一种模拟卷曲螺旋形成的Polo盒-肽相互作用的新模式。对结构指导的STIL突变体的体内分析揭示了与PLK 4-PB 3和L1的不同结合模式，以及STIL寡聚化与PLK 4结合的相互作用。我们认为，STIL-CC/PLK 4相互作用介导PLK 4的激活以及中心粒PLK 4的稳定，并在中心粒复制中起着关键作用。DOI：http://dx.doi.org/10.7554/eLife.07888.001 Centrioles是组织细胞内分子支架的结构，这对细胞的形状和活性以及细胞分裂期间重复染色体的分离都很重要。中心粒也构成了被称为纤毛的触角状结构的基部的一部分，纤毛从细胞表面伸出，允许细胞感知化学物质，触摸甚至移动。不分裂的细胞含有一对中心粒。在分裂的细胞中，两个中心粒在每个分裂周期复制一次，新的中心粒在每个现有中心粒旁边形成。中心粒只复制一次是至关重要的，因为额外的拷贝可能导致可能导致出生缺陷和癌症的问题。中心粒需要两种蛋白质，称为PLK 4和STIL，以复制。这两种蛋白质中的任何一种过量都会导致额外的中心粒。另一方面，如果缺少这些，就不能进行重复。PLK 4属于称为激酶的酶的大家族。激酶将磷酸基团连接到其他蛋白质上，这可以激活或灭活其他蛋白质。PLK 4可以将磷酸基团添加到STIL上，但尚不清楚这两种蛋白质如何相互作用。Arquint，Gabryjonczyk，Imseng，Böhm等人分析了人类细胞中的这种相互作用，发现PLK 4和STIL直接相互结合。STIL蛋白的一部分采用了所谓的“卷曲螺旋”结构，其中扭曲的蛋白质长度像一根绳子一样相互缠绕。卷曲螺旋与PLK 4的两个不同部分相互作用。根据这些观察结果，使用X射线晶体学和核磁共振可视化与STIL结合的PLK-4的三维结构。这些技术揭示了STIL的卷曲螺旋区域形成细长结构，PLK-4沿其整个长度沿着相互作用。Arquint，Gabryjonczyk，Imseng，Böhm等人然后分析了PLK 4和STIL是否需要彼此才能被招募到中心粒。当PLK 4在细胞中耗尽时，STIL从中心粒中丢失，表明PLK 4直接募集STIL。然而，与预期相反，当STIL水平降低时，PLK 4在中心粒积累。这表明STIL通过刺激其激酶活性来维持PLK 4的适当水平。需要进一步的工作来精确地理解PLK 4和STIL如何与下游作用的其他蛋白质相互作用，以高度受控的方式形成新的中心粒。DOI：http://dx.doi.org/10.7554/eLife.07888.002网站
Polo-like kinases (PLK) are eukaryotic regulators of cell cycle progression, mitosis and cytokinesis; PLK4 is a master regulator of centriole duplication. Here, we demonstrate that the SCL/TAL1 interrupting locus (STIL) protein interacts via its coiled-coil region (STIL-CC) with PLK4 in vivo. STIL-CC is the first identified interaction partner of Polo-box 3 (PB3) of PLK4 and also uses a secondary interaction site in the PLK4 L1 region. Structure determination of free PLK4-PB3 and its STIL-CC complex via NMR and crystallography reveals a novel mode of Polo-box–peptide interaction mimicking coiled-coil formation. In vivo analysis of structure-guided STIL mutants reveals distinct binding modes to PLK4-PB3 and L1, as well as interplay of STIL oligomerization with PLK4 binding. We suggest that the STIL-CC/PLK4 interaction mediates PLK4 activation as well as stabilization of centriolar PLK4 and plays a key role in centriole duplication. DOI: http://dx.doi.org/10.7554/eLife.07888.001 Centrioles are structures that organize the molecular scaffolding inside cells, which is important for a cell's shape and activity, as well as the segregation of duplicated chromosomes during cell division. Centrioles also form part of the base of the antenna-like structures called cilia, which project out from the cell's surface and allow cells to sense chemicals and touch or even to move. A cell that is not dividing contains a pair of centrioles. In dividing cells, the two centrioles duplicate once per cycle of division and a new centriole forms next to each of the existing ones. It is essential that centrioles duplicate only once, because extra copies can lead to problems that may cause birth defects and cancer. Centrioles require two proteins, called PLK4 and STIL, in order to duplicate. An excess of either of these proteins results in extra centrioles. On the other hand, if these are missing, duplication cannot take place. PLK4 belongs to a large family of enzymes called kinases. A kinase attaches a phosphate group to other proteins, which can either activate or deactivate the other protein. PLK4 can add phosphate groups onto STIL, but it is not known precisely how these two proteins interact with each other. Arquint, Gabryjonczyk, Imseng, Böhm et al. have analyzed this interaction in human cells and found that PLK4 and STIL bind directly to one another. Part of the STIL protein adopts a so-called ‘coiled-coil’ structure in which twisted lengths of protein wrap around each other like a piece of string. The coiled-coil interacts with two different parts of PLK4. Following on from these observations, the three-dimensional structure of PLK-4 bound to STIL was visualized using X-ray crystallography and nuclear magnetic resonance. These techniques revealed that the coiled-coil region of STIL forms an elongated structure and PLK-4 interacts along its entire length. Arquint, Gabryjonczyk, Imseng, Böhm et al. then analyzed whether PLK4 and STIL need one another in order to get recruited to centrioles. When PLK4 was depleted in cells, STIL was lost from centrioles, suggesting that PLK4 directly recruits STIL. However, contrary to expectations, when STIL levels were reduced, PLK4 accumulated at centrioles. This suggests that STIL maintains appropriate levels of PLK4 via stimulation of its kinase activity. Further work is needed to precisely understand how PLK4 and STIL interact with other proteins that act downstream to lead to the formation of new centrioles in a highly controlled manner. DOI: http://dx.doi.org/10.7554/eLife.07888.002