Section of Physiology and Genetics, Department of Biology, University of Turku, Vesilinnantie 5, FI-20500, Turku, Finland.
Turku Bioscience, University of Turku and Åbo Akademi University, 20520, Turku, Finland.
Cell Commun Signal. 2020 Aug 8;18(1):121. doi: 10.1186/s12964-020-00618-6.
The PIM family kinases promote cancer cell survival and motility as well as metastatic growth in various types of cancer. We have previously identified several PIM substrates, which support cancer cell migration and invasiveness. However, none of them are known to regulate cellular movements by directly interacting with the actin cytoskeleton. Here we have studied the phosphorylation-dependent effects of PIM1 on actin capping proteins, which bind as heterodimers to the fast-growing actin filament ends and stabilize them.
Based on a phosphoproteomics screen for novel PIM substrates, we have used kinase assays and fluorescence-based imaging techniques to validate actin capping proteins as PIM1 substrates and interaction partners. We have analysed the functional consequences of capping protein phosphorylation on cell migration and adhesion by using wound healing and real-time impedance-based assays. We have also investigated phosphorylation-dependent effects on actin polymerization by analysing the protective role of capping protein phosphomutants in actin disassembly assays.
We have identified capping proteins CAPZA1 and CAPZB2 as PIM1 substrates, and shown that phosphorylation of either of them leads to increased adhesion and migration of human prostate cancer cells. Phosphorylation also reduces the ability of the capping proteins to protect polymerized actin from disassembly.
Our data suggest that PIM kinases are able to induce changes in actin dynamics to support cell adhesion and movement. Thus, we have identified a novel mechanism through which PIM kinases enhance motility and metastatic behaviour of cancer cells. Video abstract.
PIM 家族激酶促进了多种类型癌症中的肿瘤细胞存活、运动和转移性生长。我们先前已经鉴定了几种支持肿瘤细胞迁移和侵袭的 PIM 底物。然而，目前还没有已知的底物能够通过与肌动蛋白细胞骨架直接相互作用来调节细胞运动。在此，我们研究了 PIM1 通过磷酸化对肌动蛋白封端蛋白的影响，这些蛋白作为异二聚体与快速生长的肌动蛋白丝末端结合并稳定它们。
基于对新型 PIM 底物的磷酸化蛋白质组学筛选，我们使用激酶测定和基于荧光的成像技术验证了肌动蛋白封端蛋白是 PIM1 的底物和相互作用伙伴。我们通过划痕愈合和实时阻抗基础测定，分析了封端蛋白磷酸化对细胞迁移和黏附的功能后果。我们还通过分析封端蛋白磷酸突变体在肌动蛋白解聚测定中的保护作用，研究了磷酸化依赖性对肌动蛋白聚合的影响。
我们鉴定了 CAPZA1 和 CAPZB2 作为 PIM1 的底物，并且表明它们中的任一个的磷酸化都会导致人前列腺癌细胞的黏附和迁移增加。磷酸化还降低了封端蛋白保护聚合肌动蛋白免受解聚的能力。
我们的数据表明 PIM 激酶能够诱导肌动蛋白动力学的变化，以支持细胞黏附和运动。因此，我们已经确定了一种新的机制，通过该机制 PIM 激酶增强了肿瘤细胞的迁移和转移行为。视频摘要。
