Kohno Takayuki, Ninomiya Takafumi, Kikuchi Shin, Konno Takumi, Kojima Takashi
Department of Cell Science, Research Institute for Frontier Medicine (T.Koh., T.Kon., T.Koj.), and Department of Anatomy, Sapporo Medical University, Sapporo, Japan (T.N., S.K.)
Department of Cell Science, Research Institute for Frontier Medicine (T.Koh., T.Kon., T.Koj.), and Department of Anatomy, Sapporo Medical University, Sapporo, Japan (T.N., S.K.).
Mol Pharmacol. 2015 May;87(5):815-24. doi: 10.1124/mol.114.096982. Epub 2015 Feb 13.
Staurosporine (STS) has been known as a classic protein kinase C inhibitor and is a broad-spectrum inhibitor targeting over 250 protein kinases. In this study, we observed that STS treatment induced drastic morphologic changes, such as elongation of a very large number of nonbranched, actin-based long cell protrusions that reached up to 30 µm in an hour without caspase activation or PARP cleavage in fibroblasts and epithelial cells. These cell protrusions were elongated not only from the free cell edge but also from the cell-cell junctions. The elongation of STS-dependent protrusions was required for ATP hydrolysis and was dependent on myosin-X and fascin but independent of Cdc42 and VASP. Interestingly, in the presence of an actin polymerization inhibitor, namely, cytochalasin D, latrunculin A, or jasplakinolide, STS treatment induced excess tubulin polymerization, which resulted in the formation of many extra-long microtubule (MT)-based protrusions toward the outside of the cell. The unique MT-based protrusions were thick and linear compared with the STS-induced filaments or stationary filopodia. These protrusions, which were composed of microtubules, have been scarcely observed in cultured non-neuronal cells. Taken together, our findings revealed that STS-sensitive kinases are essential for the maintenance of normal cell morphology, and a common unidentified molecular mechanism is involved in the formation of the following two different types of protrusions: actin-based filaments and MT-based shafts.
星形孢菌素（STS）一直被认为是一种经典的蛋白激酶C抑制剂，是一种靶向超过250种蛋白激酶的广谱抑制剂。在本研究中，我们观察到STS处理诱导了剧烈的形态变化，例如在成纤维细胞和上皮细胞中，大量无分支的、基于肌动蛋白的长细胞突起伸长，在一小时内可达30 µm，且无半胱天冬酶激活或聚（ADP-核糖）聚合酶（PARP）裂解。这些细胞突起不仅从游离的细胞边缘伸长，也从细胞-细胞连接处伸长。依赖于STS的突起伸长需要ATP水解，并且依赖于肌球蛋白-X和丝束蛋白，但不依赖于Cdc42和血管舒张刺激磷蛋白（VASP）。有趣的是，在存在肌动蛋白聚合抑制剂（即细胞松弛素D、拉特罗毒素A或茉莉酮酸甲酯）的情况下，STS处理诱导了过量的微管蛋白聚合，导致许多超长的基于微管（MT）的突起向细胞外形成。与STS诱导的细丝或静止丝状伪足相比，独特的基于MT的突起厚且呈线性。这些由微管组成的突起在培养的非神经元细胞中很少被观察到。综上所述，我们的研究结果表明，对STS敏感的激酶对于维持正常细胞形态至关重要，并且一种共同的未确定分子机制参与了以下两种不同类型突起的形成：基于肌动蛋白的细丝和基于MT的轴。
