Department of Biochemistry and Signal Transduction, University Medical Center Hamburg-Eppendorf, Martinistrasse 52, D-20246 Hamburg, Germany.
Institue of Anatomy and Experimental Morphology, University Medical Center Hamburg-Eppendorf, Martinistrasse 52, D-20246 Hamburg, Germany.
Biosci Rep. 2023 Feb 27;43(2). doi: 10.1042/BSR20222150.
Expression of Ins(1,4,5)P3-kinase-A (ITPKA), the neuronal isoform of Ins(1,4,5)P3-kinases, is up-regulated in many tumor types. In particular, in lung cancer cells this up-regulation is associated with bad prognosis and it has been shown that a high level of ITPKA increases migration and invasion of lung cancer cell lines. However, since ITPKA exhibits actin bundling and Ins(1,4,5)P3-kinase activity, it was not clear which of these activities account for ITPKA-promoted migration and invasion of cancer cells. To address this issue, we inhibited endogenous actin bundling activity of ITPKA in lung cancer H1299 cells by overexpressing the dominant negative mutant ITPKAL34P. Analysis of actin dynamics in filopodia as well as wound-healing migration revealed that ITPKAL34P inhibited both processes. Moreover, the formation of invasive protrusions into collagen I was strongly blocked in cells overexpressing ITPKAL34P. Furthermore, we found that ATP stimulation slightly but significantly (by 13%) increased migration of cells overexpressing ITPKA while under basal conditions up-regulation of ITPKA had no effect. In accordance with these results, overexpression of a catalytic inactive ITPKA mutant did not affect migration, and the Ins(1,4,5)P3-kinase-inhibitor GNF362 reversed the stimulating effect of ITPKA overexpression on migration. In summary, we demonstrate that under basal conditions the actin bundling activity controls ITPKA-facilitated migration and invasion and in presence of ATP the Ins(1,4,5)P3-kinase activity slightly enhances this effect.
Ins(1,4,5)P3-激酶-A（ITPKA）的表达在许多肿瘤类型中上调，其为 Ins(1,4,5)P3-激酶的神经元同工酶。特别是在肺癌细胞中，这种上调与预后不良相关，并且已经表明高水平的 ITPKA 增加了肺癌细胞系的迁移和侵袭。然而，由于 ITPKA 具有肌动蛋白成束和 Ins(1,4,5)P3-激酶活性，因此不清楚这些活性中的哪一种导致 ITPKA 促进癌细胞的迁移和侵袭。为了解决这个问题，我们通过过表达显性负突变体 ITPKAL34P 抑制肺癌 H1299 细胞中的内源性肌动蛋白成束活性。对片状伪足中的肌动蛋白动力学和划痕愈合迁移的分析表明，ITPKAL34P 抑制了这两个过程。此外，在过表达 ITPKAL34P 的细胞中，侵入胶原蛋白 I 的侵袭性突起的形成强烈受阻。此外，我们发现 ATP 刺激虽然轻微但显著（增加 13%）了过表达 ITPKA 的细胞的迁移，而在基础条件下 ITPKA 的上调没有影响。与这些结果一致，过表达无催化活性的 ITPKA 突变体不会影响迁移，并且 Ins(1,4,5)P3-激酶抑制剂 GNF362 逆转了 ITPKA 过表达对迁移的刺激作用。总之，我们证明在基础条件下，肌动蛋白成束活性控制 ITPKA 促进的迁移和侵袭，并且在存在 ATP 的情况下，Ins(1,4,5)P3-激酶活性略微增强了这种效应。
