Department of Biochemistry and Molecular Biology, Monash University, Clayton, Victoria, Australia.
Department of Anatomical Pathology, Alfred Hospital, Prahran, Victoria, Australia.
Oncogene. 2015 Jul;34(28):3711-27. doi: 10.1038/onc.2014.303. Epub 2014 Sep 22.
Glioblastoma is the most common and lethal primary malignant brain tumor in adults. The tumor suppressor gene PTEN is deleted, mutated or hypermethylated in more than 60% of glioblastoma cases resulting in hyperactivation of the phosphoinositide 3-kinase pathway, which leads to sustained PI(3,4,5)P3 signaling, and thereby hyperactivation of Akt and other effectors. PI(3,4,5)P3 is also hydrolyzed to PI(3,4)P2 by inositol polyphosphate 5-phosphatases such as SKIP, but the role this pathway has in glioblastoma is unknown. Microarray expression profiling of SKIP in human glioblastoma has revealed both increased and decreased SKIP gene expression. Here we have screened PTEN-deficient glioblastoma for SKIP protein expression by immunohistochemistry and report that SKIP expression is increased in some cases or decreased relative to normal brain. Using the U-87MG PTEN-deficient cell line we show that SKIP knockdown did not further enhance cell proliferation or survival. However, SKIP overexpression in U-87MG cells suppressed anchorage-independent cell growth and growth factor-induced PI(3,4,5)P3/Akt signaling. Although, SKIP knockdown did not affect cell proliferation or survival, cell migration was significantly retarded, associated with significantly increased PI(4,5)P2 signals, and decreased phosphorylation of the actin-regulatory protein cofilin, a PI(4,5)P2-binding protein. Notably, overexpression of SKIP also inhibited migration of U-87MG cells to a similar degree as observed with PTEN reconstitution, however, via distinct mechanisms. PTEN reconstitution promoted sustained lamellipodia generation and focal adhesion formation. In contrast, SKIP overexpression reduced sustained lamellipodia formation, talin incorporation into focal adhesions and recruitment of PI(4,5)P2-binding proteins to the plasma membrane. Notably, analysis of two independent ONCOMINE microarray data sets revealed a significant correlation between increased SKIP mRNA expression in glioblastoma and improved long-term survival. Therefore, SKIP expression in glioblastoma may affect the local invasion of PTEN-deficient tumors.
胶质母细胞瘤是成人中最常见和最致命的原发性恶性脑肿瘤。在超过 60%的胶质母细胞瘤病例中，肿瘤抑制基因 PTEN 缺失、突变或超甲基化，导致磷酸肌醇 3-激酶途径的过度激活，从而导致 PI(3,4,5)P3 信号的持续激活，进而导致 Akt 和其他效应物的过度激活。PI(3,4,5)P3 也可被肌醇多磷酸 5-磷酸酶（如 SKIP）水解为 PI(3,4)P2，但该途径在胶质母细胞瘤中的作用尚不清楚。SKIP 在人类胶质母细胞瘤中的基因表达谱微阵列分析显示，SKIP 基因表达既有增加也有减少。在这里，我们通过免疫组织化学筛选了 PTEN 缺陷型胶质母细胞瘤中的 SKIP 蛋白表达，并报告 SKIP 表达在一些病例中增加或相对于正常大脑减少。使用 U-87MG PTEN 缺陷细胞系，我们表明 SKIP 敲低不会进一步增强细胞增殖或存活。然而，U-87MG 细胞中 SKIP 的过表达抑制了锚定非依赖性细胞生长和生长因子诱导的 PI(3,4,5)P3/Akt 信号。尽管 SKIP 敲低不会影响细胞增殖或存活，但细胞迁移明显延迟，与 PI(4,5)P2 信号显著增加以及肌动蛋白调节蛋白丝束蛋白的磷酸化减少相关，丝束蛋白是一种 PI(4,5)P2 结合蛋白。值得注意的是，SKIP 的过表达也以与观察到的 PTEN 重建相似的程度抑制了 U-87MG 细胞的迁移，但通过不同的机制。PTEN 重建促进了持续的片状伪足生成和焦点形成。相比之下，SKIP 过表达减少了持续的片状伪足形成、talin 整合到焦点形成中以及 PI(4,5)P2 结合蛋白向质膜的募集。值得注意的是，对两个独立的 ONCOMINE 微阵列数据集的分析表明，胶质母细胞瘤中 SKIP mRNA 表达增加与长期生存改善之间存在显著相关性。因此，胶质母细胞瘤中 SKIP 的表达可能影响 PTEN 缺陷型肿瘤的局部浸润。
