Novellino Luisa, De Filippo Annamaria, Deho Paola, Perrone Federica, Pilotti Silvana, Parmiani Giorgio, Castelli Chiara
Unit of Immunotherapy of Human Tumors, Department of Experimental Oncology, Fondazione IRCCS Istituto Nazionale dei Tumori, Milan, Italy.
Cell Signal. 2008 May;20(5):872-83. doi: 10.1016/j.cellsig.2007.12.024. Epub 2008 Jan 12.
Previous reports showed that receptor-type protein-tyrosine phosphatase PTPRK co-localizes with beta-catenin at adherens junctions, and in vitro experiments suggested that beta-catenin could be substrate of PTPRK-mediated phosphatase activity. beta-catenin is a molecule endowed with a dual function being involved both in cell adhesion and in Wnt signaling pathway. Here we provide evidence for the role of PTPRK in negatively regulating the beta-catenin transcriptional activity by modulating its intracellular and membrane distribution. Expression of PTPRK protein in HEK293 cells and in PTPRK-null melanoma cell lines, one of which harbors a mutated oncogenic beta-catenin, impairs nuclear accumulation of wild type and oncogenic forms of beta-catenin, limits cytosolic levels of tyrosine-phosphorylated beta-catenin, and leads to re-localization of E-cadherin/beta-catenin complexes in ordered membrane phase along cell-cell contacts. This re-modulation of beta-catenin cellular distribution results in the inhibition of cyclin D1 and c-myc protein expression, whose genes are targets of beta-catenin. Tumor cells upon re-expression of PTPRK have a reduced proliferative and migration capacity. Moreover we show that PTPRK is also active in negatively regulating the transactivating function of beta-catenin in normal melanocytes as confirmed by experiments with silenced PTPRK by specific siRNA. Our data show that PTPRK influences transactivating activity of beta-catenin in non-tumoral and neoplastic cells by regulating the balance between signaling and adhesive beta-catenin, thus providing biochemical basis for the hypothesis of PTPRK as a tumor suppressor gene.
先前的报道显示，受体型蛋白酪氨酸磷酸酶PTPRK与β-连环蛋白在黏附连接处共定位，体外实验表明β-连环蛋白可能是PTPRK介导的磷酸酶活性的底物。β-连环蛋白是一种具有双重功能的分子，既参与细胞黏附又参与Wnt信号通路。在此，我们提供证据表明PTPRK通过调节β-连环蛋白的细胞内和膜分布，对其转录活性起负调节作用。PTPRK蛋白在HEK293细胞和PTPRK基因缺失的黑色素瘤细胞系（其中之一含有突变的致癌性β-连环蛋白）中的表达，会损害野生型和致癌形式的β-连环蛋白的核内积累，限制酪氨酸磷酸化β-连环蛋白的胞质水平，并导致E-钙黏蛋白/β-连环蛋白复合物沿细胞间接触重新定位于有序的膜相。β-连环蛋白细胞分布的这种重新调节导致细胞周期蛋白D1和c-myc蛋白表达受到抑制，其基因是β-连环蛋白的靶标。重新表达PTPRK后的肿瘤细胞增殖和迁移能力降低。此外，我们还表明，通过用特异性siRNA沉默PTPRK的实验证实，PTPRK在正常黑素细胞中对β-连环蛋白的反式激活功能也具有负调节作用。我们的数据表明，PTPRK通过调节信号传导性β-连环蛋白和黏附性β-连环蛋白之间的平衡，影响非肿瘤细胞和肿瘤细胞中β-连环蛋白的反式激活活性，从而为PTPRK作为肿瘤抑制基因的假说提供了生化依据。
