Institute of Anatomy and Cell Biology, Justus-Liebig-University, Giessen, Germany.
Institute of Pathology, Justus-Liebig-University, Giessen, Germany.
PLoS One. 2019 May 10;14(5):e0216642. doi: 10.1371/journal.pone.0216642. eCollection 2019.
LDC3/Dynarrestin, an aminothiazole derivative, is a recently developed small molecule, which binds protein tyrosine phosphatase interacting protein 51 (PTPIP51). PTPIP51 interacts with various proteins regulating different signaling pathways leading to proliferation and migration. Her2 positive breast cancer cells (SKBR3) express high levels of PTPIP51. Therefore, we investigated the effects of LDC3/Dynarrestin on PTPIP51 and its interactome with 12 different proteins of various signal pathways including the interaction with dynein in SKBR3 cells. The localization and semi-quantification of PTPIP51 protein and the Tyr176 phosphorylated PTPIP51 protein were evaluated. Protein-protein-interactions were assessed by Duolink proximity ligation assays. Interactions and the activation of signal transduction hubs were examined with immunoblots. LDC3/Dynarrestin led to an increased PTPIP51 tyrosine 176 phosphorylation status while the overall amount of PTPIP51 remained unaffected. These findings are paralleled by an enhanced interaction of PTPIP51 with its crucial kinase c-Src and a reduced interaction with the counteracting phosphatase PTP1B. Furthermore, the treatment results in a significantly augmented interaction of PTPIP51/14-3-3β and PTPIP51/Raf1, the link to the MAPK pathway. Under the influence of LDC3/Dynarrestin, the activity of the MAPK pathway rose in a concentration-dependent manner as indicated by RTK assays and immunoblots. The novel small molecule stabilizes the RelA/IκB/PTPIP51 interactome and can abolish the effects caused by TNFα stimulation. Moreover, LDC3/Dynarrestin completely blocked the Akt signaling, which is essential for tumor growth. The data were compared to the recently described interactome of PTPIP51 in LDC3/Dynarrestin treated non-cancerous keratinocyte cells (HaCaT). Differences were identified exclusively for the mitochondrial-associated ER-membranes (MAM) interactions and phospho-regulation related interactome of PTPIP51.LDC3/Dynarrestin gives the opportunity/possibility to influence the MAPK signaling, NFkB signaling and probably calcium homeostasis in breast cancer cells by affecting the PTPIP51 interactome.
LDC3/Dynarrestin，一种氨基噻唑衍生物，是一种新开发的小分子，它与蛋白酪氨酸磷酸酶相互作用蛋白 51（PTPIP51）结合。PTPIP51 与调节增殖和迁移的各种信号通路的各种蛋白质相互作用。Her2 阳性乳腺癌细胞（SKBR3）表达高水平的 PTPIP51。因此，我们研究了 LDC3/Dynarrestin 对 PTPIP51 及其与 12 种不同信号通路的蛋白质相互作用体的影响，包括与 SKBR3 细胞中的动力蛋白的相互作用。评估了 PTPIP51 蛋白的定位和半定量以及 Tyr176 磷酸化的 PTPIP51 蛋白。通过 Duolink 邻近连接测定法评估蛋白质-蛋白质相互作用。用免疫印迹法检查信号转导枢纽的相互作用和激活。LDC3/Dynarrestin 导致 PTPIP51 酪氨酸 176 磷酸化状态增加，而 PTPIP51 的总量不受影响。这些发现与 PTPIP51 与其关键激酶 c-Src 的相互作用增强以及与其拮抗磷酸酶 PTP1B 的相互作用减弱相平行。此外，该治疗导致 PTPIP51/14-3-3β 和 PTPIP51/Raf1 的相互作用明显增加，与 MAPK 途径相关。在 LDC3/Dynarrestin 的影响下，MAPK 途径的活性呈浓度依赖性增加，如 RTK 测定和免疫印迹所示。这种新型小分子稳定了 RelA/IκB/PTPIP51 相互作用组，并可以消除 TNFα 刺激引起的作用。此外，LDC3/Dynarrestin 完全阻断了 Akt 信号转导，这对肿瘤生长至关重要。将数据与最近描述的 LDC3/Dynarrestin 处理非癌性角质形成细胞（HaCaT）中 PTPIP51 的相互作用组进行了比较。仅在与线粒体相关的内质网（MAM）相互作用和 PTPIP51 的磷酸化调节相互作用组方面发现了差异。LDC3/Dynarrestin 为通过影响 PTPIP51 相互作用组来影响乳腺癌细胞中的 MAPK 信号转导、NFkB 信号转导和可能的钙稳态提供了机会/可能性。
