Kiely Maeve, Adams David R, Hayes Sheri L, O'Connor Rosemary, Baillie George S, Kiely Patrick A
Graduate Entry Medical School, Materials and Surface Science Institute and Health Research Institute, University of Limerick, Ireland.
Institute of Chemical Sciences, Heriot-Watt University, Riccarton Campus, Edinburgh EH14AS, UK.
Cell Signal. 2017 Jul;35:290-300. doi: 10.1016/j.cellsig.2016.09.001. Epub 2016 Sep 4.
Conflicting reports implicate the scaffolding protein RACK1 in the progression of breast cancer. RACK1 has been identified as a key regulator downstream of growth factor and adhesion signalling and as a direct binding partner of PP2A. Our objective was to further characterise the interaction between PP2A and RACK1 and to advance our understanding of this complex in breast cancer cells. We examined how the PP2A holoenzyme is assembled on the RACK1 scaffold in MCF-7 cells. We used immobilized peptide arrays representing the entire PP2A-catalytic subunit to identify candidate amino acids on the C subunit of PP2A that might be involved in binding of RACK1. We identified the RACK1 interaction sites on PP2A. Stable cell lines expressing PP2A with FR69/70AA, R214A and Y218F substitutions were generated and it was confirmed that the RACK1/PP2A interaction is essential to stabilise PP2A activity. We used Real-Time Cell Analysis and a series of assays to demonstrate that disruption of the RACK1/PP2A complex also reduces the adhesion, proliferation, migration and invasion of breast cancer cells and plays a role in maintenance of the cancer phenotype. This work has significantly advanced our understanding of the RACK1/PP2A complex and suggests a pro-carcinogenic role for the RACK1/PP2A interaction. This work suggests that approaches to target the RACK1/PP2A complex are a viable option to regulate PP2A activity and identifies a novel potential therapeutic target in the treatment of breast cancer.
相互矛盾的报道表明支架蛋白RACK1与乳腺癌的进展有关。RACK1已被确定为生长因子和黏附信号下游的关键调节因子，也是PP2A的直接结合伴侣。我们的目标是进一步表征PP2A与RACK1之间的相互作用，并加深我们对乳腺癌细胞中这种复合物的理解。我们研究了PP2A全酶如何在MCF-7细胞中的RACK1支架上组装。我们使用代表整个PP2A催化亚基的固定化肽阵列来识别PP2A C亚基上可能参与RACK1结合的候选氨基酸。我们确定了PP2A上的RACK1相互作用位点。构建了表达具有FR69/70AA、R214A和Y218F替代的PP2A的稳定细胞系，并证实RACK1/PP2A相互作用对于稳定PP2A活性至关重要。我们使用实时细胞分析和一系列试验来证明RACK1/PP2A复合物的破坏也会降低乳腺癌细胞的黏附、增殖、迁移和侵袭，并在维持癌症表型中发挥作用。这项工作显著推进了我们对RACK1/PP2A复合物的理解，并表明RACK1/PP2A相互作用具有促癌作用。这项工作表明，靶向RACK1/PP2A复合物的方法是调节PP2A活性的可行选择，并确定了一种治疗乳腺癌的新的潜在治疗靶点。
