Semaan Crystal, Henderson Beric R, Molloy Mark P
Department of Chemistry and Biomolecular Sciences, Macquarie University, Sydney, Australia.
Gene and Stem Cell Therapy Program Centenary Institute, The University of Sydney, Camperdown, 2050, NSW, Australia.
Biochem Biophys Rep. 2019 Jul 10;19:100662. doi: 10.1016/j.bbrep.2019.100662. eCollection 2019 Sep.
Beta-catenin is well-known as a key effector of Wnt signalling and aberrant expression is associated with several human cancers. Stabilisation of and atypical subcellular localisation of beta-catenin, regulated in part through specific protein-protein interactions has been linked to cancer development, however the mechanisms behind these pathologies is yet to be fully elucidated. Affinity purification and mass spectrometry were used to identify potential β-catenin interacting proteins in SW480 colon cancer cells. Recombinant β-catenin constructs were used to co-isolate interacting proteins from stable isotope labelled cells followed by detection using mass spectrometry. Several known and new putative interactors were observed. In particular, we identified interaction with a set of coatomer complex I subunits implicated in retrograde transport at the Golgi, and confirmed endogenous interaction of β-catenin with coatomer subunit COPB using immunoprecipitation assays and immunofluorescence microscopy. These observations suggest a hitherto unrecognised role for β-catenin in the secretory pathway and warrant further functional studies to unravel its activity at this cellular location.
β-连环蛋白是众所周知的Wnt信号传导的关键效应因子，其异常表达与多种人类癌症相关。β-连环蛋白的稳定化和非典型亚细胞定位，部分通过特定的蛋白质-蛋白质相互作用进行调节，这与癌症发展有关，然而这些病理背后的机制尚未完全阐明。亲和纯化和质谱法用于鉴定SW480结肠癌细胞中潜在的β-连环蛋白相互作用蛋白。重组β-连环蛋白构建体用于从稳定同位素标记的细胞中共分离相互作用蛋白，随后使用质谱法进行检测。观察到了几种已知的和新的假定相互作用蛋白。特别地，我们鉴定出与一组参与高尔基体逆行转运的外套膜蛋白复合物I亚基存在相互作用，并使用免疫沉淀试验和免疫荧光显微镜证实了β-连环蛋白与外套膜蛋白亚基COPB的内源性相互作用。这些观察结果表明β-连环蛋白在分泌途径中具有迄今未被认识的作用，需要进一步的功能研究来揭示其在该细胞位置的活性。
