Department of General and Gastroenterological Surgery, Osaka Medical College, Osaka, Japan.
Department of General and Gastroenterological Surgery, Osaka Medical College, Osaka, Japan.
Biochim Biophys Acta Mol Basis Dis. 2018 Aug;1864(8):2600-2609. doi: 10.1016/j.bbadis.2018.04.018. Epub 2018 Apr 21.
Nogo-B is a member of the Nogo/Reticulon-4 family and has been reported to be an inducer of apoptosis in certain types of cancer cells. However, the role of Nogo-B in human cancer remains less understood. Here, we demonstrated the functions of Nogo-B in colorectal cancer cells. In clinical colorectal cancer specimens, Nogo-B was obviously overexpressed, as determined by immunohistochemistry; and Western blot analysis showed its expression level to be significantly up-regulated. Furthermore, knockdown of Nogo-B in two colorectal cancer cell lines, SW480 and DLD-1, by transfection with si-RNA (siR) resulted in significantly reduced cell viability and a dramatic increase in apoptosis with insistent overexpression of cleaved caspase-8 and cleaved PARP. The transfection with Nogo-B plasmid cancelled that apoptosis induced by siRNogoB in SW480 cells. Besides, combinatory treatment with siR-Nogo-B/staurosporine (STS) or siR-Nogo-B/Fas ligand (FasL) synergistically reduced cell viability and increased the expression of apoptotic signaling proteins in colorectal cancer cells. These results strongly support our contention that Nogo-B most likely played an oncogenic role in colorectal cancer cells, mainly by negatively regulating the extrinsic apoptotic pathway in them. Finally, we revealed that suppression of Nogo-B caused down-regulation of c-FLIP, known as a major anti-apoptotic protein, and activation of caspase-8 in the death receptor pathway. Interaction between Nogo-B and c-FLIP was shown by immunoprecipitation and immunofluorescence studies. In conclusion, Nogo-B was shown to play an important negative role in apoptotic signaling through its interaction with c-FLIP in colorectal cancer cells, and may thus become a novel therapeutic target for colorectal cancer.
Nogo-B 是 Nogo/Reticulon-4 家族的一员，据报道它能诱导某些类型的癌细胞凋亡。然而，Nogo-B 在人类癌症中的作用还不太清楚。在这里，我们研究了 Nogo-B 在结直肠癌细胞中的功能。通过免疫组织化学法，我们在临床结直肠癌标本中发现 Nogo-B 明显过表达；Western blot 分析显示其表达水平显著上调。此外，通过 siRNA（siR）转染，在两种结直肠癌细胞系 SW480 和 DLD-1 中敲低 Nogo-B，导致细胞活力明显降低，凋亡明显增加，同时 cleaved caspase-8 和 cleaved PARP 的表达持续上调。Nogo-B 质粒的转染取消了 siRNogoB 在 SW480 细胞中诱导的凋亡。此外，siR-Nogo-B/Staurosporine (STS) 或 siR-Nogo-B/Fas Ligand (FasL) 的联合处理协同降低结直肠癌细胞的活力并增加凋亡信号蛋白的表达。这些结果有力地支持了我们的观点，即 Nogo-B 很可能在结直肠癌细胞中发挥致癌作用，主要通过负调控它们的外在凋亡途径。最后，我们揭示了抑制 Nogo-B 导致已知的主要抗凋亡蛋白 c-FLIP 的下调，并激活死亡受体途径中的 caspase-8。免疫沉淀和免疫荧光研究表明 Nogo-B 和 c-FLIP 之间存在相互作用。总之，Nogo-B 通过与结直肠癌细胞中的 c-FLIP 相互作用，在凋亡信号中发挥重要的负调控作用，因此可能成为结直肠癌的新治疗靶点。
