Chen Cheng, Zhang Xuejun
Department of Plastic Surgery, Zhongshan Hospital, Fudan University, Shanghai, 200032, China.
J Transl Med. 2017 Feb 21;15(1):42. doi: 10.1186/s12967-017-1147-2.
The IRE1α-XBP1 pathway is the most conserved branch of the unfolded protein response pathways, which are activated during endoplasmic reticulum (ER) stress caused by the accumulation of unfolded/misfolded proteins in the ER lumen. The IRE1α-XBP1 pathway plays a critical role in various cancers. However, the role of this pathway in melanoma cell growth remains unclear.
Sixty-one pairs of melanoma specimens and corresponding normal tissues from patients were stained with XBP1. Then, XBP1 splicing levels were detected in human tissues and cell lines at the mRNA level. IL-6 expression levels were determined in both melanocytes (HEMn-MP) and melanoma cells (Mel-RMu) overexpressing the spliced form of XBP1 (XBP1s). IL-6 expression was also examined in 4μ8C-treated HEMn-MP and Mel-RMu cells overexpressing IRE1α. Next, we analyzed potential XBP1s binding sites within the IL-6 promoter and conducted ChIP experiments. IL-6/STAT3 signaling was detected by western blotting. Melanoma cell proliferation was examined by CCK8 and BrdU assays.
The mRNA and protein expression levels of XBP1s were significantly elevated in human melanoma tissues and cell lines compared with normal tissues or melanocytes, thus indicating the activation of the IRE1α-XBP1 branch in melanoma. Ectopic expression of IRE1α or XBP1s robustly enhanced IL-6 expression in HEMn-MP and Mel-RMu cells. Moreover, the inhibition of the RNase activity of IRE1α also abolished the effect of IRE1α in promoting IL-6 expression. Mechanistically, XBP1 binds the IL-6 promoter and activates its expression. Furthermore, secreted IL-6 functions in an autocrine/paracrine manner, activates the intracellular JAK/STAT3 pathway and promotes the proliferation of melanoma cells.
Our results reveal that the IRE1α-XBP1 pathway regulates Mel-RMu cell proliferation and progression by activating IL-6/STAT3 signaling.
IRE1α-XBP1通路是未折叠蛋白反应通路中最保守的分支，在内质网（ER）腔中未折叠/错误折叠蛋白积累导致的内质网应激期间被激活。IRE1α-XBP1通路在多种癌症中起关键作用。然而，该通路在黑色素瘤细胞生长中的作用仍不清楚。
用XBP1对61例患者的黑色素瘤标本及相应正常组织进行染色。然后，在mRNA水平检测人组织和细胞系中的XBP1剪接水平。在过表达XBP1剪接形式（XBP1s）的黑素细胞（HEMn-MP）和黑色素瘤细胞（Mel-RMu）中测定IL-6表达水平。还在4μ8C处理的过表达IRE1α的HEMn-MP和Mel-RMu细胞中检测IL-6表达。接下来，我们分析了IL-6启动子内潜在的XBP1s结合位点并进行了染色质免疫沉淀实验。通过蛋白质印迹法检测IL-6/STAT3信号传导。通过CCK8和BrdU试验检测黑色素瘤细胞增殖。
与正常组织或黑素细胞相比，人黑色素瘤组织和细胞系中XBP1s的mRNA和蛋白表达水平显著升高，表明黑色素瘤中IRE1α-XBP1分支被激活。IRE1α或XBP1s的异位表达在HEMn-MP和Mel-RMu细胞中强烈增强IL-6表达。此外，IRE1α核糖核酸酶活性的抑制也消除了IRE1α促进IL-6表达的作用。机制上，XBP1结合IL-6启动子并激活其表达。此外，分泌的IL-6以自分泌/旁分泌方式发挥作用，激活细胞内JAK/STAT3通路并促进黑色素瘤细胞增殖。
我们的结果表明，IRE1α-XBP1通路通过激活IL-6/STAT3信号传导调节Mel-RMu细胞增殖和进展。
