Abstract
BackgroundThe 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.MethodsSixty-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.ResultsThe 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.ConclusionOur results reveal that the IRE1α-XBP1 pathway regulates Mel-RMu cell proliferation and progression by activating IL-6/STAT3 signaling.
Highlights
The IRE1α-X-box-binding protein 1 (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
Immunohistochemical staining to determine XBP1s expression in human melanoma tissues To investigate the expression of XBP1s in human melanoma, immunohistochemical staining for XBP1s was conducted on a melanoma TMA containing 61 pairs of melanoma specimens and corresponding normal tissues
The inositol-requiring enzyme 1 (IRE1)/XBP1 pathway regulates interleukin‐6 expression in melanocytes and melanoma cell lines The results shown in Fig. 1 indicate that the IRE1α-XBP1 pathway is constitutively activated in human melanoma tissues
Summary
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. The role of this pathway in melanoma cell growth remains unclear. One of the most malignant tumors, is increasing in incidence worldwide. The UPR of mammalian cells is initiated by three ER transmembrane proteins: activating transcription factor 6 (ATF6), inositol-requiring enzyme 1 (IRE1), and doublestranded RNA-activated protein kinase-like ER kinase (PERK), which act as proximal sensors of ER stress. Under ER stress, sequestration of GRP78 by unfolded proteins activates these sensors by inducing the phosphorylation and homodimerization of IRE1 and PERK along with the relocalization of ATF6 to the Golgi, where it is cleaved by Site 1 and 2 proteases, leading to its activation as a transcriptional factor [7,8,9]. IRE1 catalyzes the non-conventional splicing of the mRNA encoding X-box-binding protein 1 (XBP1) by removing a 26 nt intron, thereby producing an active spliced form (XBP1s), initiating an essential UPR program [10]
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