LAUSR

Abstract The X‐box binding protein 1 (XBP1) is a pivotal transcription factor in the endoplasmic reticulum stress response. Our previous studies have proven that XBP1 is involved in vascular endothelial growth factor (VEGF)‐mediated endothelial cell (EC) proliferation and angiogenesis. In this study, we used EC monolayer wound healing, tube formation and transwell migration models to explore the role of XBP1splicing in EC migration. We found that scratching on EC monolayer triggered XBP1splicing, which was attenuated by the presence of SU5416and LY294002, suggesting that VEGF signalling pathways may be involved. Over‐expression of the spliced XBP1 (XBP1s) via Ad‐XBP1s gene transfer increased while knockdown of IRE1&agr;or XBP1 by ShRNA lentivirus suppressed EC migration. Over‐expression of XBP1s up‐regulated the nitric oxide synthase 3 (NOS3)mRNA through the 3’UTR‐mediated stabilisation and increased eNOS protein translation. Further experiments demonstrated that miR‐24 participated in the XBP1s‐induced eNOSup‐regulation and EC migration. Further co‐IP and immunofluorescence staining assays revealed that protein kinase B (Akt), eNOS and XBP1s form a complex, resulting in Akt and eNOS nucleus relocation. These results suggest that XBP1 splicing can regulate eNOS expression and cellular location, leading to EC migration and therefore contributing to wound healing and angiogenesis. HighlightsWound in EC monolayer induced XBP1 splicing, which was involved in EC migration.XBP1s increased NOS3 mRNA stability via down‐regulating microRNA miR‐24.ENOS was involved in EC migration via forming a complex with AKT and XBP1s. Abbreviations: XBP1: X‐box binding protein 1; VEGF: vascular endothelial growth factor; EC: endothelial cell; XBP1s: XBP1; NOS3: nitric oxide synthase 3; (Ad‐XBP1s): adenovirus ‐XBP1s; Akt: Protein kinase B; NOS3: nitric oxide synthase 3; NO: nitric oxide; bZIP: basic‐region leucine zipper; IRE1&agr;: inositol requiring enzyme 1 alpha; ER: endoplasmic reticulum; XBP1u: unsplicedXBP1; HUVEC: human umbilical vein EC; pGL3‐NOS3‐Luc reporter: NOS3 promoter reporter expression; ATF6N: mature activating transcription factor 6; L‐NAME: NG‐nitro‐L‐arginine methyl ester; KDR: the kinase insert domain receptor; Dox/: doxycycline; DMSO, DM: Dimethyl sulfoxide.