Endothelial SRSF1 directly regulates ATF3-KLF2 pathway to promote ischemia-induced angiogenesis

Peripheral artery disease is a severe ischemic vascular pathology without effective pharmacological approaches and improving angiogenesis to recover blood perfusion is a promising therapeutic strategy. Endothelial cells (ECs) are the primary cell type contributing to angiogenesis in response to ischemia. However, the molecular mechanisms regulating ischemia-induced angiogenesis remain elusive. We used a discovery-driven approach to identify elevated serine/arginine splicing factor 1 (SRSF1) expression in endothelial cells after ischemia. Proteomic analyses identified endogenous SRSF1 accumulated in ECs of the ischemic muscle and responded to hypoxia. Mice deficient in endothelial SRSF1 exhibited impaired blood flow recovery and impaired new capillary formation following HLI. Importantly, overexpression of SRSF1 enhanced blood flow recovery and angiogenesis after HLI. SRSF1 overexpression facilitated, while SRSF1 knockdown suppressed, the angiogenic function of human ECs. To explore the mechanism the mechanisms contributing to SRSF1-promoted endothelial angiogenesis,we conducted transcriptome RNA-seq searching for downstream molecules and verified it through RNA-pull down and chromatin immunoprecipitation qPCR analysis. Mechanistically, SRSF1 modulated ATF3 alternative splicing, wherein SRSF1 directly bound to ATF3 pre-mRNA and elevated full-length ATF3 transcript at the expense of truncated ATF3Δzip2 transcript, and in turn, ATF3 directly bound to the KLF2 promoter and suppressed KLF2-S1PR1 signaling, through which mediated the SRSF1-promoted endothelial tube formation and angiogenesis. Additionally, alprostadil, the prostaglandin E1 analog, could activate the SRSF1 signaling to improve endothelial angiogenesis in vitro and in vivo. Our findings identified SRSF1 as a novel regulator of ischemia-induced angiogenesis that enhances endothelial angiogenic functions by regulating the ATF3-KLF2/SIPR1 pathway. These results suggest that modulation of endothelial SRSF1 may represent a promising therapeutic approach for treating ischemic vascular diseases.