Ablation of endothelial <i>Atg7</i> inhibits ischemia-induced angiogenesis by upregulating <i>Stat1</i> that suppresses <i>Hif1a</i> expression.

Ischemia-induced angiogenesis is critical for blood flow restoration and tissue regeneration, but the underlying molecular mechanism is not fully understood. ATG7 (autophagy related 7) is essential for classical degradative macroautophagy/autophagy and cell cycle regulation. However, whether and how ATG7 influences endothelial cell (EC) function and regulates post-ischemic angiogenesis remain unknown. Here, we showed that in mice subjected to femoral artery ligation, EC-specific deletion of <i>Atg7</i> significantly impaired angiogenesis, delayed the recovery of blood flow reperfusion, and displayed reduction in HIF1A (hypoxia inducible factor 1 subunit alpha) expression. In addition, in cultured human umbilical vein endothelial cells (HUVECs), overexpression of <i>HIF1A</i> prevented <i>ATG7</i> deficiency-reduced tube formation. Mechanistically, we identified STAT1 (signal transducer and activator of transcription 1) as a transcription suppressor of <i>HIF1A</i> and demonstrated that ablation of <i>Atg7</i> upregulated STAT1 in an autophagy independent pathway, increased STAT1 binding to <i>HIF1A</i> promoter, and suppressed <i>HIF1A</i> expression. Moreover, lack of ATG7 in the cytoplasm disrupted the association between ATG7 and the transcription factor ZNF148/ZFP148/ZBP-89 (zinc finger protein 148) that is required for STAT1 constitutive expression, increased the binding between ZNF148/ZFP148/ZBP-89 and KPNB1 (karyopherin subunit beta 1), which promoted ZNF148/ZFP148/ZBP-89 nuclear translocation, and increased STAT1 expression. Finally, inhibition of STAT1 by fludarabine prevented the inhibition of HIF1A expression, angiogenesis, and blood flow recovery in <i>atg7</i> KO mice. Our work reveals that lack of ATG7 inhibits angiogenesis by suppression of HIF1A expression through upregulation of STAT1 independently of autophagy under ischemic conditions, and suggest new therapeutic strategies for cancer and cardiovascular diseases.<b>Abbreviations</b>: ATG5: autophagy related 5; ATG7: autophagy related 7; <i>atg7</i> KO: endothelial cell-specific <i>atg7</i> knockout; BECN1: beclin 1; ChIP: chromatin immunoprecipitation; CQ: chloroquine; ECs: endothelial cells; EP300: E1A binding protein p300; HEK293: human embryonic kidney 293 cells; HIF1A: hypoxia inducible factor 1 subunit alpha; HUVECs: human umbilical vein endothelial cells; IFNG/IFN-&#x3b3;: Interferon gamma; IRF9: interferon regulatory factor 9; KPNB1: karyopherin subunit beta 1; MAP1LC3A: microtubule associated protein 1 light chain 3 alpha; MEFs: mouse embryonic fibroblasts; MLECs: mouse lung endothelial cells; NAC: N-acetyl-l-cysteine; NFKB1/NF&#x3ba;B: nuclear factor kappa B subunit 1; PECAM1/CD31: platelet and endothelial cell adhesion molecule 1; RELA/p65: RELA proto-oncogene, NF-kB subunit; ROS: reactive oxygen species; SP1: Sp1 transcription factor; SQSTM1/p62: sequestosome 1; STAT1: signal transducer and activator of transcription 1; ULK1: unc-51 like autophagy activating kinase 1; <i>ulk1</i> KO: endothelial cell-specific <i>ulk1</i> knockout; VSMCs: mouse aortic smooth muscle cells; WT: wild type; ZNF148/ZFP148/ZBP-89: zinc finger protein 148.