Whole Genome and Embryo Transcriptome Analysis of Vertebrate Identifies nxhl Controlling Angiogenesis by Targeting VE-PTP

BACKGROUND Angiogenesis is closely associated with angiogenesis-dependent diseases including cancers and ocular diseases. Anti-angiogenic therapeutics have been focusing on the (VEGF)/VEGFR signaling axis. However, the clinical resistance, high cost and frequent administration of anti-VEGF drugs make it urgent to discover novel angiogenic pathways.VE-PTP (ptprb) is a novel target with great anti-angiogenic potential. However, it is unclear whether upstream signaling pathways targeting VE-PTP exist in angiogenesis. METHODS Whole genome and embryo transcriptome sequencing were applied to discover the new gene nxhl. Transgenic zebrafish model, morpholino knockdown and small interfering RNA were used to explore the role of nxhl in angiogenesis both in vitro and in vivo. RNA pulldown, RIP and ChIRP-MS were used to identify interactions between RNA and protein. RESULTS We discovered a novel zebrafish gene nxhl which is a homologue of the conserved gene nxh that co-expressed with some key genes essential for embryo development in vertebrate. Nxhl deletion causes angiogenesis defects in embryo. Moreover, nxhl is essential to mediate effects of angiogenesis in vivo and in vitro, and ptprb depletion duplicates the phenotypes of nxhl deficiency. Importantly, nxhl acts upstream of ptprb and regulates many extreme important ptprb-linked angiogenic genes by targeting VE-PTP (ptprb) through interactions with NCL. Notably, nxhl deletion decreases the phosphorylation of NCL T76 and increases the acetylation of NCL K88, suggesting nxhl may regulate downstream VE-PTP signaling pathways by mediation of NCL posttranslational modification. This is the first description of the interaction between nxhl and NCL, NCL and VE-PTP (ptprb), uncovering a novel nxhl-NCL-VE-PTP signaling pathway on angiogenesis regulation. CONCLUSIONS Our study identifies nxhl controlling angiogenesis by targeting VE-PTP through interactions with NCL, uncovering novel upstream controllers of VE-PTP. This nxhl-NCL-VE-PTP pathway may be a therapeutic target in the treatment of angiogenesis-dependent diseases. Clinical Perspective What Is New? We report a novel nxhl-NCL-VE-PTP signaling pathway that controls angiogenesis. We for the first time demonstrate that nxhl interacts with NCL which simultaneously binds to VE-PTP that plays key roles on EC adherens junction, integrity and vascular homeostasis. Nxhl also controls some other crucial VE-PTP-linked downstream angiogenic genes (such as Tie2, VEGFaa, VEGFR2, Erbb2, S1pr1 and Hey2) which explain the phenotypes induced by the nxhl deficiency. Our study indicates the key role of nxhl on controlling angiogenesis as an upstream regulator of VE-PTP. What Are the Clinical Implications? There are several ongoing researches investigating the utility of VE-PTP or NCL inhibitors on treatment of angiogenesis-dependent diseases including a range of cancers and nonneoplastic diseases, such as AMD, DME, RA and atherosclerosis. Targeting the nxhl-NCL-VE-PTP pathway may facilitate therapeutic angiogenesis in patients with cancers or ocular diseases such as DME. Our study highlights the great potential of nxhl on anti-angiogenic therapeutics by targeting VE-PTP.