Regulation of Cyr61/Ccn1 Expression by Hypoxia through Cooperation of C- Jun/Ap-1 and Hif-1 Alpha in Retinal Vascular Endothelial Cells

Hypoxia is the most important factor in the pathogenesis of diabetic retinopathy. Cysteine-rich 61 (Cyr61) is one of the angiogenic factors involved in the development of proliferative diabetic retinopathy (PDR). The aim of this study was to investigate the mechanism of hypoxia-induced Cyr61 expression in retinal vascular endothelial cells. The hypoxia- induced expression of mRNA and protein of Cyr61 was studied in monkey choroidal retinal vascular endothelial (RF /6A) cells. Luciferase reporter assays and electrophoretic mobility shift assays were used to identify the hypoxia responsible region and transcription factors in the Cyr61 promoter. Chromatin immunoprecipitation and immunoprecipitation were performed to study the role of hypoxia-inducible factor (HIF)-1 alpha and c-Jun/activator protein-1 (AP-1) in Cyr61 transcriptional regulation. The results showed that hypoxia significantly induced Cyr61 mRNA and protein expression in RF/6A cells. The effect was mediated through phosphorylation of c-Jun. Luciferase assays , electrophoretic mobility shift assays, chromatin immunoprecipitation and immunoprecipitation showed that HIF- 1 alpha interacted with c-Jun/AP-1 and their binding on the AP-1 binding motif within the Cyr61 promoter induced the expression of Cyr61. In conclusion, hypoxia controlled the transcriptional regulation of the Cyr61 gene in RF/6A cells by cooperation of HIF-1 alpha and c-Jun/AP-1. Cyr61 might play an important role in ischemic retinal diseases, such as PDR.