Transcriptional regulation of platelet-derived growth factor-B chain by thrombin in endothelial cells: involvement of Egr-1 and CREB-binding protein

Abstract

Thrombin and platelet-derived growth factor-B chain (PDGF-B) are key factors in the stimulation of atherosclerosis. The effect of thrombin on PDGF-B production has been characterized. However, the underlying mechanism is still far clear. Here, we investigate the transcription factors and regulators that are involved in PDGF-B production caused by thrombin in endothelial cells (ECs). Levels of PDGF were analyzed by real-time RT-PCR and ELISA, while levels of early growth response-1 (Egr-1) were analyzed by real-time RT-PCR and western blot. To evaluate the function of CBP and Egr-1 involved in regulation of PDGF-B, small interfering RNA (siRNA) were used to down-regulate their expression in mRNA and protein level. Interaction of Egr-1 and CBP was measured with immunoprecipitation and western blot. Thrombin induced an early and transient up-regulation of transcription factor early Egr-1, which was followed by a delayed increase of PDGF-B. siRNA against Egr-1-inhibited thrombin-induced PDGF-B production. Furthermore, thrombin could enhance the interaction of Egr-1 with its co-activator CREB-binding protein (CBP). CBP knockdown attenuated this interaction, and led to a reduction of PDGF-B expression induced by thrombin. Our results suggest that CBP might be one of the main interaction targets for Egr-1, and the transient activation of Egr-1 and recruitment of CBP are required for thrombin-induced PDGF-B in ECs.