COX‐2 mRNA stability in Monocytes by Ligation of RAGE. Novel roles of hnRNPK and miR‐16

Abstract

Advanced glycation end products (AGEs) act via their receptor, RAGE, and play major roles in diabetic complications. We recently showed that AGEs and a RAGE ligand, S100b, can induce the inflammatory cycoloxygenase‐2 (COX‐2) gene in monocytes by both mRNA stability and transcription mechanisms. In this study we examined the mRNA stabilizing mechanisms. S100b treatment of THP‐1 monocytes led to significant accumulation of COX‐2 mRNA at 2 hr. This was mainly due to RNA stability as also confirmed by S100b‐induced increase in luciferase activity of a COX‐2‐3’UTR‐Luc construct. Heterogenous nuclear ribonucleoprotein‐K (hnRNP K) promotes cross‐talk among multiple nuclear factors. RNA immunoprecipitation assays showed that S100b increased binding of hnRNP K to COX‐2 UTR RNA in THP‐1 cells. This was further confirmed by RNA‐EMSA and supershift with hnRNP K antibody. siRNAs to hnRNPK blocked S100b‐induced COX‐2 mRNA stability. Furthermore, micro RNA(miR)‐16 was also involved in S100b induced COX‐2 stability. In untreated cells, siRNA to miR‐16 enhanced COX‐2 3’UTR‐Luc activity and mRNA stability suggesting that miR‐16 can destabilize COX‐2 mRNA. Upon S100b treatment, miR16 levels (QPCR and N.blots) were reduced, while COX‐2 stability was enhanced. These new findings show that diabetic stimuli like AGEs can stabilize inflammatory genes such as COX‐2 by novel RNA regulatory mechanims involving hnRNP K and miR16.