Study the mechanism of HuR-mediated regulation on FGF9 expression under hypoxia

Abstract

Fibroblast growth factor 9 (FGF9) is an autocrine/paracrine growth factor and involved in many important physiological processes The expression of FGF9 is known to be under tightly controlled and aberrant expression of FGF9 was associated with many human diseases including cancers Our pervious study showed that AUF1 an AU rich element binding protein (ARE-BP) binds to FGF9 3’UTR and decreases FGF9 mRNA stability HuR is another ARE-BP that functions as a stabilizing factor to regulate mRNA turnover and translational efficiency in stress conditions We hypothesized that HuR may bind to FGF9 ARE to increase FGF9 expression under stress conditions like hypoxia The objective of this study is to characterize the mechanism of HuR-mediated regulation on FGF9 expression under hypoxia Using RNA-Immunoprecipitation and pull down assays our data demonstrated that HuR specifically binds to the second AUUUA element of FGF9 ARE Furthermore firefly luciferase reporter assay and ELISA showed that overexpression of HuR increased the FGF9 3’UTR-mediated reporter gene activity and endogenous FGF9 expression under hypoxia Using mRNA stability assay our data showed that changing the expression of HuR influences FGF9 mRNA half-life but the dynamic interaction between HuR and FGF9 mRNA was changed under hypoxia As HuR is a shuttle protein to carry HuR-bound RNA transporting between nuclear and cytoplasm we examined cellular distributions of HuR in normoxia and hypoxia conditions and our data showed that cytoplasm HuR was significantly increased in response to hypoxia Using RNA interference to knockdown HuR we demonstrated that HuR and AUF1 are collaboratively control FGF9 expression under normoxia However hypoxia altered the interplay between HuR and AUF1 and these two proteins turns to compete for FGF9 ARE binding and consequently HuR carries FGF9 mRNA to the cytoplasm for translation and thus increase FGF9 protein expression Results from this study provide more insights on post-transcriptional regulation of FGF9 mRNA to fine tune FGF9 homeostasis under hypoxia

Date of Award	2014 Aug 26
Original language	English
Supervisor	Hsiao-Fang Sun (Supervisor)