Gene Regulation And Interference In Developing Chicken Inner Ear

The decision of the cell fate specification in the developing vertebrate inner ear is dependent on complex regulatory genetic pathways involving in the Wnt families. Axin2 is a negative regulator of the canonical Wnt pathway that suppresses signal transduction by promoting degradation of β-Catenin. We have identified a chickhomolog gene, cAxin2, which is expressed in the otic cup beginning at stage HH14+ and exhibits a dynamic expression pattern during formation and further differentiation of the otic vesicle. In this study, we explored the possibility of using RNAinterference (RNAi) -induced targeted gene silencing introduced by in ovo electroporation to manipulate gene expression in the otic vesicle. We have demonstrated that a strong enhanced green fluorescent protein (EGFP) signal can be detected in the otic vesicle after transfection with pEGFP-cAxin2-shRNA constructs by in ovo electroporation. The further investigation with in situ hybridization forgenes cAxin2, Pared box2(cPax2), and bone morphogenetic protein 4(cBMP4) showthat the cAxin2 expression was inhibited by short hairpin RNA (shRNA), cPax2 wasdown regulated and cBMP4 was up regulated as well as an abnormally enlarged otic vesicle in the transfected otic vesicle. Our results suggest that cAxin2 is important molecular required for inner ear development.