Congenital hearing loss affects ~ 1:1000 children, making it one of the most commonly diagnosed sensory birth defects. The long-term objective of this study is to use Xenopus laevis as a vertebrate model organism to investigate the genetic mechanisms of inner ear development and deafness. Targeted alteration of gene expression in Xenopus embryos, which has been successfully used in the investigation of other organ systems, is an ideal method to investigate the developmental dynamics of inner ear formation. We will initially focus on the role of EYA1 in inner ear development, since EYA1 mutations result in branchio-otorenal syndrome (BOR), a common syndromic forms of hearing loss with a unique phenotype that may be reproducible and easily recognizable in a model organism. This study has 3 specific aims: 1) to determine the spatial and temporal developmental expression patterns of EYA1 in the Xenopus laevis inner ear. A comparison will be made of the stage specific expression patterns of both EYA1 mRNA and protein in the inner ear, with specific attention to the presence and functional significance of alternative splicing in EYA1 expression. 2) To perform targeted EYA1 mRNA degradation with modified oligonucleotide injection of Xenopus embryos to determine its molecular and morphologic effects in the developing inner ear. If these experiments result in inner ear abnormalities comparable to that found in BOR, this would indicate haploin sufficiency as a likely mechanism for the development of the BOR phenotype. 3) To model the effects of EYA1 mRNA misexpression by embryo injection experiments involving excess normal message or altered mRNAs corresponding to mutations found in BOR. If inner ear malformations result from inappropriate EYA1 mRNA levels, this would indicate that dominant negative or abnormal protein activity underlies the dominant phenotype of BOR.
| Li, You E; Allen, Bryan G; Weeks, Daniel L (2012) Using ?C31 integrase to mediate insertion of DNA in Xenopus embryos. Methods Mol Biol 917:219-30 |
| Li, Youe; Manaligod, Jose M; Weeks, Daniel L (2010) EYA1 mutations associated with the branchio-oto-renal syndrome result in defective otic development in Xenopus laevis. Biol Cell 102:277-92 |
| Allen, Bryan G; Weeks, Daniel L (2009) Bacteriophage phiC31 integrase mediated transgenesis in Xenopus laevis for protein expression at endogenous levels. Methods Mol Biol 518:113-22 |
| Allen, Bryan G; Weeks, Daniel L (2005) Transgenic Xenopus laevis embryos can be generated using phiC31 integrase. Nat Methods 2:975-9 |
