Purchase this article with an account.
B. Appukuttan, T. J. McFarland, A. J. Stempel, Y. Zhang, M. H. Davies, J. B. Kassem, M. R. Powers, J. T. Stout; Analysis of RTEF-1 Isoforms in a Mouse Model of Laser-Induced Choroidal Neovascularization. Invest. Ophthalmol. Vis. Sci. 2009;50(13):1173.
Download citation file:
© ARVO (1962-2015); The Authors (2016-present)
Recently we identified novel alternatively spliced isoforms of the TEA domain transcription factor RTEF-1 within human ocular vascular endothelial cells and mouse neural retinal tissue. We determined that certain human RTEF-1 isoforms (1305, 936, 447) were capable of enhancing expression of the vascular endothelial growth factor (VEGF) gene whereas another novel isoform (651) can repress production of VEGF. The production of VEGF mediates angiogenesis and can drastically influence the development of choroidal neovascularization (CNV). In this study we investigated whether RTEF-1 is present and alternatively spliced within the choroid of a laser induced mouse model of CNV.
Mice (C57BL/6) between the ages of 6 to 8 weeks old were subjects. A 532-nm argon green laser was used to produce 4 laser burn spots on the retina in both eyes of a mouse. Eyes were enucleated at 3, 6, 24 and 72 hours after laser irradiation, dissected and the neural retina and choroidal tissues were isolated. Total RNA was isolated from choroid and neural retinal tissue separately and cDNA prepared. Consensus primers spanning the TEA/ATTS DNA binding (TEA), proline rich (PRD) and two STY domain were designed and RT-PCR was performed. Amplified products were purified from an agarose gel and sequenced.
Twelve exons are predicted to code for the full length 1305 bp RTEF-1 mRNA, with exons 2 to 10 containing the TEA, PRD and both STY domains. RT-PCR analysis of choroidal tissue at the 3 hour time point indicated the presence of a 542bp product amplified with primers designed to exon 1 and exon 10. This 542bp product was present in untreated choroid but absent from the lasered eye. Sequencing confirmed that this product is another novel alternatively spliced RTEF-1 isoform, which lacks exons 5-9 resulting in loss of the PRD and one STY domain. This particular isoform has not been identified in human tissue and may be a murine ocular-specific isoform.
A novel murine specific isoform of RTEF-1 exists within the choroid. This isoform shows most similarity to the human 651 isoform that can inhibit transcriptional activity of the VEGF gene. Whether this new mouse isoform can influence VEGF gene expression is of interest and has implications for the regulation of angiogenesis in the eye.
This PDF is available to Subscribers Only