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B. Appukuttan, T.J. McFarland, Y. Zhang, B. Babra, M.H. Davies, L.–O. Atchaneeyasakul, M.R. Powers, J.T. Rosenbaum, J.T. Stout; Characterization of Novel mRNA Isoforms of TEA Domain Transcription Factors Within the Mammalian Eye . Invest. Ophthalmol. Vis. Sci. 2005;46(13):3117.
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© ARVO (1962-2015); The Authors (2016-present)
Purpose:The TEA/ATTS DNA binding domain gene family is highly conserved from Drosophila, mice to human. Expression of specific members of these genes has been identified in various mammalian tissues, including heart, skeletal muscle, pancreas, brain and lung. Isoforms, of these transcription factors, arising from alternative splicing of mRNA from a single gene have been identified within a single tissue. This family of transcription factors is responsible for activation of an assortment of genes. However the expression profile of any of these genes within the mammalian eye has not been reported. We aim to identify whether any genes from the TEA/ATTS DNA binding family are expressed within human retinal and iris endothelial cells, mouse neural retina and whether alternative spliced isoforms exist, and characterize the function of these isoforms within the eye. Methods: Primary culture of vascular endothelial cells isolated from human retina and iris were established and used as a source for RNA. Total RNA was isolated from the neural retina of C57BL mice. Consensus primers spanning the TEA/ATTS DNA binding domain were designed and RT–PCR was performed. Amplified products were purified from an agarose gel and sequenced. Results: RT–PCR from human ocular endothelial cells gave one major product of 1300bp, and two minor products at 900bp and 600bp. RT–PCR from mouse neural retina gave 2 products, one at 1,220bp and a smaller band at 1091bp. Sequencing revealed that the 900bp and the 600bp transcripts from human RNA are alternate spliced products of the 1300bp transcript. Four of the 11 exons predicted to code for the 1300bp transcript are spliced out of the 900bp transcript. An unusual in frame splice event within the middle of exon 3 joins the middle of exon 10 for the 600bp transcript, resulting in loss of part of the TEA domain and loss of a putative nuclear localization signal. The 1091bp mouse product is an alternative spliced version of the larger 1,220bp mouse transcript. The mouse 1091bp transcript is lacking a 129bp sequence that corresponds in exact size and sequence homology to exon 5 of the human ocular endothelial product. Conclusions: A gene from the TEA/ATTS DNA family is expressed within the mammalian retina. Alternative isoforms exist that may confer different transcriptional activity. The predicted protein from the 600bp transcript from endothelial cells is unlikely to bind to DNA or be transported into the nucleus. Which genes these transcription factors activate within the retina is under investigation.
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