Abstract
Abstract: :
Background:AIPL1 is the fourth gene involved in Leber’s Congenital Amaurosis (LCA4), the most frequent cause of congenital blindness in children. Mutations in AIPL1 cause a severe form of LCA, including earliest onset and rapid loss of vision. Methods:Full length transcripts of AIPL1 were amplified by RT–PCR from human and mammalian neuroretina. Samples were obtained from female and male human donors between 20 and 90 years of age, who underwent surgery for trauma of orbital bones or extended facial bone tumors. RT–PCR products were subcloned and sequencing confirmed the completeness of the plasmid inserts. Evolutionary conservation of splice–variations was tested by RT–PCR using murine, rat, porcine and bovine retinal RNA preparations. Results:At least 4 different human transcripts of AIPL1 were identified at various amounts by RT–PCR. The major variant corresponded to the full–length transcript. The minor variants were present in approximately 20% of the gene’s transcripts and lacked either the entire exons 2 or 3 or the first parts of exons 2 or 5 respectively. Examination of AIPL1 transcription in several mammals demonstrated alternative splicing in all those species with both splice variants either identical to or different from human variants. Conclusions: The identified splice variants present transcriptional in–frame deletions of the protein coding region, presumably giving rise to different isoforms of the AIPL1 protein. Interestingly, the alternative splicing events seem to affect important protein functions since the peptidyl–prolyl–isomerase domain and the binding site of cell cycle regulator NUB1 are involved. We expect the respective alternative AIPL1 isoforms to fulfill individual functions and even possess mutually exclusive chaperone activity and NUB1 interaction ability. Conservation throughout evolution implies a functional importance of certain splice variants which we isolated from human, porcine, murine, and rat retina. To support our hypothesis, we are currently producing isoform specific antibodies to evaluate cell specific expression and/or isoform specific localization within photoreceptor cells.
Keywords: retinal degenerations: hereditary • gene/expression • transcription