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ABSTRACT: In the protein 4.1R gene, alternative first exons splice differentially to alternative 3' splice sites far downstream in exon 2'/2 (E2'/2). We describe a novel intrasplicing mechanism by which exon 1A (E1A) splices exclusively to the distal E2'/2 acceptor via two nested splicing reactions regulated by novel properties of exon 1B (E1B). E1B behaves as an exon in the first step, using its consensus 5' donor to splice to the proximal E2'/2 acceptor. A long region of downstream intron is excised, juxtaposing E1B with E2'/2 to generate a new composite acceptor containing the E1B branchpoint/pyrimidine tract and E2 distal 3' AG-dinucleotide. Next, the upstream E1A splices over E1B to this distal acceptor, excising the remaining intron plus E1B and E2' to form mature E1A/E2 product. We mapped branchpoints for both intrasplicing reactions and demonstrated that mutation of the E1B 5' splice site or branchpoint abrogates intrasplicing. In the 4.1R gene, intrasplicing ultimately determines N-terminal protein structure and function. More generally, intrasplicing represents a new mechanism by which alternative promoters can be coordinated with downstream alternative splicing.
The EMBO Journal 02/2008; 27(1):122-31. · 9.20 Impact Factor
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ABSTRACT: The human genome uses alternative pre-mRNA splicing as an important mechanism to encode a complex proteome from a relatively small number of genes. An unknown number of these genes also possess multiple transcriptional promoters and alternative first exons that contribute another layer of complexity to gene expression mechanisms. Using a collection of more than 100 erythroid-expressed genes as a test group, we used genome browser tools and genetic databases to assess the frequency of alternative first exons in the genome. Remarkably, 35% of these erythroid genes show evidence of alternative first exons. The majority of the candidate first exons are situated upstream of the coding exons, whereas a few are located internally within the gene. Computational analyses predict transcriptional promoters closely associated with many of the candidate first exons, supporting their authenticity. Importantly, the frequent presence of consensus translation initiation sites among the alternative first exons suggests that many proteins have alternative N-terminal structures whose expression can be coupled to promoter choice. These findings indicate that alternative promoters and first exons are more widespread in the human genome than previously appreciated and that they may play a major role in regulating expression of selected protein isoforms in a tissue-specific manner.
Blood 04/2006; 107(6):2557-61. · 9.90 Impact Factor
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ABSTRACT: Recent studies have shown that transcription and alternative splicing can be mechanistically coupled. In the EPB41 (protein 4.1R) and EPB41L3 (protein 4.1B) genes, we showed previously that promoter/alternative first exon choice is coupled to downstream splicing events in exon 2. Here we demonstrate that this coupling is conserved among several vertebrate classes from fish to mammals. The EPB41 and EPB41L3 genes from fish, bird, amphibian, and mammal genomes exhibit shared features including alternative first exons and differential splice acceptors in exon 2. In all cases, the 5'-most exon (exon 1A) splices exclusively to a weaker internal acceptor site in exon 2, skipping a fragment designated as exon 2'. Conversely, alternative first exons 1B and 1C always splice to the stronger first acceptor site, retaining exon 2'. These correlations are independent of cell type or species of origin. Since exon 2' contains a translation initiation site, splice variants generate protein isoforms with distinct N-termini. We propose that these genes represent a physiologically relevant model system for mechanistic analysis of transcription-coupled alternative splicing.
Genomics 01/2006; 86(6):701-7. · 3.02 Impact Factor
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ABSTRACT: Human seizure disorders are a major health concern due to the large number of affected individuals, the potentially devastating consequences of untreated seizure occurrences, and the lack of an effective treatment for all patients. Although anticonvulsants have proven very helpful in treating seizures and remain the best option available for treatment, not all afflicted individuals respond to medication and many only do so in unique drug combinations or at the cost of adverse side-effects. Therefore, new and more effective anticonvulsants are continually sought after to combat this illness. In this study, we present results which offer the possibility of using Drosophila bang-sensitive (BS) mutants as a tool to screen anticonvulsants. By feeding the BS mutants a known anticonvulsant, potassium bromide, we have demonstrated that the drug dramatically reduces the seizures of bang senseless, the most severe of the BS mutants. This methodology suggests that the Drosophila system can potentially be a powerful instrument for assaying and testing new compounds with anticonvulsant properties.
Brain Research 10/2004; 1020(1-2):45-52. · 2.73 Impact Factor
