Dataset Information

Dual-specificity splice sites function alternatively as 5' and 3' splice sites.

ABSTRACT: As a result of large-scale sequencing projects and recent splicing-microarray studies, estimates of mammalian genes expressing multiple transcripts continue to increase. This expansion of transcript information makes it possible to better characterize alternative splicing events and gain insights into splicing mechanisms and regulation. Here, we describe a class of splice sites that we call dual-specificity splice sites, which we identified through genome-wide, high-quality alignment of mRNA/EST and genome sequences and experimentally verified by RT-PCR. These splice sites can be alternatively recognized as either 5' or 3' splice sites, and the dual splicing is conceptually similar to a pair of mutually exclusive exons separated by a zero-length intron. The dual-splice-site sequences are essentially a composite of canonical 5' and 3' splice-site consensus sequences, with a CAG|GURAG core. The relative use of a dual site as a 5' or 3' splice site can be accurately predicted by assuming competition for specific binding between spliceosomal components involved in recognition of 5' and 3' splice sites, respectively. Dual-specificity splice sites exist in human and mouse, and possibly in other vertebrate species, although most sites are not conserved, suggesting that their origin is recent. We discuss the implications of this unusual splicing pattern for the diverse mechanisms of exon recognition and for gene evolution.

SUBMITTER: Zhang C

PROVIDER: S-EPMC1986607 | biostudies-literature | 2007 Sep

REPOSITORIES: biostudies-literature

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Dual-specificity splice sites function alternatively as 5' and 3' splice sites.

Zhang Chaolin C Hastings Michelle L ML Krainer Adrian R AR Zhang Michael Q MQ

Proceedings of the National Academy of Sciences of the United States of America 20070911 38

As a result of large-scale sequencing projects and recent splicing-microarray studies, estimates of mammalian genes expressing multiple transcripts continue to increase. This expansion of transcript information makes it possible to better characterize alternative splicing events and gain insights into splicing mechanisms and regulation. Here, we describe a class of splice sites that we call dual-specificity splice sites, which we identified through genome-wide, high-quality alignment of mRNA/EST ...[more]

PMID: 17848517

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Project description:UnlabelledHerpesviruses have large and complex DNA genomes. The largest among the herpesviruses, those of the cytomegaloviruses, include over 170 genes. Although most herpesvirus gene products are expressed from unspliced transcripts, a substantial number of viral transcripts are spliced. Some viral transcripts are subject to alternative splicing, which leads to the expression of several proteins from a single gene. Functional analysis of individual proteins derived from an alternatively spliced gene is difficult, as deletion and nonsense mutagenesis, both common methods used in the generation of viral gene knockout mutants, affect several or all gene products at the same time. Here, we show that individual gene products of an alternatively spliced herpesvirus gene can be inactivated selectively by mutagenesis of the splice donor or acceptor site and by intron deletion or substitution mutagenesis. We used this strategy to dissect the essential M112/113 gene of murine cytomegalovirus (MCMV), which encodes the MCMV Early 1 (E1) proteins. The expression of each of the four E1 protein isoforms was inactivated individually, and the requirement for each isoform in MCMV replication was analyzed in fibroblasts, endothelial cells, and macrophages. We show that the E1 p87 isoform, but not the p33, p36, and p38 isoforms, is essential for viral replication in cell culture. Moreover, the presence of one of the two medium-size isoforms (p36 or p38) and the presence of intron 1, but not its specific sequence, are required for viral replication. This study demonstrates the usefulness of splice site mutagenesis for the functional analysis of alternatively spliced herpesvirus genes.ImportanceHerpesviruses include up to 170 genes in their DNA genomes. The functions of most viral gene products remain poorly defined. The construction of viral gene knockout mutants has thus been an important tool for functional analysis of viral proteins. However, this strategy is of limited use when viral gene transcripts are alternatively spliced, leading to the expression of several proteins from a single gene. In this study, we showed, as a proof of principle, that each protein product of an alternatively spliced gene can be eliminated individually by splice site mutagenesis. Mutant viruses lacking individual protein products displayed different phenotypes, demonstrating that the products of alternatively spliced genes have nonredundant functions.

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Dual-specificity splice sites function alternatively as 5' and 3' splice sites.

Publications

Dual-specificity splice sites function alternatively as 5' and 3' splice sites.

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