A novel bipartite intronic splicing enhancer promotes the inclusion of a mini-exon in the AMP deaminase 1 gene.
Alternative splicing of the 12-base exon 2 of the adenosine monophosphate deaminase (AMPD) gene is subject to regulation by both cis- and trans-regulatory signals. The extent of exon 2 inclusion is stage- and cell type-specific and is subject to the physiological state of the cell. In adult skeletal muscle, a cell type that regulates the activity of this allosteric enzyme at several levels, the exon 2-plus form of AMPD, predominates. We have performed a systematic analysis of the cis-acting regulatory sequences that reside in the intron immediately downstream of this mini-exon. A complex element comprising sequences that enhance exon 2 inclusion and sequences that counteract this effect resides in the middle of this intron. We demonstrate that the enhancing component is bipartite, with more than a kilobase of sequence separating the two functional sites. The presence of even minimal levels the mini-exon in the fully processed AMPD mRNA requires both of these sites, neither of which appears in any other published splicing enhancer. An RNA binding activity derived from a muscle cell line requires both of the enhancing sites. Mutations in either of the sites that eliminate exon 2 inclusion abrogate this binding activity.
Genetta
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Joseph P. Stokes Research Institute, Children's Hospital of Philadelphia, Philadelphia, PA 19104, USA.
Morisaki
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Morisaki
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Holmes
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D016475:3T3 Cells; D000659:AMP Deaminase / Q000235:genetics*; D017398:Alternative Splicing; D000818:Animals; D001483:Base Sequence; D004742:Enhancer Elements, Genetic; D005091:Exons / Q000235:genetics*; D007438:Introns; D051379:Mice; D008807:Mice, Inbred BALB C; D008969:Molecular Sequence Data; D009132:Muscles / Q000166:cytology / Q000378:metabolism; D016297:Mutagenesis, Site-Directed; D012313:RNA / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction
D012313:RNA; D000659:AMP Deaminase