The calcitonin exon and its flanking intronic sequences are sufficient for the regulation of human calcitonin/calcitonin gene-related peptide alternative RNA splicing.
The primary transcript of the calcitonin (CT)/calcitonin gene-related peptide (CGRP) is alternatively spliced in a cell-specific fashion to produce CT in thyroid C cells and CGRP in neuronal cells. The key step in this regulatory process is the recognition and inclusion of exon 4 to produce CT mRNA or nonrecognition and exclusion of exon 4 to produce CGRP mRNA. To determine whether inclusion/exclusion of CT exon is regulated independently of its position, we created a series of minigene constructs containing decreasing amounts of CT gene sequence. A human glioblastoma cell line, T98G, was tested and used as a CT exon exclusion cell line, while HeLa cells were used as a CT exon inclusion cell line. CT exon inclusion/exclusion was regulated when either the relative position of exon 4 within the CT gene was changed or when the exon with flanking sequence was inserted into a completely heterologous gene. Our results demonstrate that CT exon functions as a unit in a position-independent fashion in regulating its own inclusion/exclusion. We believe that the heterologous fusion gene containing only exon 4 and part of its flanking intron sequences will be useful for further defining the sequence elements involved in the regulation of CT/CGRP splicing.
Lou
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Department of Medical Specialties, University of Texas M.D. Anderson Cancer Center, Houston 77030.
Cote
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GJ
Gagel
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D017398:Alternative Splicing; D001483:Base Sequence; D002116:Calcitonin / Q000235:genetics*; D015740:Calcitonin Gene-Related Peptide / Q000235:genetics*; D005091:Exons; D005786:Gene Expression Regulation; D005909:Glioblastoma; D006367:HeLa Cells; D006801:Humans; D007438:Introns; D008969:Molecular Sequence Data; D016296:Mutagenesis; D016133:Polymerase Chain Reaction; D012333:RNA, Messenger / Q000378:metabolism; D012194:RNA-Directed DNA Polymerase; D014162:Transfection; D014407:Tumor Cells, Cultured
D012333:RNA, Messenger; D002116:Calcitonin; D012194:RNA-Directed DNA Polymerase; D015740:Calcitonin Gene-Related Peptide