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Molecular analysis of mutations affecting hprt mRNA splicing in human T‐lymphocytes in vivo
Author(s) -
Rossi Anna M.,
Tates Ad D.,
Van Zeeland Albert A.,
Vrieling Harry
Publication year - 1992
Publication title -
environmental and molecular mutagenesis
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1
H-Index - 87
eISSN - 1098-2280
pISSN - 0893-6692
DOI - 10.1002/em.2850190103
Subject(s) - exon , splice site mutation , minigene , rna splicing , biology , intron , genetics , exon skipping , splice , exon trapping , mutant , microbiology and biotechnology , exonic splicing enhancer , alternative splicing , exon shuffling , hypoxanthine guanine phosphoribosyltransferase , genomic dna , mutation , gene , rna
Molecular analysis of hypoxanthine‐guanine phosphoribosyltransferase ( hprt ) cDNA from 6‐thioguanine‐resistant T‐lymphocytes cloned from smoking and non‐smoking adult donors showed that 35% of these mutants were defective in splicing of hprt mRNA. Among a set of 42 hprt splice mutants, we observed i) complete loss of one or more exons, ii) partial loss of one exon, or iii) inclusion of part of an intron sequence between adjacent exons. Loss of exon 4 was significantly more frequent than of the other exons, suggesting that the sequences that regulate splicing of this exon are either larger than those of the other exons or especially prone to mutation. In order to identify the molecular nature of DNA alterations causing aberrant splicing of hprt mRNA, 17 splice mutants were analyzed in more detail by sequencing the genomic regions flanking the mis‐spliced exon. Base pair substitutions or small deletions causing defective splicing were either detected in exon sequences or in splice site consensus sequences of introns. Furthermore, genomic deletions encompassing entire exons were found. In some mutants, the alteration responsible for incorrect splicing could not be identified, suggesting that the target sequence for splice mutations is larger than merely the splice junctions. Molecular characterization of hprt splice mutations will lead to the identification of specific sequences regulating splicing of hprt mRNA and will reveal whether the mutational spectrum in splice mutants is similar to that found in the hprt coding region.