title: A silent mutation induces exon skipping in the phenylalanine hydroxylase gene in phenylketonuria.
abstract: An A-->T substitution in cDNA nucleotide 1197 (c.1197A/T) of the human phenylalanine hydroxylase (PAH) gene has been regarded as a silent mutation, because both the wild-type (GUA) and the mutant (GUU) alleles encode a valine residue at codon 399 (V399 V). The nucleotide c.1197 is located at the 3'-end of exon 11at position -3 of the exon-intron junction. To explore whether the substitution exerts any effects on the processing of the PAH mRNA, illegitimate PAH transcripts from lymphoblast cultures of a phenylketonuria (PKU) patient heterozygous for c.1197A/T were analyzed by the polymerase chain reaction following reverse-transcription (RT-PCR). mRNAs with an exon 11 deletion were revealed. Furthermore, by using an R408 W mutation in the paternal allele as a marker, sequence analysis of the RT-PCR products indicates that virtually all PAH transcripts from the maternal allele with the c. 1197A/T substitution do not contain exon 11. To address whether this substitution is the main determinant for exon skipping, PAH minigenes with or without the substitution were constructed and transfected to a human hepatoma cell line. Analysis of the transcription products by S1 nuclease mapping clearly indicated that such exon 11 skipping was directly associated with the c.1197A/T substitution. Thus, this study demonstrates that the c.1197A/T substitution in the PAH gene is not just a neutral polymorphism but a mutation that induces post-transcriptional skipping of exon 11 leading to a PKU phenotype.
authors.affiliation: Department of Medical Research and Education, Veterans General Hospital-Taipei, Taiwan, Republic of China.
authors.forename: H K
authors.initials: HK
authors.lastname: Chao
authors.forename: K J
authors.initials: KJ
authors.lastname: Hsiao
authors.forename: T S
authors.initials: TS
authors.lastname: Su
mesh_terms: D001483:Base Sequence; D005091:Exons; D006801:Humans; D009154:Mutation; D010651:Phenylalanine Hydroxylase / Q000235:genetics*; D010661:Phenylketonurias / Q000201:enzymology / Q000235:genetics*; D012333:RNA, Messenger / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014162:Transfection; D014407:Tumor Cells, Cultured
chemical_list: D012333:RNA, Messenger; D010651:Phenylalanine Hydroxylase