Premature termination codons do not affect the rate of splicing of neighboring introns.
Introduction of a premature termination codon (PTC) into an exon of a gene can lead to nonsense-mediated decay of the mRNA, which is best characterized as a cytoplasmic event. However, increasing evidence has suggested that PTCs may also influence the nuclear processing of an RNA transcript, leading to models of nuclear surveillance perhaps involving translating nuclear ribosomes. We used quantitative RT-PCR to measure the in vivo steady-state levels of every exon-intron junction in wild-type, PTC-containing, and missense-containing precursor mRNAs of both the nonrearranging dihydrofolate reductase (DHFR) and the somatically rearranging Ig- micro genes. We find that each exon-intron junction's abundance and, therefore, the rate of intron removal, is not significantly affected by the presence of a PTC in a neighboring exon in either the DHFR or Ig- micro pre-mRNA. Similarly, the abundance of the uncleaved Ig- micro polyadenylation sites does not differ between wild-type and PTC-containing Ig- micro pre-mRNAs. Our Ig- micro data were confirmed by RNase protection analyses, and multiple cell isolates were examined to resolve differences with previously published data on steady-state pre-mRNA levels. We conclude that the presence of a PTC affects the rate of neither splicing nor the cleavage step of 3' end formation during pre-mRNA processing in the nucleus. Our results are discussed with respect to existing evidence for nuclear surveillance mechanisms.
Lytle
J Robin
JR
Department of Molecular Biophysics and Biochemistry, Howard Hughes Medical Institute, Yale University School of Medicine, New Haven, Connecticut 06536, USA.
Steitz
Joan A
JA
D000818:Animals; D018389:Codon, Nonsense / Q000378:metabolism*; D006801:Humans; D007148:Immunoglobulin mu-Chains / Q000235:genetics; D007438:Introns / Q000502:physiology*; D026723:Polyadenylation / Q000502:physiology; D012322:RNA Precursors / Q000378:metabolism*; D012326:RNA Splicing; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012260:Ribonucleases / Q000378:metabolism; D013762:Tetrahydrofolate Dehydrogenase / Q000235:genetics
D018389:Codon, Nonsense; D007148:Immunoglobulin mu-Chains; D012322:RNA Precursors; D013762:Tetrahydrofolate Dehydrogenase; D012260:Ribonucleases