Selective degradation of nonsense beta-phosphodiesterase mRNA in the heterozygous rd mouse.
OBJECTIVE
To investigate the molecular mechanism relating phenotype and genotype in the rd mouse, mRNA and pre-mRNA levels derived from the wild-type and position-347 nonsense mutant beta-phosphodiesterase (beta-PDE) genes were determined and compared with the corresponding gene copy ratios.


METHODS
Total RNA and genomic DNA was isolated from the retinas of three heterozygous rd/+ mouse strains. For each, quantitative reverse transcription-polymerase chain reaction (RT-PCR) was used to determine the ratio of wild-type and rd beta-phosphodiesterase pre-mRNA and mature mRNA. The gene copy ratio between wild-type and rd beta-PDE was also determined by quantitative PCR.


RESULTS
The pre-mRNA ratio of wild-type versus nonsense mutant was close to 1:1, whereas the corresponding mRNA ratio was greater than 3:1, even though the gene copy ratio was confirmed to be 1:1.


CONCLUSIONS
The equivalence of pre-mRNA ratio level for wild-type and nonsense mutant in the rd/+ retina indicates that both genes were transcribed at similar levels. Thus, neither the nonsense mutation at position 347 nor the intron 1 retroviral insertion also present in the rd gene seem to have affected gene transcription. In contrast, the strain-independent bias favoring wild-type mature mRNA in vivo suggests a specific degradation of mutant transcript during or after pre-mRNA splicing. This allele-specific degradation serves to decrease mutant transcript levels dramatically in all rd strains, and suggests that photoreceptor cells have the capacity to reduce the level of an mRNA containing a nonsense mutation.
Yan
W
W
Department of Molecular Genetics and Microbiology, University of Florida, Gainesville 32610, USA.
Lewin
A
A
Hauswirth
W
W
D015106:3',5'-Cyclic-GMP Phosphodiesterases / Q000235:genetics / Q000378:metabolism*; D000818:Animals; D054707:Cyclic Nucleotide Phosphodiesterases, Type 6; D004247:DNA / Q000032:analysis; D017931:DNA Primers / Q000737:chemistry; D005136:Eye Proteins / Q000235:genetics / Q000378:metabolism*; D005260:Female; D018628:Gene Dosage; D051379:Mice; D008817:Mice, Mutant Strains; D012313:RNA / Q000032:analysis; D012333:RNA, Messenger / Q000378:metabolism*; D012160:Retina / Q000201:enzymology*; D012162:Retinal Degeneration / Q000201:enzymology* / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction
D017931:DNA Primers; D005136:Eye Proteins; D012333:RNA, Messenger; D012313:RNA; D004247:DNA; D015106:3',5'-Cyclic-GMP Phosphodiesterases; D054707:Cyclic Nucleotide Phosphodiesterases, Type 6; C517438:Pde6b protein, mouse