Molecular analysis of X-ray-induced alcohol dehydrogenase (ADH) null mutations in Drosophila melanogaster.
We have attempted to analyze at the molecular level mutants previously determined as having intragenic lesions caused by X-ray mutagenesis. C.S. Aaron isolated 33 null mutations at the Adh locus and in collaboration with other investigators classified 23 as deletions. Of the eight mutants analyzed here, only two produced a detectable ADH protein using the two-dimensional electrophoresis technique. Restriction endonuclease and Southern blot analysis showed that three of the mutants were normal compared to the wild-type restriction pattern, with one of the three producing a mutant ADH protein. Among the five mutants that had altered restriction patterns, only one mutant produced a detectable mutant ADH protein. All the mutants produced a hybridizable mRNA when probed with the genomic clones, suggesting that the mutant phenotype was not due to transcriptional inhibition. Two probable explanations proposed for these observations are (1) mutations may be due to deletions of one or a few bases resulting in frameshifts to nonsense codons and premature termination of ADH peptide synthesis or (2) mutations may be a result of transitions to nonsense codons, again producing shortened ADH proteins. Those mutants producing a mutant polypeptide may have resulted from mutations to missense rather than nonsense codons. The five mutants showing an abnormal endonuclease Southern blot along with the 23 mutants previously shown to be deletions (28/33 or 85%) are associated with multiple DNA chain breaks. Although all of the DNA chain breaks are not necessarily associated with the mutant phenotype of the Adh locus, multiple DNA chain breaks are the most consistent characteristic of ionizing radiation damage to DNA.
Kelley
M R
MR
Mims
I P
IP
Farnet
C M
CM
Dicharry
S A
SA
Lee
W R
WR
D000426:Alcohol Dehydrogenase; D000429:Alcohol Oxidoreductases / Q000235:genetics*; D000818:Animals; D004262:DNA Restriction Enzymes; D004331:Drosophila melanogaster / Q000201:enzymology / Q000235:genetics*; D005796:Genes; D007526:Isoelectric Point; D008970:Molecular Weight; D009154:Mutation / Q000528:radiation effects; D012333:RNA, Messenger / Q000235:genetics; D014965:X-Rays
D012333:RNA, Messenger; D000429:Alcohol Oxidoreductases; D000426:Alcohol Dehydrogenase; D004262:DNA Restriction Enzymes