Characterization of the functional gene encoding mouse class III alcohol dehydrogenase (glutathione-dependent formaldehyde dehydrogenase) and an unexpressed processed pseudogene with an intact open reading frame.
Multiple forms of vertebrate alcohol dehydrogenase (ADH) have been identified, but only one form, class III ADH, has been conserved in all organisms studied. Class III ADH functions in vitro as a glutathione-dependent formaldehyde dehydrogenase, which suggests that this was the original function that drove the evolution of ADH. Genetic analysis of class III ADH in yeast supports this view, but such studies are lacking in higher eukaryotes. The mouse ADH family has been previously analyzed and it contains three forms of ADH including the class III enzyme. We have initiated a molecular genetic analysis of the mouse class III ADH gene (Adh-2) by screening a genomic library with a full-length cDNA. Two overlapping clones contained the complete Adh-2 gene composed of nine exons in a 12-kb region, with the placement of introns matching that observed in other mammalian ADH genes. In this screening, we also isolated a clone (psi Adh-2) that lacks introns and which resembles a processed pseudogene. psi Adh-2 contained 25 point mutations relative to the previously analyzed Adh-2 cDNA, but still retained an intact open reading frame. Northern blot analysis using gene-specific probes provided evidence that psi Adh-2 does not produce a mRNA in either liver or kidney, whereas Adh-2 does. The functionality of the two genes was also compared by fusion of their 5'-flanking regions to a lacZ reporter gene. Reporter gene expression following transfection into mouse F9 embryonal carcinoma cells indicated that only Adh-2 possesses promoter activity.
Foglio
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La Jolla Cancer Foundation Research Foundation, CA 92037, USA.
Duester
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D000445:Aldehyde Oxidoreductases / Q000235:genetics*; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D003001:Cloning, Molecular; D015342:DNA Probes / Q000235:genetics; D018076:DNA, Complementary / Q000235:genetics; D019143:Evolution, Molecular; D005796:Genes; D017930:Genes, Reporter; D015698:Genomic Library; D006801:Humans; D007763:Lac Operon; D051379:Mice; D008969:Molecular Sequence Data; D016366:Open Reading Frames; D011401:Promoter Regions, Genetic; D011544:Pseudogenes; D012333:RNA, Messenger / Q000235:genetics; D015183:Restriction Mapping; D012689:Sequence Homology, Nucleic Acid; D013045:Species Specificity; D014162:Transfection
D015342:DNA Probes; D018076:DNA, Complementary; D012333:RNA, Messenger; C020145:formaldehyde dehydrogenase (glutathione); D000445:Aldehyde Oxidoreductases