A strain of Saccharomyces cerevisiae has been constructed which is deficient in the four alcohol dehydrogenase (ADH) isozymes known at present. This strain (adho), being irreversibly mutated in the genes ADHI, ADH3, and ADH4 and carrying a point mutation in the gene ADH2 coding for the glucose-repressible isozyme ADHII, still produces up to one third of the theoretical maximum yield of ethanol in a homofermentative conversion of glucose to ethanol. Analysis of the glucose metabolism of adho cells shows that the lack of all known ADH isozymes results in the formation of glycerol as a major fermentation product, accompanied by a significant production of acetaldehyde and acetate. Treatment of glucose-growing adho cells with the respiratory-chain inhibitor antimycin A leads to an immediate cessation of ethanol production, demonstrating that ethanol production in adho cells is dependent on mitochondrial electron -transport. Reduction of acetaldehyde to ethanol in isolated mitochondria could also be demonstrated. This reduction is apparently linked to the oxidation of acetaldehyde to acetate. Preliminary data suggest that this novel type of ethanol formation in S. cerevisiae is associated with the inner mitochondrial membrane.
The mouse seminal vesicle shape (svs) mutation is a spontaneous recessive mutation that causes branching morphogenesis defects in the prostate gland and seminal vesicles. Unlike many other mutations that reduce prostatic and/or seminal vesicle branching, the svs mutation dramatically reduces branching without reducing organ growth. Using a positional cloning approach, we identified the svs mutant lesion as a 491 bp insertion in the tenth intron of Fgfr2 that results in changes in the pattern of Fgfr2 alternative splicing. An engineered null allele of Fgfr2 failed to complement the svs mutation proving that a partial loss of FGFR2(IIIb) isoforms causes svs phenotypes. Thus, the svs mutation represents a new type of adult viable Fgfr2 allele that can be used to elucidate receptor function during normal development and in the adult. In the developing seminal vesicles, sustained activation of ERK1/2 was associated with branching morphogenesis and this was absent in svs mutant seminal vesicles. This defect appears to be the immediate downstream effect of partial loss of FGFR2(IIIb) because activation of FGFR2(IIIb) by FGF10 rapidly induced ERK1/2 activation, and inhibition of ERK1/2 activation blocked seminal vesicle branching morphogenesis. Partial loss of FGFR2(IIIb) was also associated with down-regulation of several branching morphogenesis regulators including Shh, Ptch1, Gli1, Gli2, Bmp4, and Bmp7. Together with previous studies, these data suggest that peak levels of FGFR2(IIIb) signaling are required to induce branching and sustain ERK1/2 activation, whereas reduced levels support ductal outgrowth in the prostate gland and seminal vesicles.
The reduction of acetaldehyde to ethanol plays a key role in sugar metabolism of baker's yeast, Saccharomyces cerevisiae, by allowing regeneration of NAD+ from glycolytic NADH+H+. The in vivo significance of the reaction catalyzed by alcohol dehydrogenase (ADH) has been verified by mutants (adhl) deficient in ADH I, one of the four known ADH isozymes. Such mutant cells grow slowly on glucose, and growth is entirely dependent on mitochondrial functions (23). Despite this requirement for a functional respiratory chain in adhi mutants, glucose is largely converted to the major fermentation products, glycerol and ethanol, and to minor products such as acetaldehyde and acetate. We showed recently that the residual ethanol production in adhl mutant cells (approximately 30% of wild-type level) cannot be attributed to any of the known ADH isozymes (ADH II through ADH IV) by using mutant cells carrying null alleles at the four ADH loci (9). The existence of another cytoplasmic NAD+-dependent ADH isozyme was also ruled out since adh°cells cannot grow in a medium containing ethanol as the sole source of carbon. Rather, acetaldehyde reduction in adh/ cells occurs inside mitochondria. In fact, mitochondrial preparations from glucose-grown cells can quantitatively convert acetaldehyde to ethanol and acetate. This dismutation of acetaldehyde requires a functional respiratory chain, which is consistent with the finding that ethanol formation in adh/ cells is blocked by antimycin A.
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