1887

Abstract

Glucose-transport-deficient mutants of pombe were obtained by treatment of wild-type cells (972h) with -methyl-s'-nitro--nitrosoguanidine, and by selection of resulting mutants on gluconate medium containing 0.05% 2-deoxy-D-glucose (2DG). One mutant, designated , was unable to grow on D-glucose and/or D-fructose as a carbon source (Glc/Fru), and was resistant to 2DG; hence, none of the three sugars was taken up by the mutant cells. The hexokinase activity in the wild-type and the mutant cells was equal. Genetic purification of by back-crossing with a leucine-auxotrophic mutant and the wild-type resulted in two strains: , with reduced 2DG resistance, and , which had lost 2DG-resistance. grew in D-glucose-containing media, albeit very slowly. No measurable sugar uptake was detectable in either of the two mutants within the 1 h test interval. Tetrad analyses proved a Mendelian segregation of growth on D-glucose and leucine auxotrophy. However, 2DG resistance did not co-segregate with the Glc/Fruphenotype, indicating that the transport deficiency and 2DG resistance characters are not encoded on the same genomic locus. Using a genomic bank of , two transformants, and , were found which had regained the wild-type growth and transport phenotype by complementation. Correspondingly, both D-glucose uptake and 2DG accumulation were restored in the transformed strains. Restriction analysis and Northern blots suggested that the G7 genomic fragment and the 4.1 kb I restriction fragment of the G12 genomic fragment both contain a complete structural symporter gene.

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1994-10-01
2021-10-27
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