Publicação
Constitutive expression and functionality of the gene JEN1 of Saccharomyces cerevisiae in the presence of glucose
| Resumo: | The product of the gene JEN1 is a plasma membrane permease for lactate, acetate, propionate and pyruvate in S. cerevisiae that is absent in glucose grown cells (Casal et al., 1999. J. Bacteriol. 181: 2620-2623). The aim of the present work was to study the expression and functionality of JEN1 in the presence of glucose. The entire ORF was cloned by PCR into the centromeric plasmid p416GPD, under the control of the constitutive promotor derived from the gene encoding glyceraldehyde-3-phosphate dehydrogenase. The recipient strain used in the present work was W303-1A jen1: strain L23 contains JEN1 under the control of the above mentioned promotor, and strain L19 contains the empty plasmid. In cells of the strain L23, grown in YNB with glucose (2%, w/v) pH 4.0, the expression of JEN1 was confirmed by RT-PCR. Activity for the monocarboxylate permease, measured with 14[C]-lactic acid at pH 5.0 was also present, therefore indicating that the carrier is operational. In cells of strain L19, neither JEN1 mRNA nor transport activity was found under the above described conditions. Both strains had identical growth parameters in all the conditions tested, and showed similar substrate consumption patterns. In the present work it was verified that, in the presence of glucose, the main catabolic repressor of the monocarboxylate permease, the constitutive expression of JEN1 can restore the transport activity. |
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| Autores principais: | Schuller, Dorit Elisabeth |
| Outros Autores: | Casal, Margarida; Leão, Cecília |
| Ano: | 2000 |
| País: | Portugal |
| Tipo de documento: | outro |
| Tipo de acesso: | acesso aberto |
| Instituição associada: | Universidade do Minho |
| Idioma: | inglês |
| Origem: | RepositóriUM - Universidade do Minho |
| Resumo: | The product of the gene JEN1 is a plasma membrane permease for lactate, acetate, propionate and pyruvate in S. cerevisiae that is absent in glucose grown cells (Casal et al., 1999. J. Bacteriol. 181: 2620-2623). The aim of the present work was to study the expression and functionality of JEN1 in the presence of glucose. The entire ORF was cloned by PCR into the centromeric plasmid p416GPD, under the control of the constitutive promotor derived from the gene encoding glyceraldehyde-3-phosphate dehydrogenase. The recipient strain used in the present work was W303-1A jen1: strain L23 contains JEN1 under the control of the above mentioned promotor, and strain L19 contains the empty plasmid. In cells of the strain L23, grown in YNB with glucose (2%, w/v) pH 4.0, the expression of JEN1 was confirmed by RT-PCR. Activity for the monocarboxylate permease, measured with 14[C]-lactic acid at pH 5.0 was also present, therefore indicating that the carrier is operational. In cells of strain L19, neither JEN1 mRNA nor transport activity was found under the above described conditions. Both strains had identical growth parameters in all the conditions tested, and showed similar substrate consumption patterns. In the present work it was verified that, in the presence of glucose, the main catabolic repressor of the monocarboxylate permease, the constitutive expression of JEN1 can restore the transport activity. |
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