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We have isolated the KlNOT4 gene of the yeast Kluyveromyces lactis, which encodes a component of the evolutionarily conserved CCR4-NOT complex. We show that inactivation of the gene leads to pleiotropic defects that were differentially suppressed by the NOT4 gene of S. cerevisiae, indicating that these genes have overlapping, but not identical, functions. K. lactis strains lacking Not4p are defective in fermentation and show reduced transcription of glucose transporter and glycolytic genes, which are phenotypes that are not found in the corresponding mutant of S. cerevisiae. We also show that Not4 proteins control the respiratory pathway in both yeasts, although with some differences. They activate transcription of KlACS2 and KlCYC1, but repress KlICL1, ScICL1, ScACS1, and ScCYC1. Altogether, our results indicate that Not4p is a pivotal factor involved in the regulation of carbon metabolism in yeast.

The Inactivation of KlNOT4, a Kluyveromyces lactis Gene Encoding a Component of the CCR4-NOT Complex, Reveals New Regulatory Functions