The expression of the major glucose transporter gene, RAG1, is induced by glucose in Kluyveromyces lactis. This regulation involves several pathways, including one that is similar to Snf3/Rgt2-ScRgt1 in Saccharomyces cerevisiae. We have identified missing key components of the K. lactis glucose signaling pathway by comparison to the same pathway of S. cerevisiae. We characterized a new mutation, rag19, which impairs RAG1 regulation. The Rag19 protein is 43% identical to the F-box protein ScGrr1 of S. cerevisiae and is able to complement an Scgrr1 mutation. In the K. lactis genome, we identified a single gene, SMS1 (for similar to Mth1 and Std1), that encodes a protein showing an average of 50% identity with Mth1 and Std1, regulators of the ScRgt1 repressor. The suppression of the rag4 (glucose sensor), rag8 (casein kinase I), and rag19 mutations by the ⌬sms1 deletion, together with the restoration of RAG1 transcription in the double mutants, demonstrates that Sms1 is a negative regulator of RAG1 expression and is acting downstream of Rag4, Rag8, and Rag19 in the cascade. We report that Sms1 regulates KlRgt1 repressor activity by preventing its phosphorylation in the absence of glucose, and that SMS1 is regulated by glucose, both at the transcriptional and the posttranslational level. Two-hybrid interactions of Sms1 with the glucose sensor and KlRgt1 repressor suggest that Sms1 mediates the glucose signal from the plasma membrane to the nucleus. All of these data demonstrated that Sms1 was the K. lactis homolog of MTH1 and STD1 of S. cerevisiae. Interestingly, MTH1 and STD1 were unable to complement a ⌬sms1 mutation.

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Sensing of extracellular glucose is necessary for cells to adapt to glucose variation in their environment. In the respiratory yeast Kluyveromyces lactis, extracellular glucose controls the expression of major glucose permease gene RAG1 through a cascade similar to the Saccharomyces cerevisiae Snf3/Rgt2/Rgt1 glucose signaling pathway. This regulation depends also on intracellular glucose metabolism since we previously showed that glucose induction of the RAG1 gene is abolished in glycolytic mutants. Here we show that glycolysis regulates RAG1 expression through the K. lactis Rgt1 (KlRgt1) glucose signaling pathway by targeting the localization and probably the stability of Rag4, the single Snf3/Rgt2-type glucose sensor of K. lactis. Additionally, the control exerted by glycolysis on glucose signaling seems to be conserved in S. cerevisiae. This retrocontrol might prevent yeasts from unnecessary glucose transport and intracellular glucose accumulation. Sensing and adaption to environmental variations and stresses is fundamental for any cell to live and to grow properly. Among the environmental signals that cells have to consider, nutrients, and especially glucose, are of particular importance. Indeed, glucose is the principal carbon and energy source for most living organisms. Glucose signaling is a key pathway allowing cells to adapt their sugar transport system and metabolism to the quality and quantity of carbon source present in their environment.Glucose transport and the glucose signaling network have been widely studied in the fermentative yeast model Saccharomyces cerevisiae (1). The yeast Kluyveromyces lactis is an excellent alternate and complementary model organism to investigate glucose signaling (2). Indeed, the pathway is simpler, with only two glucose permeases and little if any gene redundancy. Moreover, unlike S. cerevisiae, the K. lactis living style is preferably respiratory, making it closer to superior eukaryotes. In K. lactis, the fermentative growth on medium containing 5% glucose plus antimycin A (respiration inhibitor) defines the Rag-positive (Rag ϩ ) phenotype and is informative about the functionality of glucose signaling, glucose transport, and glucose metabolism.In K. lactis, two glucose permeases are known: Rag1, with a low affinity for glucose, and Hgt1, having a high affinity for glucose (3,4). Only RAG1 expression is regulated by the extracellular glucose concentration (5). In the absence of extracellular glucose, the transcriptional repressor K. lactis Rgt1 (KlRgt1), bound to the corepressor Sms1, represses RAG1 expression (6, 7) (see Fig. 1A for a model). Extracellular glucose is sensed by the plasma membrane glucose sensor Rag4, a glucose permease homolog unable to transport glucose (8) and exhibiting a characteristic long cytoplasmic C-terminal tail thought to have a role in glucose signaling. Glucose binding to Rag4 probably leads to a conformational change allowing interaction with the casein kinase I (CKI) Rag8 (6, 9). When activated by glucose, Rag8 is supposed to phosphor...