The molecular mechanisms that enable yeast cells to detect and transmit cold signals and their physiological significance in the adaptive response to low temperatures are unknown. Here, we have demonstrated that the MAPK Hog1p is specifically activated in response to cold. Phosphorylation of Hog1p was dependent on Pbs2p, the MAPK kinase (MAPKK) of the high osmolarity glycerol (HOG) pathway, and Ssk1p, the response regulator of the two-component system Sln1p-Ypd1p. However, Sho1p was not required. Interestingly, phosphorylation of Hog1p was stimulated at 30°C in cells exposed to the membrane rigidifier agent dimethyl sulfoxide. Moreover, Hog1p activation occurred specifically through the Sln1 branch. This suggests that Sln1p monitors changes in membrane fluidity caused by cold. Quite remarkably, activation of Hog1p at low temperatures affected the transcriptional response to cold shock. Indeed, the absence of Hog1p impaired the cold-instigated expression of genes for trehalose-and glycerol-synthesizing enzymes and small chaperones. Moreover, a downward transfer to 12 or 4°C stimulated the overproduction of glycerol in a Hog1p-dependent manner. However, hog1⌬ mutant cells showed no growth defects at 12°C as compared with the wild type. On the contrary, deletion of HOG1 or GPD1 decreased tolerance to freezing of wildtype cells preincubated at a low temperature, whereas no differences could be detected in cells shifted directly from 30 to ؊20°C. Thus, exposure to low temperatures triggered a Hog1p-dependent accumulation of glycerol, which is essential for freeze protection.Variations in the surrounding temperature are probably the most common stress for all living organisms. In particular, a downshift in temperature leads to a reduction in membrane fluidity, impaired protein biosynthesis, and stabilization of secondary structures of DNA and RNA (1). Consequently, the adaptive response to cold shock in most organisms, studied so far, includes a change in the lipid composition of membranes and the remodeling of the transcriptional and translational machinery. These changes are mainly triggered by a drastic variation in the gene expression program, which leads to both survival and adaptation to low temperatures.Unlike other stress conditions, the biochemical mechanisms by which eukaryotic cells sense changes and respond to a downshift in temperature are poorly understood. Using the cyanobacteria Synechocystis as an experimental model, Murata and Los (2) suggested that a phase transition of the plasma membrane upon cold shock would trigger a conformational change of a putative cold sensor, this being the primary event in the transduction of the temperature signal. According to this hypothesis, a membrane-bound histidine kinase, Hik33, has been identified as a cold sensor in this organism, which is able to detect changes in the physical state of the membrane and regulate the expression of most cold-induced genes (3, 4). Other putative cold sensors, such as DesK in Bacillus subtilis (5) and transient receptor potential (T...

The response of yeast cells to sudden temperature downshifts has received little attention compared with other stress conditions. Like other organisms, both prokaryotes and eukaryotes, in Saccharomyces cerevisiae a decrease in temperature induces the expression of many genes involved in transcription and translation, some of which display a cold-sensitivity phenotype. However, little is known about the role played by many cold-responsive genes, the sensing and regulatory mechanisms that control this response or the biochemical adaptations at or near 0 degrees C. This review focuses on the physiological significance of cold-shock responses, emphasizing the molecular mechanisms that generate and transmit cold signals. There is now enough experimental evidence to conclude that exposure to low temperature protects yeast cells against freeze injury through the cold-induced accumulation of trehalose, glycerol and heat-shock proteins. Recent results also show that changes in membrane fluidity are the primary signal triggering the cold-shock response. Notably, this signal is transduced and regulated through classical stress pathways and transcriptional factors, the high-osmolarity glycerol mitogen-activated protein kinase pathway and Msn2/4p. Alternative cold-stress generators and transducers will also be presented and discussed.

The HXK2 gene is required for a variety of regulatory effects leading to an adaptation for fermentative metabolism in Saccharomyces cerevisiae. However, the molecular basis of the specific role of Hxk2p in these effects is still unclear. One important feature in order to understand the physiological function of hexokinase PII is that it is a phosphoprotein, since protein phosphorylation is essential in most metabolic signal transductions in eukaryotic cells. Here we show that Hxk2p exists in vivo in a dimeric-monomeric equilibrium which is affected by phosphorylation. Only the monomeric form appears phosphorylated, whereas the dimer does not. The reversible phosphorylation of Hxk2p is carbon source dependent, being more extensive on poor carbon sources such as galactose, raffinose, and ethanol. In vivo dephosphorylation of Hxk2p is promoted after addition of glucose. This effect is absent in glucose repression mutants cat80/grr1, hex2/reg1, and cid1/glc7. Treatment of a glucose crude extract from cid1-226 (glc7-T152K) mutant cells with -phosphatase drastically reduces the presence of phosphoprotein, suggesting that CID1/GLC7 phosphatase together with its regulatory HEX2/REG1 subunit are involved in the dephosphorylation of the Hxk2p monomer. An HXK2 mutation encoding a serine-to-alanine change at position 15 [HXK2 (S15A)] was to clarify the in vivo function of the phosphorylation of hexokinase PII. In this mutant, where the Hxk2 protein is unable to undergo phosphorylation, the cells could not provide glucose repression of invertase. Glucose induction of HXT gene expression is also affected in cells expressing the mutated enzyme. Although we cannot rule out a defect in the metabolic state of the cell as the origin of these phenomena, our results suggest that the phosphorylation of hexokinase is essential in vivo for glucose signal transduction.