High Osmolarity Pathway in Budding Yeast
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High Osmolarity Pathway in Budding Yeast
The internal osmolarity of a growing yeast cell is maintained higher than the external osmolarity. The resulting osmotic gradient across the plasma membrane brings in water for cell expansion and creates turgor. Yeast cells are quite resistant to various types of stress including hypertonic stress. This adaptability can be traced to stress-activated signaling pathways that sense the stress condition and activate expression of proteins that resist the toxic effects of the stress and promote survival and eventual cell growth under the new conditions. In Saccharomyces cerevisiae, changes in the osmolarity of the medium have been reported to affect different signaling pathways. The best-characterized signaling system by far involves the MAPK (Mitogen-Activated Protein Kinase) Hog1, a relative of the p38 and JNK (c-Jun N-terminal kinase) families of SAPKs (Stress-Activated Protein Kinases). In yeast, specific osmosensing devices seem to be responsible for detecting changes in osmolarity. High osmolarity in budding yeast activates at least two different sensors, which are connected to different signal transduction branches that converge at the level of the MAP Kinase Kinase Pbs2. One signaling branch is defined by the Sln1Ypd1Ssk1 multicomponent system, the other by Sho1, a plasma-membrane protein that contains an SH3 domain. These two sensors regulate two different MAPK pathways utilizing common kinase elements that will lead to the transcription of genes necessary for survival in hyperosmotic conditions such as those required for the synthesis of glycerol to increase the internal osmolarity (Ref.1).

Sln1 Pathway is considered as two-component osmosensor pathway, which is composed of three proteins, Sln1, Ypd1, and Ssk1, which functionally behave as two linked two-component systems. Sln1 corresponds to the first two-component system, since it contains both a histidine kinase domain and a receiver domain. The Ypd1-Ssk1 pair functions as the second two-component system. Ypd1 is phosphorylated on a histidine residue as a result of a transfer of the phosphate on the aspartic acid of Sln1. This phosphate is then transferred to Ssk1 on an aspartic acid. Sln1 and Ypd1 are predicted to act as negative regulators of the Hog pathway MAPK cascade. Ssk1 has a kinase activity, and it interacts with and regulates the Ssk2 and Ssk22 MAPKKKs and subsequent Pbs2 activation. Ssk1 is inactive when phosphorylated (i.e., in low-osmolarity conditions) and activated by high osmolarity (when the sensor Sln1 is inactive), which eventually leads to stimulation of Hog1 and transcription of genes necessary for survival in hyperosmotic conditions (Ref.2).

Pbs2 activation can also be achieved by a second, independent mechanism that involves the transmembrane protein Sho1, the MAPKKK Ste11, the Ste11-binding protein Ste50, the Ste20 PAK (p21-activated kinase) and the small GTPase CDC42 (Cell Division Cycle-42). Sho1 contains four transmembrane domains and a COOH-terminal cytoplasmic region with a SH3 (Src Homology-3) domain. In high-osmolarity conditions, Ste11 is activated in a Sho1-dependent manner. Ste11 then phosphorylates and activates Pbs2, which activates Hog1 in the module. Phosphorylation causes a rapid and marked concentration of Hog1 in the nucleus, while under normal conditions Hog1 appears to be evenly distributed between the cytosol and the nucleus. Activated Hog1 regulates the transcription of genes required for survival in hyperosmotic conditions (Ref.3).

In budding yeast, Hog1 MAPK plays a key role in global gene regulation. These osmostress-regulated genes are implicated mainly in carbohydrate metabolism, general stress protection, protein production and signal transduction. This global change in transcription could account, at least in part, for the metabolic adjustments required for osmostress adaptation. In yeast, five transcription factors are known to be controlled by the Hog1 MAPK. Hot1, Smp1, Msn2 and Msn4 activate, whereas Sko1 represses or activates, different subsets of osmotic-inducible and Hog1-regulated genes. Sko1 is an ATF/CREB factor whose repressive activity via the Ssn6Tup1 complex is inhibited by Hog1 in response to osmostress. Msn2 and Msn4 are generic stress factors controlled by PKA and Hog1 by an unknown mechanism. Hot1 physically interacts with Hog1, and its binding to DNA and subsequent transactivation activity are regulated by its phosphorylation by the kinase. Genetic evidence and gene expression data link additional transcription factors to Hog pathway function: Msn1, Sgd1, and Gcn4. In addition, the Sln1-Ypd1 sensor system controls a second response regulator, Skn7. This protein is apparently involved in multiple cellular processes and genetically interacts with both the Hog and the cell integrity pathways (Ref.4).

The Hog pathway is also negatively regulated by the action of the tyrosine-specific protein phosphatases Ptp2 and Ptp3, which dephosphorylate the MAPK Hog1. Three phosphoserine/threonine phosphatases (Ptc1, Ptc2 and Ptc3) also negatively regulates Hog pathway, whose substrate is unknown. Removal or inactivation of these negative regulators causes reduced cell growth through hyperactivation of the Hog pathway. The Hog pathway plays a role in mediating the hyperosmotic stress-induced expression of stress response genes, recovery of cell morphogenesis, and repression of the pheromone response pathway. Not only is the Hog pathway required for various responses to hyperosmotic stress, but also hyperosmotic stress activates the pathway, measured as an increase in tyrosine phosphorylation of the MAPK Hog1. The Hog pathway is activated by increasing the concentration of different solutes (e.g., NaCl, KCl, sorbitol, or glucose), showing that the activating stimulus is truly related to the osmotic change rather than an increase in concentration of a specific solute. How the MAPK cascade is activated by hyperosmotic stress stands out as a fascinating problem for which there are now several important molecular clues (Ref.5).