In this paper, we present data on sugar-induced cell death (SICD) in the yeast Saccharomyces cerevisiae in the exponential phase of growth. We suggest that the nature of SICD in exponentially grown yeast is primary necrosis, in contrast to cells in the stationary growth phase, which exhibit apoptotic SICD. The following findings confirm this conclusion: (i) the process rate; (ii) the impairments of plasma membrane integrity; (iii) the drastic morphological changes in the intracellular content; (iv) the absence of chromatin condensation; (v) the absence of externalization of phosphotidylserine (PS) on the outer leaflet of plasma membrane and (vi) the insensitivity of the SICD process to cycloheximide (CHX). Research shows that SICD occurs in a subpopulation of cells in the S-phase.

Inorganic polyphosphate (polyP) is crucial for adaptive reactions and stress response in microorganisms. A convenient model to study the role of polyP in yeast is the Saccharomyces cerevisiae strain CRN/PPN1 that overexpresses polyphosphatase Ppn1 with stably decreased polyphosphate level. In this study, we combined the whole-transcriptome sequencing, fluorescence microscopy, and polyP quantification to characterize the CRN/PPN1 response to manganese and oxidative stresses. CRN/PPN1 exhibits enhanced resistance to manganese and peroxide due to its pre-adaptive state observed in normal conditions. The pre-adaptive state is characterized by up-regulated genes involved in response to an external stimulus, plasma membrane organization, and oxidation/reduction. The transcriptome-wide data allowed the identification of particular genes crucial for overcoming the manganese excess. The key gene responsible for manganese resistance is PHO84 encoding a low-affinity manganese transporter: Strong PHO84 down-regulation in CRN/PPN1 increases manganese resistance by reduced manganese uptake. On the contrary, PHM7, the top up-regulated gene in CRN/PPN1, is also strongly up-regulated in the manganese-adapted parent strain. Phm7 is an unannotated protein, but manganese adaptation is significantly impaired in Δphm7, thus suggesting its essential function in manganese or phosphate transport.

Enzyme I (EI) is the first protein in the phosphoryl transfer sequence from phosphoenolpyruvate (PEP) to sugar in carbohydrate uptake via the bacterial PEP:glycose phosphotransferase system. The EI monomer͞ dimer transition may regulate the phosphotransferase system because only the EI dimer is autophosphorylated by PEP. We previously showed that the EI monomer comprises two major domains: (i) a compact, protease-resistant N-terminal domain (EI-N), containing the active site His, and (ii) a f lexible, protease-sensitive C-terminal domain (EI-C), which is required for EI dimerization. EI-N interacts with the second protein, HPr, and phospho-HPr, but EI-N neither dimerizes nor is phosphorylated by PEP. We report here the molecular cloning and some properties of EI-C. EI-C is rapidly proteolyzed in vivo. Therefore, two different overexpression vectors encoding fusion proteins were constructed. Fusion Xa contains MalE (the maltose-binding protein), the four-amino acid sequence required by protease factor Xa, followed by EI-C. The phosphoenolpyruvate (PEP):glycose phosphotransferase system (PTS) has an essential role in several physiological processes in the bacterial cell. These include sugar transport, chemotaxis to its sugar substrates, regulation of expression of certain operons, etc. (for reviews, see refs. 1-3). A given function is mediated by a particular ensemble of PTS proteins. For example, uptake͞phosphorylation of different sugars requires from three to six PTS proteins. However, the common, pivotal component of each system is enzyme I (EI), the first protein in the phosphoryl transfer sequence.Some properties of the Escherichia coli and Salmonella typhimurium EI proteins (which are virtually identical) have been reviewed (4) and are briefly summarized as follows. (i) EI undergoes a highly temperature-sensitive monomer͞dimer transition; the E. coli monomer molecular mass is 63.5 kDa. The dimer accepts the phosphoryl group from PEP, but the monomer does not. The rates of association͞dissociation are surprisingly slow, much slower than the catalytic rate of the enzyme (5). (ii) The monomer contains two domains, a thermally stable, protease-resistant N-terminal domain, EI-N, and a flexible, highly protease-sensitive C-terminal domain, EI-C (6). (iii) EI-N contains the active site His residue and was originally isolated after protease digestion of intact EI (6). It was subsequently cloned from the intact gene ptsI (7,8). Both the solution and crystal structures of cloned EI-N have been established (9, 10). (iv) The C-terminal domain contains all four Cys residues and both Trp residues of intact EI. Because EI-N does not dimerize, it was originally suggested that the C-terminal domain is responsible for dimer formation (6). The flexibility of EI-C has been well documented in thermal unfolding experiments (6) and by the reactivity of Cys-SH residues, which vary in the presence of EI ligands (11,12). It has been suggested that the EI-C domain in intact EI confers species specificity for HPr in the N-term...

Extracellular pH has a signi cant impact on the physiology of the yeast cell, but its role in cell death has not been thoroughly investigated. We studied the effect of extracellular pH on the development of primary necrosis in Saccharomyces cerevisiae yeast under two general conditions leading to cell death. The rst is sugar induced cell death (SICD), and the second is death caused by several speci c gene deletions, which have been recently identi ed in a systematic screen. It was shown that in both cases, primary necrosis is suppressed at neutral pH. SICD was also inhibited by the protonophore dinitrophenol (DNP) and 150 mM extracellular K+, with the latter condition also bene ting survival of cell dying due to gene mutations. Thus, we show that neutral pH can suppress different types of primary necrosis. We suggest that changes to the cellular membrane potential can play a central role in yeast cell death.