The vacuolar H + -ATPase (V-ATPase) has long been appreciated to function as an electrogenic H + pump. By altering the pH of intracellular compartments, the V-ATPase dictates enzyme activity, governs the dissociation of ligands from receptors and promotes the coupled transport of substrates across membranes, a role often aided by the generation of a transmembrane electrical potential. In tissues where the V-ATPase is expressed at the plasma membrane, it can serve to acidify the extracellular microenvironment. More recently, however, the V-ATPase has been implicated in a bewildering variety of additional roles that seem independent of its ability to translocate H + . These non-canonical functions, which include fusogenicity, cytoskeletal tethering and metabolic sensing, are described in this Cell Science at a Glance article and accompanying poster, together with a brief overview of the conventional functions of the V-ATPase.

SummaryThe phage-shock-protein (Psp) stress-response system is conserved in many bacteria and has been linked to important phenotypes in Escherichia coli , Salmonella enterica and also Yersinia enterocolitica , where it is essential for virulence. It is activated by specific extracytoplasmic stress events such as the mislocalization of secretin proteins. From studies of the Psp system in E. coli , the cytoplasmic membrane proteins PspB and PspC have only been proposed to act as positive regulators of psp gene expression. However, in this study we show that PspB and PspC of Y. enterocolitica are dual function proteins, acting both as regulators and effectors of the Psp system. Consistent with the current model, they positively control psp gene expression in response to diverse inducing cues. PspB and PspC must work together to achieve this regulatory function, and bacterial twohybrid (BACTH) analysis demonstrated a specific interaction between them, which was confirmed by in vivo cross-linking. We also show that PspB and PspC play a second role in supporting growth when a secretin protein is overexpressed. This function is independent from their role as regulators of psp gene expression. Furthermore, whereas PspB and PspC must work together for their regulatory function, they can apparently act independently to support growth during secretin production. This study expands the current understanding of the roles played by PspB and PspC, and demonstrates that they cannot be considered only as positive regulators of psp gene expression in Y. enterocolitica .

Known inducers of the phage shock protein (Psp) system suggest that it is an extracytoplasmic stress response, as are the well-studied RpoE and Cpx systems. However, a random approach to identify conditions and proteins that induce the Psp system has not been attempted. It is also unknown whether the proteins or mutations that induce Psp are specific or if they also activate the RpoE and Cpx systems. This study addressed these issues for the Yersinia enterocolitica Psp system. Random transposon mutagenesis identified null mutations and overexpression mutations that increase ⌽(pspA-lacZ) operon fusion expression. The results suggest that Psp may respond exclusively to extracytoplasmic stress. Null mutations affected glucosamine-6-phosphate synthetase (glmS), which plays a role in cell envelope biosynthesis, and the F 0 F 1 ATPase (atp operon). The screen also revealed that in addition to several secretins, the overexpression of three novel putative inner membrane proteins ( Misfolding and/or mislocalization of envelope proteins induce extracytoplasmic stress responses in bacteria. The RpoE and Cpx systems of Escherichia coli and its relatives are wellstudied examples (reviewed in references 41 and 42). These systems control many genes, with some overlap between their regulons, which encode proteases, envelope protein folding factors, and several proteins of unknown function (10,41,43). Mounting a response to extracytoplasmic stress is extremely important. rpoE is an essential gene in E. coli (14) and Yersinia enterocolitica (21), although this is apparently not the case in the closely related Salmonella genus (23).The RpoE response is important for virulence in Salmonella enterica serovar Typhimurium (23,45) and Vibrio cholerae (30). In addition, the RpoE and Cpx responses are induced by the overproduction of P pilus subunits from uropathogenic E. coli (25), and the Cpx system affects assembly and expression of the P pilus (24). The Cpx system is also important for the attachment of E. coli to surfaces (37), which is a critical step during biofilm formation.The phage shock protein (Psp) system may be another example of an extracytoplasmic stress response. pspA operon expression, studied most extensively in E. coli K-12, is induced by the mislocalization of secretin proteins and by environmental conditions that induce the RpoE response (reviewed in reference 35).

Exposure of Cryptococcus neoformans cells to gamma radiation results in a gradual release of capsular polysaccharide, in a dose-dependent manner. This method allows the systematic exploration of different capsular regions. Using this methodology, capsule density was determined to change according to the radial distribution of glucuronoxylomannan and total polysaccharide, becoming denser at the inner regions of the capsule. Scanning electron microscopy of cells following gamma radiation treatment confirmed this finding. The zeta potential of the capsule also increased as the capsule size decreased. However, neither charge nor density differences were correlated with any change in sugar composition (xylose, mannose, and glucuronic acid) in the different capsular regions, since the proportions of these sugars remained constant throughout the capsule. Analysis of the capsular antigenic properties by monoclonal antibody binding and Scatchard analysis revealed fluctuations in the binding affinity within the capsule but not in the number of antibody binding sites, suggesting that the spatial organization of high-and low-affinity epitopes within the capsule changed according to radial position. Finally, evidence is presented that the structure of the capsule changes with capsule age, since the capsule of older cells became more resistant to gamma radiation-induced ablation. In summary, the capsule of C. neoformans is heterogeneous in its spatial distribution and changes with age. Furthermore, our results suggest several mechanisms by which the capsule may protect the fungal cell against exogenous environmental factors.Capsules are a common feature among microorganisms, especially pathogenic bacteria such as Bacillus anthracis, Streptococcus pneumoniae, and Neisseria meningitidis. Microbial capsules can confer particular characteristics, such as protection against stress conditions (64), and are prominent virulence factors. In contrast to the situation in bacteria, extracellular capsules are rare in fungi. The only encapsulated pathogenic fungus is the basidiomycetous yeast Cryptococcus neoformans. This fungus is commonly found in the environment, inhabiting various niches such as pigeon droppings, trees, and water (reviewed in reference 8). The pathogenesis of C. neoformans has been well studied. The yeast is commonly acquired by the host via inhalation. The infection is asymptomatic in immunocompetent hosts. However, in cases of immune suppression, pulmonary infection can be followed by extrapulmonary dissemination of the yeast into other organs, such as spleen, liver, and brain. Untreated cryptococcal meningitis is invariably fatal.The polysaccharide capsule of C. neoformans is considered the main virulence factor of this pathogen (37). Acapsular C. neoformans strains manifest greatly reduced virulence (10, 31), and mutants that produce a larger capsule are hypervirulent (19). The capsule of this yeast is believed to function in protection from desiccation, radiation, and predation by phagocytic organisms (reviewed in...