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Summary Release of cytoplasmic proteins into the supernatant occurs both in bacteria and eukaryotes. Since the underlying mechanism remains unclear, the excretion of cytoplasmic proteins (ECP) has been referred to as ‘non-classical protein secretion’. We show that none of the known specific protein transport systems of Gram-positive bacteria are involved in ECP. However, the expression of the cationic and amphipathic α-type phenol-soluble modulins (PSMs), particularly of PSMα2, significantly increased ECP; while PSMβ peptides or δ-toxin have no effect on ECP. Since psm expression is strictly controlled by the accessory gene regulator (agr), ECP was also reduced in agr-negative mutants. PSMα peptides damage the cytoplasmic membrane as indicated by the release of not only CPs, but also lipids, nucleic acids and ATP. Thus, our results show that in Staphylococcus aureus, PSMα peptides non-specifically boost the translocation of CPs by their membrane damaging activity.

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The mechanisms behind carbon dioxide (CO 2 ) dependency in non-autotrophic bacterial isolates are unclear. Here we show that the Staphylococcus aureus mpsAB operon, known to play a role in membrane potential generation, is crucial for growth at atmospheric CO 2 levels. The genes mpsAB can complement an Escherichia coli carbonic anhydrase (CA) mutant, and CA from E. coli can complement the S. aureus delta- mpsABC mutant. In comparison with the wild type, S. aureus mps mutants produce less hemolytic toxin and are less virulent in animal models of infection. Homologs of mpsA and mpsB are widespread among bacteria and are often found adjacent to each other on the genome. We propose that MpsAB represents a dissolved inorganic carbon transporter, or bicarbonate concentrating system, possibly acting as a sodium bicarbonate cotransporter.

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dUnderstanding the mechanisms of how bacteria become tolerant toward antibiotics during clinical therapy is a very important object. In a previous study, we showed that increased daptomycin (DAP) tolerance of Staphylococcus aureus was due to a point mutation in pitA (inorganic phosphate transporter) that led to intracellular accumulation of both inorganic phosphate (P i ) and polyphosphate (polyP). DAP tolerance in the pitA6 mutant differs from classical resistance mechanisms since there is no increase in the MIC. In this follow-up study, we demonstrate that DAP tolerance in the pitA6 mutant is not triggered by the accumulation of polyP. Transcriptome analysis revealed that 234 genes were at least 2.0-fold differentially expressed in the mutant. Particularly, genes involved in protein biosynthesis, carbohydrate and lipid metabolism, and replication and maintenance of DNA were downregulated. However, the most important change was the upregulation of the dlt operon, which is induced by the accumulation of intracellular P i . The GraXRS system, known as an activator of the dlt operon (D-alanylation of teichoic acids) and of the mprF gene (multiple peptide resistance factor), is not involved in DAP tolerance of the pitA6 mutant. In conclusion, DAP tolerance of the pitA6 mutant is due to an upregulation of the dlt operon, triggered directly or indirectly by the accumulation of P i .

Alterations in the expression of apoptosis-related proteins, like the inhibitor of apoptosis (IAP) protein family, display a pivotal pathway by which cancer cells acquire resistance to therapeutic treatment. Among this family, survivin, the smallest and structural unique member, deserves growing attention due to its universal over-expression in human tumors, and its prominent role in disparate networks of cellular division, intracellular signaling and apoptosis. Several preclinical studies have demonstrated that targeting survivin expression by the use of small interfering RNAs, dominant negative mutants, antisense-oligonucleotides and small molecule repressors sensitized tumor cells towards chemotherapy and irradiation and reduced tumor growth potential. Due to these properties, survivin has been proposed as a molecular target for anticancer therapies. Recent studies further revealed that radio-sensitization achieved by survivin inhibition seems to be multifaceted and involves caspase-dependent and caspase-independent mechanisms. In general, an enhanced rate of apoptosis, and pronounced cell cycle arrest have been observed. More recently, a hampered DNA-damage response has been noted, indicating a distinct role of the protein in radiation-induced double strand break repair. These properties were linked to a nuclear import and physical interrelationship with members of the DNA-DSB repair machinery such as phospho-histone H2AX and DNA dependent Protein Kinase (DNA-PKcs). The applicability of survivin-driven strategies in clinical practice is currently under investigation as the first survivin inhibitors successfully entered phase I/II trials. Although these trials do not include radiation therapy at present, survivin inhibitors may represent a novel type of molecular antagonists to improve the effectiveness of radiation therapy or chemoradio-therapy.