Ionizing radiation (IR), such as X-rays and gamma (γ)-rays, mediates various forms of cancer cell death such as apoptosis, necrosis, autophagy, mitotic catastrophe, and senescence. Among them, apoptosis and mitotic catastrophe are the main mechanisms of IR action. DNA damage and genomic instability contribute to IR-induced cancer cell death. Although IR therapy may be curative in a number of cancer types, the resistance of cancer cells to radiation remains a major therapeutic problem. In this review, we describe the morphological and molecular aspects of various IR-induced types of cell death. We also discuss cytogenetic variations representative of IR-induced DNA damage and genomic instability. Most importantly, we focus on several pathways and their associated marker proteins responsible for cancer resistance and its therapeutic implications in terms of cancer cell death of various types and characteristics. Finally, we propose radiation-sensitization strategies, such as the modification of fractionation, inflammation, and hypoxia and the combined treatment, that can counteract the resistance of tumors to IR.

In Salmonella enterica serovar Typhimurium, the membrane-localized CadC is a transcriptional activator of the cadBA operon, which contributes to the acid tolerance response. Unlike in Escherichia coli, in which transcription of cadC is constitutive, in S. enterica serovar Typhimurium cadC expression is induced by low pH and lysine. Inactivation of cadC suppresses the acid-sensitive phenotype of a cadA mutation, suggesting the existence of other CadC-dependent genes in addition to the cadBA operon. Using a proteomic approach, we identified 8 of the putative CadC-induced proteins and 15 of the putative CadC-repressed proteins. The former include porin proteins OmpC and OmpF. The latter include proteins involved in glycolysis, energy production, and stress tolerance. To better understand the altered levels of OmpC and OmpF, we compared expression of ompR in S. enterica serovar Typhimurium wild-type and cadC mutant strains and determined that CadC exerted a negative influence on ompR transcription. Taken together, our findings strongly suggest that CadC may be a global regulator involved in the OmpR regulatory system during acid adaptation.

The glutamate dehydrogenase RocG of Bacillus subtilis is a bifunctional protein with both enzymatic and regulatory functions. Here we show that the rocG null mutant is sensitive to ␤-lactams, including cefuroxime (CEF), and to fosfomycin but that resistant mutants arise due to gain-of-function mutations in gudB, which encodes an otherwise inactive glutamate dehydrogenase. In the presence of CEF, ⌬rocG ⌬gudB mutant cells exhibit growth arrest when they reach mid-exponential phase. Using microarray-based transcriptional profiling, we found that the W regulon was downregulated in the ⌬rocG ⌬gudB null mutant. A survey of W -controlled genes for effects on CEF resistance identified both the NfeD protein YuaF and the flotillin homologue YuaG (FloT). Notably, overexpression of yuaFG in the rocG null mutant prevents the growth arrest induced by CEF. The YuaG flotillin has been shown previously to localize to defined lipid microdomains, and we show here that the yuaFGI operon contributes to a W -dependent decrease in membrane fluidity. We conclude that glutamate dehydrogenase activity affects the expression of the W regulon, by pathways that are yet unclear, and thereby influences resistance to CEF and other antibiotics. In Bacillus subtilis, a model system for the Gram-positive bacteria (36), the synthesis of glutamate is catalyzed uniquely by the heterodimeric product of the gltAB operon. Glutamate acts as a central metabolite providing the link between carbon and nitrogen metabolism (11,40). The degradation of glutamate is catalyzed by the strictly catabolic glutamate dehydrogenase RocG (2). In addition to rocG, B. subtilis has a second glutamate dehydrogenase gene, gudB, whose product is cryptic due to an insertion of three amino acids close to the active site of this enzyme. However, null mutants of rocG rapidly accumulate spontaneous gain-offunction suppressor mutations in gudB that remove the repeat sequence encoding the three-amino-acid insertion, thereby resulting in the synthesis of active GudB (3,12).Recent studies have shown that RocG has a second activity as a regulatory protein. RocG, if glutamate is available, directly interacts with GltC, the transcription activator of the gltAB operon, thus inhibiting its activity (10, 15). However, whether it has additional functions remains largely unknown. In addition to RocG, several other bacterial enzymes are now known to regulate gene expression. Some act as transcription factors by direct binding to either DNA or RNA, and others modulate the activity of transcription factors either by covalent modification or by protein-protein interactions (9).Cefuroxime (CEF) belongs to the group of broad-spectrum ␤-lactam cephalosporin antibiotics, with antimicrobial activity against both Gram-positive and Gram-negative bacteria (31). The mode of action of CEF is conventional: by binding to specific penicillin-binding proteins (PBPs), it inhibits the third and final stage of bacterial cell wall synthesis. In Gram-negative bacteria such as Escherichia coli, CEF shows high affin...

Undecaprenyl pyrophosphate synthase (UppS) catalyzes the formation of the C 55 lipid carrier (UPP) that is essential for bacterial peptidoglycan biosynthesis. We selected here a vancomycin (VAN)-resistant derivative of Bacillus subtilis W168 that contains a single-point mutation in the ribosome-binding site of the uppS gene designated uppS1. Genetic reconstruction experiments demonstrate that the uppS1 allele is sufficient to confer low-level VAN resistance and causes reduced UppS translation. The decreased level of UppS renders B. subtilis slightly more susceptible to many late-acting cell wall antibiotics, including ␤-lactams, but significantly more resistant to fosfomycin and D-cycloserine, antibiotics that interfere with the very early steps of cell wall synthesis. We further show that the uppS1 allele leads to slightly elevated expression of the M regulon, possibly helping to compensate for the stress caused by a decrease in UPP levels. Notably, the uppS1 mutation increases resistance to VAN, fosfomycin, and D-cycloserine in wild-type cells, but this effect is greatly reduced or eliminated in a sigM mutant background. Our findings suggest that, although UppS is an attractive antibacterial target, incomplete inhibition of UppS function may lead to increased resistance to some cell wall-active antibiotics.

bIn bacteria, mutations affecting the major catalytic subunits of RNA polymerase (encoded by rpoB and rpoC) emerge in response to a variety of selective pressures. Here we isolated a Bacillus subtilis strain with high-level resistance to cefuroxime (CEF). Whole-genome resequencing revealed only one missense mutation affecting an invariant residue in close proximity to the C-terminal DNA-binding domain of RpoC (G1122D). Genetic reconstruction experiments demonstrate that this substitution is sufficient to confer CEF resistance. The G1122D mutation leads to elevated expression of stress-responsive regulons, including those of extracytoplasmic function (ECF) factors ( M , W , and X ) and the general stress factor ( B ). The increased CEF resistance of the rpoC G1122D strain is lost in the sigM rpoC G1122D double mutant, consistent with a major role for M in CEF resistance. However, a sigM mutant is very sensitive to CEF, and this sensitivity is still reduced by the G1122D mutation, suggesting that other regulatory effects are also important. Indeed, the ability of the G1122D mutation to increase CEF resistance is further reduced in a triple mutant strain lacking three ECF factors ( M , W , and X ), which are known from prior studies to control overlapping sets of genes. Collectively, our findings highlight the ability of mutations in RNA polymerase to confer antibiotic resistance by affecting the activity of alternative factors that control cell envelope stress-responsive regulons.