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Henry Rosen

Publications and source records attributed to Henry Rosen.

9 recordsLinked to original sources
PubMed2006

Selective silencing of foreign DNA with low GC content by the H-NS protein in Salmonella.

Horizontal gene transfer plays a major role in microbial evolution. However, newly acquired sequences can decrease fitness unless integrated into preexisting regulatory networks. We found that the histone-like nucleoid structuring protein (H-NS) selectively silences horizontally acquired genes by targeting sequences with GC content lower than the resident genome. Mutations in hns are lethal in Salmonella unless accompanied by compensatory mutations in other regulatory loci. Thus, H-NS provides a previously unrecognized mechanism of bacterial defense against foreign DNA, enabling the acquisition of DNA from exogenous sources while avoiding detrimental consequences from unregulated expression of newly acquired genes. Characteristic GC/AT ratios of bacterial genomes may facilitate discrimination between a cell's own DNA and foreign DNA.

Bacterial Proteins↗
PubMed2006

Effect of inactivation of the global oxidative stress regulator oxyR on the colonization ability of Escherichia coli O1:K1:H7 in a mouse model of ascending urinary tract infection.

To survive within the host urinary tract, Escherichia coli strains that cause urinary tract infection (UTI) presumably must overcome powerful oxidant stresses, including the oxygen-dependent killing mechanisms of neutrophils. Accordingly, we assessed the global oxygen stress regulator OxyR of Escherichia coli as a possible virulence factor in UTI by determining the impact of oxyR inactivation on experimental urovirulence in CBA/J and C57BL (both wild-type and p47(phox-/-)) mice. The oxyR and oxyS genes of wild-type E. coli strain Ec1a (O1:K1:H7) were replaced with a kanamycin resistance cassette to produce an oxyRS mutant. During in vitro growth in broth or human urine, the oxyRS mutant exhibited the same log-phase growth rate (broth) and plateau density (broth and urine) as Ec1a, despite its prolonged lag phase (broth) or initial decrease in concentration (urine). The mutant, and oxyRS mutants of other wild-type ExPEC strains, exhibited significantly increased in vitro susceptibility to inhibition by H(2)O(2), which, like the altered growth kinetics observed with oxyRS inactivation, were reversed by restoration of oxyR on a multiple-copy-number plasmid. In CBA/J mice, Ec1a significantly outcompeted its oxyRS mutant (by >1 log(10)) in urine, bladder, and kidney cultures harvested 48 h after perurethral inoculation of mice, whereas an oxyR-complemented mutant exhibited equal or greater colonizing ability than that of the parent. Although C57BL mice were less susceptible to experimental UTI than CBA/J mice, wild-type and p47(phox-/-) C57BL mice were similarly susceptible, and the oxyR mutant of Ec1a was similarly attenuated in C57BL mice, regardless of the p47(phox) genotype, as in CBA/J mice. Within the E. coli Reference collection, 94% of strains were positive for oxyR. These findings fulfill the second and third of Koch's molecular postulates for oxyR as a candidate virulence-facilitating factor in E. coli and indicate that oxyR is a broadly prevalent potential target for future preventive interventions against UTI due to E. coli. They also suggest that neutrophil phagocyte oxidase is not critical for defense against E. coli UTI and that the major oxidative stresses against which OxyR protects E. coli within the host milieu are not phagocyte derived.

Animals↗
PubMed2004

Bacterial responses to neutrophil phagocytosis.

PURPOSE OF REVIEW: This review focuses on adaptive bacterial interactions with neutrophils, emphasizing information communicated within the past year about bacterial factors that respond to contact with or phagocytosis by PMN. RECENT FINDINGS: Since the discovery of type III and IV secretion, progress in the analysis of bacterial interactions with host phagocytes has been extensive but largely focused on the macrophage. The remarkable growth of information about bacterial subversion of macrophage metabolism has been summarized in several excellent reviews. The scope of progress on neutrophil-bacteria interactions is more limited and dominated by recent studies of the granulocyte pathogen, Anaplasma phagocytophilum, the agent of granulocytic ehrlichiosis. SUMMARY: For many pathogens, contact with or ingestion by phagocytes elicits a vigorous but varied microbial response. The response repertoire includes activation of type III and type IV secretion systems that inject effector molecules into the host cell. Effectors modify host cell signaling and metabolic pathways to favor survival of the microbe. Whereas microbial secretory structures are few in kind and relatively conserved, effector molecules are numerous and variable. Effectors may promote phagocytosis by nonphagocytic cells or suppress phagocytosis by macrophages and neutrophils. They may suppress assembly or misdirect localization of the phagocyte NADPH oxidase that is responsible for generating toxic oxidants, and they may suppress phagosome-lysosome fusion. Phagocytosed bacteria may also up-regulate the expression of defensive proteins that attenuate the effects of phagocyte-derived antimicrobial toxins. These pathogenic stratagems probably have their origins in the competition among single-celled organisms, eukaryotes versus prokaryotes, that arose early in evolution.

Anaplasma phagocytophilum↗
PubMed2004

Neutrophil microbicidal activity: screening bacterial mutants for survival after phagocytosis using quantitative PCR.

When a constant gene replacement sequence is introduced into bacteria to produce mutants and the flanking chromosomal sequences are known, it is possible to use a quantitative polymerase chain reaction method (QPCR) to compare the concurrent survival of the different bacterial mutants under identical conditions. We describe Escherichia coli survival following neutrophil phagocytosis among three mutants deleted respectively for araB, dps or oxyR. Comparisons were made both by traditional and QPCR methods with similar results and indicate that the survival defect of an oxyR and oxyS mutant described previously can be attributed to the loss of oxyR alone. Deletion of dps, a prominent member of the regulon controlled by the oxyR gene product does not engender a survival defect. We suggest that QPCR analysis can readily compare the relative survival of 10 or more mutants concurrently. QPCR analysis would seem to be especially valuable when experimental conditions are subject to a high degree of sample to sample variability or when the stress producing system involves use of expensive or scarce resources like rare patient cells, cells from children, or the use of genetically modified animal hosts.

Bacterial Outer Membrane Proteins↗
PubMed2002

Broad-spectrum caspase inhibition paradoxically augments cell death in TNF-alpha -stimulated neutrophils.

It is increasingly clear that there are caspase-dependent and -independent mechanisms for the execution of cell death and that the utilization of these mechanisms is stimulus- and cell type-dependent. Intriguingly, broad-spectrum caspase inhibition enhances death receptor agonist-induced cell death in a few transformed cell lines. Endogenously produced oxidants are causally linked to necroticlike cell death in these instances. We report here that broad-spectrum caspase inhibitors effectively attenuated apoptosis induced in human neutrophils by incubation with agonistic anti-Fas antibody or by coincubation with tumor necrosis factor-alpha (TNF-alpha) and cycloheximide ex vivo. In contrast, the same caspase inhibitors could augment cell death upon stimulation by TNF-alpha alone during the 6-hour time course examined. Caspase inhibitor-sensitized, TNF-alpha-stimulated, dying neutrophils exhibit apoptoticlike and necroticlike features. This occurred without apparent alteration in nuclear factor-kappaB (NF-kappaB) activation. Nevertheless, intracellular oxidant production was enhanced and sustained in caspase inhibitor-sensitized, TNF-alpha-stimulated neutrophils obtained from healthy subjects. However, despite reduced or absent intracellular oxidant production following TNF-alpha stimulation, cell death was also augmented in neutrophils isolated from patients with chronic granulomatous disease incubated with a caspase inhibitor and TNF-alpha. These results demonstrate that, in human neutrophils, TNF-alpha induces a caspase-independent but protein synthesis-dependent cell death signal. Furthermore, they suggest that TNF-alpha activates a caspase-dependent pathway that negatively regulates reduced nicotinamide adenine dinucleotide phosphate (NADPH) oxidase activity.

Case-Control Studies↗
PubMed2002

Human neutrophils use the myeloperoxidase-hydrogen peroxide-chloride system to chlorinate but not nitrate bacterial proteins during phagocytosis.

The generation of extracellular oxidants by neutrophils has been widely investigated, but knowledge about the chemical reactions that occur in the phagolysosome, the cellular compartment that kills pathogens, is more limited. One important pathway may involve the production of potent halogenating agents such as hypochlorous acid (HOCl) by the myeloperoxidase-hydrogen peroxide-halide system. However, explorations of the oxidation chemistry of phagolysosomes have been hampered by the organelle's inaccessibility. To overcome this limitation, we recovered Escherichia coli that had been internalized by human neutrophils. We then analyzed the bacterial proteins for 3-chlorotyrosine, a stable marker of damage by HOCl. Mass spectrometric analysis revealed that levels of 3-chlorotyrosine in E. coli proteins increased markedly after the bacteria were internalized by human neutrophils. This increase failed to occur in E. coli exposed to neutrophils deficient in NADPH oxidase or myeloperoxidase, implicating H(2)O(2) and myeloperoxidase in the halogenation reaction. The extent of protein chlorination by normal neutrophils paralleled bacterial killing. Our observations support the view that the phagolysosome of human neutrophils uses the myeloperoxidase-hydrogen peroxide-chloride system to chlorinate bacterial proteins. In striking contrast, human neutrophils failed to nitrate bacterial proteins unless the medium was supplemented with 1 mm nitrite, and the level of nitration was low. Protein chlorination associated with bacterial killing was unaffected by the presence of nitrite in the medium. Nitration required NADPH oxidase but appeared to be independent of myeloperoxidase, suggesting that neutrophils can nitrate proteins through a pathway that requires nitrite but is independent of myeloperoxidase.

Bacterial Proteins↗
PubMed2002

mRNA expression profiles for Escherichia coli ingested by normal and phagocyte oxidase-deficient human neutrophils.

To gain a better understanding of bacterial responses to complex and hostile environments generated within the neutrophil phagosome, we estimated mRNA abundance, using genomic arrays, in Escherichia coli cells ingested by normal and phagocyte oxidase-deficient human neutrophils. Genes regulated by the oxidant sensing transcription factor OxyR were among those strongly induced upon phagocytosis by normal, but not oxidase-deficient, neutrophils. Several genes related to nitrogen metabolism, especially those regulated by the NtrC and NAC proteins and transcribed via the sigma(54) alternative sigma factor, were suppressed by both normal and oxidase-deficient neutrophils. A DeltaoxyRS mutant strain of E. coli was significantly more susceptible than the parent strain to neutrophil-mediated killing, which suggests that OxyR-regulated gene products contribute a measure of resistance to neutrophil antimicrobial systems. The hypersusceptibility of the DeltaoxyRS mutant was attenuated when oxidase-deficient neutrophils were employed, suggesting that much of the protection afforded by the OxyR regulon is against oxidative antimicrobial factors. Expression profiling of phagocytosed bacteria appears to provide useful information about conditions in the phagocytic vacuole and about bacterial defenses mounted in response to this hostile environment.

DNA-Binding Proteins↗