Arginine Is a Critical Substrate for the Pathogenesis of Pseudomonas aeruginosa in Burn Wound Infections.

Environmental conditions affect bacterial behavior and can greatly influence the course of an infection. However, the environmental cues that elicit bacterial responses in specific infection sites are relatively unknown. Pseudomonas aeruginosa is ubiquitous in nature and typically innocuous. However, it is also one of the most prevalent causes of fatal sepsis in burn wound patients. The aim of this study was to determine the impact of environmental factors, specifically the availability of arginine, on the pathogenesis of P. aeruginosa in burn wound infections. Comparison of burned versus noninjured tissue revealed that l-arginine (l-Arg) was significantly depleted in burn wounds as a consequence of elevated arginase produced by myeloid-derived suppressor cells. We also observed that l-Arg was a potent chemoattractant for P. aeruginosa, and while low concentrations of l-Arg increased P. aeruginosa's swimming motility, high concentrations resulted in diminished swimming. Based on these observations, we tested whether the administration of exogenous l-Arg into the burn wound could attenuate the virulence of P. aeruginosa in thermally injured mice. Administration of l-Arg resulted in decreased P. aeruginosa spread and sepsis and increased animal survival. Taken together, these data demonstrate that the availability of environmental arginine greatly influences the virulence of P. aeruginosa in vivo and may represent a promising phenotype-modulating tool for future therapeutic avenues.IMPORTANCE Despite our growing understanding of the pathophysiology of burn wounds and the evolution of techniques and practices to manage infections, sepsis remains a significant medical concern for burn patients. P. aeruginosa continues to be a leader among all causes of bacteremic infections due to its tendency to cause complications in immunocompromised patients and its ubiquitous presence in the hospital setting. With the unforgiving emergence of multidrug-resistant strains, it is critical that alternative strategies to control or prevent septic infections in burn patients be developed in parallel with novel antimicrobial agents. In this study, we observed that administration of l-Arg significantly reduced bacterial spread and sepsis in burned mice infected with P. aeruginosa Given the safety of l-Arg in high doses and its potential wound-healing benefits, this conditionally essential amino acid may represent a useful tool to modulate bacterial behavior in vivo and prevent sepsis in burn patients.

Barrier Crossing in Escherichia coli Chemotaxis.

We study cell navigation in spatiotemporally complex environments by developing a microfluidic racetrack device that creates a traveling wave with multiple peaks and a tunable wave speed. We find that while the population-averaged chemotaxis drift speed increases with wave speed for low wave speed, it decreases sharply for high wave speed. This reversed dependence of population-averaged chemotaxis drift speed on wave speed is caused by a "barrier-crossing" phenomenon, where a cell hops backwards from one peak attractant location to the peak behind by crossing an unfavorable (barrier) region with low attractant concentrations. By using a coarse-grained model of chemotaxis, we map bacterial motility in an attractant field to the random motion of an overdamped particle in an effective potential. The observed barrier-crossing phenomenon of living cells and its dependence on the spatiotemporal profile of attractant concentration are explained quantitatively by Kramers reaction rate theory.

Singly Flagellated Pseudomonas aeruginosa Chemotaxes Efficiently by Unbiased Motor Regulation.

UNLABELLED: Pseudomonas aeruginosais an opportunistic human pathogen that has long been known to chemotax. More recently, it has been established that chemotaxis is an important factor in the ability ofP. aeruginosato make biofilms. Genes that allowP. aeruginosato chemotax are homologous with genes in the paradigmatic model organism for chemotaxis,Escherichia coli However,P. aeruginosais singly flagellated andE. colihas multiple flagella. Therefore, the regulation of counterclockwise/clockwise flagellar motor bias that allowsE. colito efficiently chemotax by runs and tumbles would lead to inefficient chemotaxis byP. aeruginosa, as half of a randomly oriented population would respond to a chemoattractant gradient in the wrong sense. HowP. aeruginosaregulates flagellar rotation to achieve chemotaxis is not known. Here, we analyze the swimming trajectories of single cells in microfluidic channels and the rotations of cells tethered by their flagella to the surface of a variable-environment flow cell. We show thatP. aeruginosachemotaxes by symmetrically increasing the durations of both counterclockwise and clockwise flagellar rotations when swimming up the chemoattractant gradient and symmetrically decreasing rotation durations when swimming down the chemoattractant gradient. Unlike the case forE. coli, the counterclockwise/clockwise bias stays constant forP. aeruginosa We describeP. aeruginosa's chemotaxis using an analytical model for symmetric motor regulation. We use this model to do simulations that show that, givenP. aeruginosa's physiological constraints on motility, its distinct, symmetric regulation of motor switching optimizes chemotaxis. IMPORTANCE: Chemotaxis has long been known to strongly affect biofilm formation by the opportunistic human pathogenP. aeruginosa, whose essential chemotaxis genes have homologues inE. coli, which achieves chemotaxis by biasing the relative probability of counterclockwise and clockwise flagellar rotation. However, the physiological difference between multiflagellatedE. coliand singly flagellatedP. aeruginosaimplies that biased motor regulation should preventP. aeruginosapopulations from chemotaxing efficiently. Here, we used experiments, analytical modeling, and simulations to demonstrate thatP. aeruginosauses unbiased, symmetric regulation of the flagellar motor to maximize its chemotaxis efficiency. This mode of chemotaxis was not previously known and demonstrates a new variant of a paradigmatic signaling system in an important human pathogen.

MDM4 regulation by the let-7 miRNA family in the DNA damage response of glioma cells.

Despite extensive investigation into the role of let-7 miRNAs in pathological tumor processes, their involvement in the DNA damage response remains unclear. Here we show that most let-7 family members down-regulate MDM4 expression via binding to MDM4 mRNA at a conserved DNA sequence. Expression of exogenous let-7 miRNA mimics decreased MDM4 protein but not mRNA levels. Several DNA damage reagents increased let-7 expression, thereby decreasing MDM4 protein levels in glioma cells. Inhibition of endogenous let-7 with antisense RNAs rescued MDM4 protein levels with or without MG132, a proteasome-dependent degradation inhibitor. An MDM4 mutation identified in a glioma patient was associated with loss of the putative MDM4 target site. Therefore, let-7 binding to MDM4 is implicated in the DNA damage response.