The MisR Response Regulator Is Necessary for Intrinsic Cationic Antimicrobial Peptide and Aminoglycoside Resistance in Neisseria gonorrhoeae

Abstract

dDuring infection, the sexually transmitted pathogen Neisseria gonorrhoeae (the gonococcus) encounters numerous host-derived antimicrobials, including cationic antimicrobial peptides (CAMPs) produced by epithelial and phagocytic cells. CAMPs have both direct and indirect killing mechanisms and help link the innate and adaptive immune responses during infection. Gonococcal CAMP resistance is likely important for avoidance of host nonoxidative killing systems expressed by polymorphonuclear granulocytes (e.g., neutrophils) and intracellular survival. Previously studied gonococcal CAMP resistance mechanisms include modification of lipid A with phosphoethanolamine by LptA and export of CAMPs by the MtrCDE efflux pump. In the related pathogen Neisseria meningitidis, a two-component regulatory system (2CRS) termed MisR-MisS has been shown to contribute to the capacity of the meningococcus to resist CAMP killing. We report that the gonococcal MisR response regulator but not the MisS sensor kinase is involved in constitutive and inducible CAMP resistance and is also required for intrinsic low-level resistance to aminoglycosides. The 4-to 8-fold increased susceptibility of misR-deficient gonococci to CAMPs and aminoglycosides was independent of phosphoethanolamine decoration of lipid A and the levels of the MtrCDE efflux pump and seemed to correlate with a general increase in membrane permeability. Transcriptional profiling and biochemical studies confirmed that expression of lptA and mtrCDE was not impacted by the loss of MisR. However, several genes encoding proteins involved in membrane integrity and redox control gave evidence of being MisR regulated. We propose that MisR modulates the levels of gonococcal susceptibility to antimicrobials by influencing the expression of genes involved in determining membrane integrity. N eisseria gonorrhoeae is a Gram-negative diplococcus and the causative agent of the sexually transmitted infection termed gonorrhea, which is currently the second most reported infection in the United States (1); an estimated 78 million new cases of gonorrhea occurred worldwide in 2012 (2). In addition to the high worldwide prevalence of gonorrhea, strains with resistance to currently or formerly used antibiotics have emerged, and concern has been voiced that without new effective antimicrobials, some cases of gonorrhea may be difficult to treat in future years (3). In addition to its ability to resist classical antibiotics used in treatment, gonococci have evolved mechanisms to evade the antimicrobial action of host compounds that participate in the innate host defense during infection. For instance, the ability of gonococci to resist the antibiotic-like action of host cationic antimicrobial peptides (CAMPs), such as defensins (4) or larger antimicrobial proteins (e.g., bactericidal permeability-increasing protein [5], cathepsin G [6], and CAP37 [7]), has been implicated in its survival within human polymorphonuclear granulocytes (PMNs) (8, 9).Broadly, there are five known ways in which gonococci res...

Transcriptome Analysis ofNeisseria gonorrhoeaeDuring Natural Infection Reveals Differential Expression of Antibiotic Resistance Determinants Between Men and Women

“…Expression of the pilQ gene, associated with antibiotic permeability (14), was significantly decreased by 1.78-fold in vivo compared to in vitro ( Figure 4A ). The MisRS two-component regulatory system is important for resistance to host-derived antimicrobial peptides and antibiotics (61); and though not statistically significant, misR had 2.08-fold higher expression in vivo than in vitro , supporting its important role in vivo . The expression of the Mtr efflux pump was also closely examined; expression of mtrA , which can influence expression of mtrCDE (62), was significantly decreased by 2.75 fold in vivo ( Figure 4B ).…”

Both MisR (CpxR) and MisS (CpxA) Are Required for Neisseria gonorrhoeae Infection in a Murine Model of Lower Genital Tract Infection

“…Bacteria often use two-component systems (TCSs) to adapt to changing microenvironments. N. gonorrhoeae contains three TCSs: NarPQ, NtrXY, and MisRS (40)(41)(42). NarPQ and NtrXY regulate gonococcal adaptation to an anaerobic environment and the expression of respiratory genes, respectively.…”

“…In the meningococcus, MisRS is required for virulence in a murine model and is involved in the regulation of over 78 genes belonging to various functional categories, such as protein folding and iron assimilation; a MisR-binding sequence was also identified (43)(44)(45)(46)(47). In N. gonorrhoeae strain FA19, MisR (NGEG_00293) is required for resistance to cationic antimicrobial peptides (CAMPs) and aminoglycosides as well as the maintenance of membrane integrity (42). Compared to its parent, 95 genes are differentially expressed in the misR mutant; these genes encode proteins involved in membrane permeability and redox control (42).…”

“…In N. gonorrhoeae strain FA19, MisR (NGEG_00293) is required for resistance to cationic antimicrobial peptides (CAMPs) and aminoglycosides as well as the maintenance of membrane integrity (42). Compared to its parent, 95 genes are differentially expressed in the misR mutant; these genes encode proteins involved in membrane permeability and redox control (42). MisRS has homology to multiple TCSs, such as PhoP/PhoQ and CpxRA (42).…”

“…Compared to its parent, 95 genes are differentially expressed in the misR mutant; these genes encode proteins involved in membrane permeability and redox control (42). MisRS has homology to multiple TCSs, such as PhoP/PhoQ and CpxRA (42). Based on the function or role of MisRS with respect to the maintenance of membrane integrity and the analysis of MisR-dependent targets, the authors of the latter study opined that MisRS functionally resembles CpxRA (42).…”