A Nitric Oxide-Responsive Transcriptional Regulator NsrR Cooperates With Lrp and CRP to Tightly Control the hmpA Gene in Vibrio vulnificus

Abstract

Nitric oxide (NO) is an important antimicrobial effector produced by the host innate immune system to counteract invading pathogens. To survive and establish a successful infection, a fulminating human pathogen Vibrio vulnificus expresses the hmpA gene encoding an NO dioxygenase in an NO-responsive manner. In this study, we identified an Rrf2-family transcriptional regulator NsrR that is predicted to contain the Fe-S cluster coordinated by three cysteine residues. Transcriptome analysis showed that NsrR controls the expression of multiple genes potentially involved in nitrosative stress responses. Particularly, NsrR acts as a strong repressor of hmpA transcription and relieves the repression of hmpA upon exposure to NO. Notably, nsrR and hmpA are transcribed divergently, and their promoter regions overlap with each other. Molecular biological analyses revealed that NsrR directly binds to this overlapping promoter region, which is alleviated by loss of the Fe-S cluster, leading to the subsequent derepression of hmpA under nitrosative stress. We further found that a leucine-responsive regulatory protein (Lrp) negatively regulates hmpA in an NsrR-dependent manner by directly binding to the promoter region, presumably resulting in a DNA conformation change to support the repression by NsrR. Meanwhile, a cyclic AMP receptor protein (CRP) positively regulates hmpA probably through repression of nsrR and lrp by directly binding to each promoter region in a sequential cascade. Altogether, this collaborative regulation of NsrR along with Lrp and CRP enables an elaborate control of hmpA transcription, contributing to survival under host-derived nitrosative stress and thereby the pathogenesis of V. vulnificus.

Complex regulatory networks of virulence factors in Vibrio vulnificus

“…In the glucose-poor later stage of infection, CRP-mediated induction of PlpA, VvhA, and VvpE would lead to further inflammation and disease development (Figure 2). CRP also upregulates the expression of hmpA encoding a nitric oxide dioxygenase HmpA essential for survival under nitrosative stress [66]. This observation suggests that CRP coordinates alternative energy metabolism and detoxification of resulting byproducts in glucose-poor conditions.…”

S-Nitrosylation of the virulence regulator AphB promotes Vibrio cholerae pathogenesis

“…cholerae deploys AphB to repress hmpA , which is only activated by NorR in the presence of NO [ 11 ]. In other pathogenic bacteria, such as Salmonella , HmpA production needs to be tightly regulated [ 23 , 24 , 34 ] because under aerobic conditions without nitrosative stress, elevated expression of hmpA increases bacterial susceptibility to ROS. We showed here that constitutive expression of hmpA in V .…”

Transcriptomic Analysis Reveals That Municipal Wastewater Effluent Enhances Vibrio vulnificus Growth and Virulence Potential

“…The role of these transcriptional regulators in the hmpA regulation was evaluated by exposing a V. vulnificus lrp -deletion mutant and a crp -deletion mutant to elevated NO concentration. The results of these studies suggest that CRP indirectly regulates hmpA through repression of nsrR and Lrp by enhancing repression activity of NnrR through DNA structure remodeling ( Choi et al, 2021 ). In the wastewater effluent-amended cultures, lrp and crp genes were downregulated while nsrR and VvhmpA genes were upregulated, suggesting that effluent may enhance the virulence potential of V. vulnificus .…”