Polyphosphate Kinase Antagonizes Virulence Gene Expression in Francisella tularensis

Abstract

The alarmone ppGpp is a critical regulator of virulence gene expression in In this intracellular pathogen, ppGpp is thought to work in concert with the putative DNA-binding protein PigR and the SspA protein family members MglA and SspA to control a common set of genes. MglA and SspA form a complex that interacts with RNA polymerase (RNAP), and PigR functions by interacting with the RNAP-associated MglA-SspA complex. Prior work suggested that ppGpp indirectly exerts its regulatory effects in by promoting the accumulation of polyphosphate in the cell, which in turn was required for formation of the MglA-SspA complex. Here we show that in , neither polyphosphate nor ppGpp is required for formation of the MglA-SspA complex but that ppGpp promotes the interaction between PigR and the MglA-SspA complex. Moreover, we show that polyphosphate kinase, the enzyme responsible for the synthesis of polyphosphate, antagonizes virulence gene expression in, a finding that is inconsistent with the notion that polyphosphate accumulation promotes virulence gene expression in this organism. Our findings identify polyphosphate kinase as a novel negative regulator of virulence gene expression in and support a model in which ppGpp exerts its positive regulatory effects by promoting the interaction between PigR and the MglA-SspA complex. In , MglA and SspA form a complex that associates with RNA polymerase to positively control the expression of key virulence genes. The MglA-SspA complex works together with the putative DNA-binding protein PigR and the alarmone ppGpp. PigR functions by interacting directly with the MglA-SspA complex, but how ppGpp exerts its effects was unclear. Prior work indicated that ppGpp acts by promoting the accumulation of polyphosphate, which is required for MglA and SspA to interact. Here we show that formation of the MglA-SspA complex does not require polyphosphate. Furthermore, we find that polyphosphate antagonizes the expression of virulence genes in Thus, ppGpp does not promote virulence gene expression in this organism through an effect on polyphosphate.

Survival of the Fittest: The Relationship of (p)ppGpp With Bacterial Virulence

“…Because of its high lethality in its pneumonic form and extremely low infectious dose, F. tularensis is classified by the United States Center for Disease Control (CDC) as a Category A select agent. Several studies have implicated (p)ppGpp with virulence gene expression of F. tularensis , with the most current model indicating that (p)ppGpp promotes virulence in this pathogen by activating transcription of the Francisella pathogenicity island (FPI) through interactions with the DNA-binding protein PigR and the MglA-SspA-RNAP complex ( Cuthbert et al, 2017 ; Rohlfing et al, 2018 ). Indeed, virulence of a double Δ relA Δ spoT strain was shown to be attenuated in intranasally infected mice ( Ma et al, 2019 ).…”