Regulation of cid and lrg expression by CodY in Streptococcus mutans

Abstract

The ability of Streptococcus mutans to persist in a variety of adverse environments and to emerge as a numerically dominant member of stable oral biofilm communities are essential elements for its cariogenicity. The S. mutans Cid/Lrg system has been studied as a key player in the integration of complex environmental signals into regulatory networks that modulate virulence and cell homeostasis. Cid/Lrg has also been shown to be closely associated with metabolic pathways of this organism, due to distinct patterns of cid and lrg expression in response to growth phase and glucose/oxygen levels. In this study, a comparison of cid and lrg promoter regions with conserved CodY (a regulator which responds to starvation stress)‐binding motifs revealed the presence of a potential CodY‐binding site, which is arranged similarly in both cid and lrg promoters. Electrophoretic mobility shift assays (EMSAs) and promoter reporter assays demonstrated that expression of the cid and lrg operons is directly mediated by the global transcriptional regulator CodY. DNase I footprinting analyses confirmed the predicted binding sequences for CodY in both the cid and the lrg promoter regions. Overexpression of CodY had no obvious effect on lrgAB expression, but deficiency of CodY still affected lrgAB expression in a lytST ‐overexpressing strain, suggesting that CodY is required for the full regulation of lrgAB by LytST. We also demonstrated that both CodY and CcpA are involved in regulating pyruvate flux and utilization. Collectively, these data show that CodY directly regulates cid and lrg expression, and together with CcpA (previously shown to directly regulate cid and lrg promoters) contributes to coordinating pyruvate uptake and utilization in response to both the external environment and the cellular metabolic status.

Understanding LrgAB Regulation of Streptococcus mutans Metabolism

“…Collectively, these findings suggest the catabolic rate of pyruvate is an important trigger to take up external pyruvate through LrgAB, and vice versa . Results, moreover, are consistent with previous results demonstrating both lrgAB and pta-ackA expression are directly under the control of two global regulators CcpA and CodY ( Kim and Burne, 2017 ; Kim et al, 2019 ; Ahn et al, 2020c ). Considering Pdh is highly regulated by various metabolic intermediates, including pyruvate ( Hansen and Henning, 1966 ; Schwartz and Reed, 1970 ; Shen and Atkinson, 1970 ; Datta, 1991 ), and that pdh genes are commonly upregulated in response to three environmental stressors (i.e., aerobic, heat and vancomycin challenge) ( Rice et al, 2017 ), it is reasonable to posit that pyruvate catabolism and lrg regulation work together and are modulated by the cell’s metabolic status and environmental condition.…”

“…Interestingly, the lrgAB mutation was also shown to affect major metabolic pathways associated with carbohydrate, amino acid, fatty acid/lipid, nucleotide metabolism and transport ( Ahn et al, 2017 ; Rice et al, 2017 ). Recently, we reported that LrgAB functions as a stationary phase pyruvate uptake system ( Ahn et al, 2019 ) and its function is regulated by two global regulators, CcpA ( Kim et al, 2019 ) and CodY ( Ahn et al, 2020c ), that are important for cellular responses to altered carbohydrate availability and amino acid limitations ( Henkin, 1996 ; Sonenshein, 2005 , 2007 ; Deutscher, 2008 ; Gorke and Stulke, 2008 ). More interestingly, we demonstrated that excess pyruvate produced during growth can be subsequently utilized as a carbon source during stationary phase ( Ahn et al, 2019 , 2020a ), presumably promoting long-term survival of S. mutans when in a metabolically competitive environment with other oral microbiota.…”

The Pta-AckA Pathway Regulates LrgAB-Mediated Pyruvate Uptake in Streptococcus mutans

“…Although the disruption of the Pta-AckA pathway inhibits acetate production, lack of acetate does not seem to directly result in the observed dramatic reduction of lrgAB induction and pyruvate uptake, because the inhibitory effect of an inactive Pta-AckA pathway on lrgAB induction was observed in both FMC11 (acetate-enriched) and TV3 (acetate-limited). Nevertheless, the fact that, like the LrgAB system [ 30 , 31 ], the Pta-AckA pathway is also tightly regulated by CcpA and CodY [ 54 ], implicates that the pyruvate uptake and acetate production systems may be linked in a metabolically complex manner which warrants further study.…”

“…We recently revealed that the S. mutans LrgAB system—which is directly and positively regulated by the LytST two-component system (TCS) [ 28 , 29 ] and hypothesized to induce cell death and lysis with its partner operon CidAB—functions as a stationary phase pyruvate uptake system, with its activity modulated in response to glucose and oxygen levels [ 26 ]. More recently, we found that LrgAB pyruvate uptake activity is under the control of two global regulators, CcpA [ 30 ] and CodY [ 31 ], and is affected by the composition (i.e., acetate and potassium ion) of the growth medium [ 27 ]. This illustrates the metabolic complexity of LrgAB activity and pyruvate uptake regulation in S. mutans.…”

Extracellular DNA released by glycine-auxotrophic Staphylococcus epidermidis small colony variant facilitates catheter-related infections

“…However, programmed cell death is mediated by a battery of coordinated effectors among which CidA and LrgA are tightly under the control of metabolic signals, such as carbon overflow 57 . Though originally identified in S. aureus , CidA and LrgA proteins are found to be conserved in several other species including S. epidermidis 54 , 58 , 59 . As CidA shares higher structural similarity with the bacteriophage-encoded holin protein, it was proposed to induce cell lysis and death in a manner similar with holin 48 , 60 .…”