Bacterial Second Messenger Cyclic di-AMP Modulates the Competence State in Streptococcus pneumoniae

Abstract

Streptococcus pneumoniae (the pneumococcus) is a naturally competent organism that causes diseases such as pneumonia, otitis media, and bacteremia. The essential bacterial second messenger cyclic di-AMP (c-di-AMP) is an emerging player in the stress responses of many pathogens. In S. pneumoniae, c-di-AMP is produced by a diadenylate cyclase, CdaA, and cleaved by phosphodiesterases Pde1 and Pde2. c-di-AMP binds a transporter of K+ (Trk) family protein, CabP, which subsequently halts K+ uptake via the transporter TrkH. Recently, it was reported that Pde1 and Pde2 are essential for pneumococcal virulence in mouse models of disease. To elucidate c-di-AMP-mediated transcription that may lead to changes in pathogenesis, we compared the transcriptomes of wild-type (WT) and Δpde1 Δpde2 strains by transcriptome sequencing (RNA-Seq) analysis. Notably, we found that many competence-associated genes are significantly upregulated in the Δpde1 Δpde2 strain compared to the WT. These genes play a role in DNA uptake, recombination, and autolysis. Competence is induced by a quorum-sensing mechanism initiated by the secreted factor competence-stimulating peptide (CSP). Surprisingly, the Δpde1 Δpde2 strain exhibited reduced transformation efficiency compared to WT bacteria, which was c-di-AMP dependent. Transformation efficiency was also directly related to the [K+] in the medium, suggesting a link between c-di-AMP function and the pneumococcal competence state. We found that a strain that possesses a V76G variation in CdaA produced less c-di-AMP and was highly susceptible to CSP. Deletion of cabP and trkH restored the growth of these bacteria in medium with CSP. Overall, our study demonstrates a novel role for c-di-AMP in the competence program of S. pneumoniae. IMPORTANCE Genetic competence in bacteria leads to horizontal gene transfer, which can ultimately affect antibiotic resistance, adaptation to stress conditions, and virulence. While the mechanisms of pneumococcal competence signaling cascades have been well characterized, the molecular mechanism behind competence regulation is not fully understood. The bacterial second messenger c-di-AMP has previously been shown to play a role in bacterial physiology and pathogenesis. In this study, we provide compelling evidence for the interplay between c-di-AMP and the pneumococcal competence state. These findings not only attribute a new biological function to this dinucleotide as a regulator of competence, transformation, and survival under stress conditions in pneumococci but also provide new insights into how pneumococcal competence is modulated.

Cyclic Di-adenosine Monophosphate Regulates Metabolism and Growth in the Oral Commensal Streptococcus mitis

“…Intracellularly, it functions as a second messenger, relaying intracellular and extracellular signals into bacterial cell responses. It regulates a broad range of physiological processes such as bacterial growth and cell size, biofilm formation, virulence, potassium homeostasis, central metabolism, antibiotic susceptibility, maintenance of DNA integrity, and natural transformation [ 1 , 2 , 3 , 4 , 5 , 6 , 7 ]. C-di-AMP can also act as an extracellular ligand involved in bacteria-host cross-talk, modulating the host’s immune response.…”

c-di-AMP is essential for the virulence of Enterococcus faecalis

“…To evaluate the impact of c-di-AMP dysregulation at a global level, we used RNaseq to compare the transcriptome of E. faecalis OG1RF, ∆dhhP∆gdpP and ∆cdaA strains. To our knowledge, this is the first time the transcriptome of a bona fide c-di-AMP 0 strain was obtained, as previous global transcriptional analysis were restricted to strains with high c-di-AMP levels (9,35,67). In B. subtilis, c-di-AMP regulates transcription by interacting with a c-di-AMPspecific riboswitch (2) with several c-di-AMP riboswitch-regulated genes involved in K + and osmolyte transport, and cell wall metabolism (73).…”

“…Second messenger nucleotides are synthesized by bacteria in response to internal or external stimuli and are used to reprogram cell physiology through physical interactions with proteins (allosteric regulation), RNA riboswitches, or both (1)(2)(3). Despite being a relatively recent discovery, cyclic di-adenosine monophosphate (c-di-AMP) has been shown to control a variety of bacterial processes including osmoregulation, cell envelope homeostasis, stress and antibiotic tolerance, biofilm formation, central metabolism, DNA repair, genetic competence and sporulation (4)(5)(6)(7)(8)(9)(10)(11)(12)(13)(14)(15)(16). In addition, c-di-AMP plays an important role in host-pathogen interactions as it can be exported to the bacterial extracellular milieu promoting a potent type I interferon immune response (17).…”

A Cyclic di-GMP Network Is Present in Gram-Positive Streptococcus and Gram-Negative Proteus Species

“…Previously, a cyclic di-AMP signaling network had been ubiquitously identified in streptococcal species . Cyclic di-AMP signaling has been mainly investigated in the human pathogens S. pneumoniae and S. pyogenes , the dental caries causing S. mutans , and the animal pathogen S. suis , where c-di-AMP signaling controls exopolysaccharide biosynthesis and biofilm formation, antimicrobial resistance, the competence status, and regulation of host immunity, among other phenotypes. − In S. gallolyticus subsp. gallolyticus , c-di-AMP signaling has been shown to promote osmoresistance, alter cell morphology, and inhibit biofilm formation and host–cell interactions .…”