Intracellular Concentrations of Borrelia burgdorferi Cyclic Di-AMP Are Not Changed by Altered Expression of the CdaA Synthase

Savage, Christina R.; Arnold, William K.; Gjevre-Nail, Alexandra; Koestler, Benjamin J.; Bruger, Eric L.; Barker, Jeffrey R.; Waters, Christopher M.; Stevenson, Brian

doi:10.1371/journal.pone.0125440

Cited by 15 publications

(25 citation statements)

References 75 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…The levels of c-di-AMP in the DAC overexpression strain were significantly altered, i.e., 1.7-fold higher ( p < 0.05), compared to the WT strain. Whereas this is lower than might be anticipated based on gene expression data, some bacterial strains which overaccumulate DAC mRNA show no upregulation of intracellular c-di-AMP levels suggesting possible negative feedback mechanisms on DAC activity in some cases ( Savage et al, 2015 ). The levels of c-di-AMP in the PDE overexpression Synechocystis strain were on average half as much as the WT strain, although this level was not significantly different.…”

Section: Resultsmentioning

confidence: 93%

Homeostasis of Second Messenger Cyclic-di-AMP Is Critical for Cyanobacterial Fitness and Acclimation to Abiotic Stress

et al. 2018

Self Cite

View full text Add to dashboard Cite

Second messengers are intracellular molecules regulated by external stimuli known as first messengers that are used for rapid organismal responses to dynamic environmental changes. Cyclic di-AMP (c-di-AMP) is a relatively newly discovered second messenger implicated in cell wall homeostasis in many pathogenic bacteria. C-di-AMP is synthesized from ATP by diadenylyl cyclases (DAC) and degraded by specific c-di-AMP phosphodiesterases (PDE). C-di-AMP DACs and PDEs are present in all sequenced cyanobacteria, suggesting roles for c-di-AMP in the physiology and/or development of these organisms. Despite conservation of these genes across numerous cyanobacteria, the functional roles of c-di-AMP in cyanobacteria have not been well-investigated. In a unique feature of cyanobacteria, phylogenetic analysis indicated that the broadly conserved DAC, related to CdaA/DacA, is always co-associated in an operon with genes critical for controlling cell wall synthesis. To investigate phenotypes regulated by c-di-AMP in cyanobacteria, we overexpressed native DAC (sll0505) and c-di-AMP PDE (slr0104) genes in the cyanobacterium Synechocystis sp. PCC 6803 (hereafter Synechocystis) to increase and decrease intracellular c-di-AMP levels, respectively. DAC- and PDE-overexpression strains, showed abnormal aggregation phenotypes, suggesting functional roles for regulating c-di-AMP homeostasis in vivo. As c-di-AMP may be implicated in osmotic responses in cyanobacteria, we tested whether sorbitol and NaCl stresses impacted expression of sll0505 and slr0104 or intracellular c-di-AMP levels in Synechocystis. Additionally, to determine the range of cyanobacteria in which c-di-AMP may function, we assessed c-di-AMP levels in two unicellular cyanobacteria, i.e., Synechocystis and Synechococcus elongatus PCC 7942, and two filamentous cyanobacteria, i.e., Fremyella diplosiphon and Anabaena sp. PCC 7120. C-di-AMP levels responded differently to abiotic stress signals in distinct cyanobacteria strains, whereas salt stress uniformly impacted another second messenger cyclic di-GMP in cyanobacteria. Together, these results suggest regulation of c-di-AMP homeostasis in cyanobacteria and implicate a role for the second messenger in maintaining cellular fitness in response to abiotic stress.

show abstract

Section: Resultsmentioning

confidence: 93%

Homeostasis of Second Messenger Cyclic-di-AMP Is Critical for Cyanobacterial Fitness and Acclimation to Abiotic Stress

et al. 2018

Self Cite

View full text Add to dashboard Cite

show abstract

“…There is limited but significant evidence that c-di-AMP is produced by (some) Gram-negative organisms. This has been noted in Chlamydia trachomatis [ 31 ] and Borrelia burgdorferi [ 32 , 33 ].…”

Section: Introductionmentioning

confidence: 79%

The Characterization of Escherichia coli CpdB as a Recombinant Protein Reveals that, besides Having the Expected 3´-Nucleotidase and 2´,3´-Cyclic Mononucleotide Phosphodiesterase Activities, It Is Also Active as Cyclic Dinucleotide Phosphodiesterase

et al. 2016

View full text Add to dashboard Cite

Endogenous cyclic diadenylate phosphodiesterase activity was accidentally detected in lysates of Escherichia coli BL21. Since this kind of activity is uncommon in Gram-negative bacteria, its identification was undertaken. After partial purification and analysis by denaturing gel electrophoresis, renatured activity correlated with a protein identified by fingerprinting as CpdB (cpdB gene product), which is annotated as 3´-nucleotidase / 2´,3´-cyclic-mononucleotide phosphodiesterase, and it is synthesized as a precursor protein with a signal sequence removable upon export to the periplasm. It has never been studied as a recombinant protein. The coding sequence of mature CpdB was cloned and expressed as a GST fusion protein. The study of the purified recombinant protein, separated from GST, confirmed CpdB annotation. The assay of catalytic efficiencies (kcat/Km) for a large substrate set revealed novel CpdB features, including very high efficiencies for 3´-AMP and 2´,3´-cyclic mononucleotides, and previously unknown activities on cyclic and linear dinucleotides. The catalytic efficiencies of the latter activities, though low in relative terms when compared to the major ones, are far from negligible. Actually, they are perfectly comparable to those of the ‘average’ enzyme and the known, bona fide cyclic dinucleotide phosphodiesterases. On the other hand, CpdB differs from these enzymes in its extracytoplasmic location and in the absence of EAL, HD and DHH domains. Instead, it contains the domains of the 5´-nucleotidase family pertaining to the metallophosphoesterase superfamily, although CpdB lacks 5´-nucleotidase activity. The possibility that the extracytoplasmic activity of CpdB on cyclic dinucleotides could have physiological meaning is discussed.

show abstract

“…The flaB transcript is frequently used as a qRT-PCR reference transcript [ 58 , 69 , 70 ]. The recA mRNA has also been used on occasion as a reference, under the same assumption of invariability [ 16 , 18 ]. However, our RNA-Seq analyses consistently revealed differences in both flaB and recA mRNA levels when comparing early or mid-exponential to stationary phases ( S4 and S5 Tables).…”

Section: Resultsmentioning

confidence: 99%

“…burgdorferi gene and protein regulation, including DNA-binding proteins, two-component sensory mechanisms, and enzymes that synthesize/degrade modified nucleotide alarmones (e.g. [ 1 , 11 – 18 ]). Detailed understanding of the regulatory networks at play during B .…”

Section: Introductionmentioning

confidence: 99%

RNA-Seq of Borrelia burgdorferi in Multiple Phases of Growth Reveals Insights into the Dynamics of Gene Expression, Transcriptome Architecture, and Noncoding RNAs

Arnold¹,

Savage²,

Brissette³

et al. 2016

PLoS ONE

Self Cite

110

View full text Add to dashboard Cite

Borrelia burgdorferi, the agent of Lyme disease, differentially expresses numerous genes and proteins as it cycles between mammalian hosts and tick vectors. Insights on regulatory mechanisms have been provided by earlier studies that examined B. burgdorferi gene expression patterns during cultivation. However, prior studies examined bacteria at only a single time point of cultivation, providing only a snapshot of what is likely a dynamic transcriptional program driving B. burgdorferi adaptations to changes during culture growth phases. To address that concern, we performed RNA sequencing (RNA-Seq) analysis of B. burgdorferi cultures at early-exponential, mid-exponential, and early-stationary phases of growth. We found that expression of nearly 18% of annotated B. burgdorferi genes changed significantly during culture maturation. Moreover, genome-wide mapping of the B. burgdorferi transcriptome in different growth phases enabled insight on transcript boundaries, operon structures, and identified numerous putative non-coding RNAs. These RNA-Seq data are discussed and presented as a resource for the community of researchers seeking to better understand B. burgdorferi biology and pathogenesis.

show abstract

Intracellular Concentrations of Borrelia burgdorferi Cyclic Di-AMP Are Not Changed by Altered Expression of the CdaA Synthase

Cited by 15 publications

References 75 publications

Homeostasis of Second Messenger Cyclic-di-AMP Is Critical for Cyanobacterial Fitness and Acclimation to Abiotic Stress

Homeostasis of Second Messenger Cyclic-di-AMP Is Critical for Cyanobacterial Fitness and Acclimation to Abiotic Stress

The Characterization of Escherichia coli CpdB as a Recombinant Protein Reveals that, besides Having the Expected 3´-Nucleotidase and 2´,3´-Cyclic Mononucleotide Phosphodiesterase Activities, It Is Also Active as Cyclic Dinucleotide Phosphodiesterase

RNA-Seq of Borrelia burgdorferi in Multiple Phases of Growth Reveals Insights into the Dynamics of Gene Expression, Transcriptome Architecture, and Noncoding RNAs

Contact Info

Product

Resources

About