GuaA and GuaB Are Essential forBorrelia burgdorferiSurvival in the Tick-Mouse Infection Cycle

Abstract

Pathogens lacking the enzymatic pathways for de novo purine biosynthesis are required to salvage purines and pyrimidines from the host environment for synthesis of DNA and RNA. Two key enzymes in purine salvage pathways are IMP dehydrogenase (GuaB) and GMP synthase (GuaA), encoded by the guaB and guaA genes, respectively. While these genes are typically found on the chromosome in most bacterial pathogens, the guaAB operon of Borrelia burgdorferi is present on plasmid cp26, which also harbors a number of genes critical for B. burgdorferi viability. Using molecular genetics and an experimental model of the tick-mouse infection cycle, we demonstrate that the enzymatic activities encoded by the guaAB operon are essential for B. burgdorferi mouse infectivity and provide a growth advantage to spirochetes in the tick. These data indicate that the GuaA and GuaB proteins are critical for the survival of B. burgdorferi in the infection cycle and highlight a potential difference in the requirements for purine salvage in the disparate mammalian and tick environments.Purine metabolism is critical for the growth and virulence in mammals of many bacterial pathogens (11,26,29,33,51). Borrelia burgdorferi, the infectious agent of Lyme borreliosis, lacks the genes encoding the enzymes required for de novo nucleotide synthesis (8, 12) and therefore must rely on salvage of purines and pyrimidines from its hosts for nucleic acid biosynthesis (21, 35). Furthermore, B. burgdorferi lacks the genes encoding key enzymes required for a classic purine salvage pathway, including hpt (hypoxanthine-guanine phosphoribosyltransferase), purA (adenylosuccinate synthase), purB (adenylosuccinate lyase), and the locus encoding a ribonucleotide reductase (4,8,12,35,66). Despite the absence of a ribonucleotide reductase, an enzyme critical for the generation of deoxynucleotides through enzymatic reduction of ribonucleotides (32), a novel purine salvage pathway that involves salvage of deoxynucleosides from the host and interconversion of purine bases to deoxynucleosides by BB0426, a deoxyribosyl transferase, has recently been demonstrated for B. burgdorferi (23) (Fig. 1).In its infection cycle, B. burgdorferi passages between two disparate environments with potentially distinct purine availabilities, the tick vector and a mammalian host. Hypoxanthine is the most abundant purine in mammalian blood (17), and it is available for salvage by B. burgdorferi during the blood meal of an infected tick and during the spirochete's transient presence in the mammalian bloodstream. Despite the absence of the hpt gene, we and others have shown that B. burgdorferi is able to transport and incorporate low levels of hypoxanthine (23,35). During mammalian infection B. burgdorferi resides in various tissues, including the skin, heart, bladder, and joints. Adenine has been shown to be ubiquitous in mammalian tissues (61) and therefore is available for salvage by B. burgdorferi. Guanine is present at low levels in mammalian blood and tissues (17, 61); however, the amount m...

Helicobacter pylori Relies Primarily on the Purine Salvage Pathway for Purine Nucleotide Biosynthesis

“…Often, numerous purine nucleotide biosynthesis genes are found in the same operons, allowing them to be controlled by a single, regulatory mechanism (34)(35)(36). Regulators for purine nucleotide biosynthesis and salvage pathways are abundant; regulatory promoter-binding proteins (33,34) and riboswitches (4,43) are utilized for many of these genes in most other bacteria.…”

“…The two currently most examined genes in purine nucleotide biosynthesis, given their importance in both de novo and salvage pathways, are guaB and guaA. Bacterial strains with mutations or deletions in these two genes have been shown to be severely attenuated for growth in Salmonella enterica serotype Typhimurium (23), Yersinia pestis (52), Francisella tularensis (63), Borrelia burgdorferi (36), and Shigella flexneri (40).…”

“…Genes comprising either pathway often are found in operons (34)(35)(36) and are often regulated by the same regulatory promoterbinding proteins (33,34) or riboswitches (4,43), generating feedback loops that allow the organism to alternate between the two synthesis pathways while maintaining adequate purine nucleotide pools. Additionally, the allosteric regulation of nucleotide pools has also been shown in key enzymes in both purine (60,61) and pyrimidine (59) nucleotide biosynthesis, indicating enzyme activity also may add an additional means of regulation.…”

BB0238, a Presumed Tetratricopeptide Repeat-Containing Protein, Is Required during Borrelia burgdorferi Mammalian Infection

“…bb0239 and bb0238, on the other hand, are oriented in the same direction on the chromosome with a relatively small intergenic region (45 bp), suggesting they may be cotranscribed. This is especially relevant considering that bb0239 is predicted to be involved in B. burgdorferi purine salvage, and this pathway is essential for mammalian infection (39)(40)(41).…”

“…Though the DMC experiments do not address tissue-specific differences in nutrient requirements or availability, the experiments may identify a general in vivo growth defect that could result in the attenuated-infectivity phenotype similar to that seen with pncA (54). As noted above, the gene adjacent to bb0239 is predicted to encode a deoxyguanosine/deoxyadenosine kinase involved in B. burgdorferi purine salvage, and this pathway is essential for mammalian infection (39,40). Due to the physical proximity of bb0238 and bb0239 in the genome, it is possible that bb0238 may play an ancillary role in purine salvage.…”