AI-3 Synthesis Is Not Dependent onluxSinEscherichia coli

Abstract

The quorum-sensing (QS) signal autoinducer-2 (AI-2) has been proposed to promote interspecies signaling in a broad range of bacterial species. AI-2 is spontaneously derived from 4,5-dihydroxy-2,3-pentanedione that, along with homocysteine, is produced by cleavage of S-adenosylhomocysteine (SAH) and S-ribosylhomocysteine by the Pfs and LuxS enzymes. Numerous phenotypes have been attributed to AI-2 QS signaling using luxS mutants. We have previously reported that the luxS mutation also affects the synthesis of the AI-3 autoinducer that activates enterohemorrhagic Escherichia coli virulence genes. Here we show that several species of bacteria synthesize AI-3, suggesting a possible role in interspecies bacterial communication. The luxS mutation leaves the cell with only one pathway, involving oxaloacetate and L-glutamate, for de novo synthesis of homocysteine. The exclusive use of this pathway for homocysteine production appears to alter metabolism in the luxS mutant, leading to decreased levels of AI-3. The addition of aspartate and expression of an aromatic amino acid transporter, as well as a tyrosine-specific transporter, restored AI-3-dependent phenotypes in an luxS mutant. The defect in AI-3 production, but not in AI-2 production, in the luxS mutant was restored by expressing the Pseudomonas aeruginosa S-adenosylhomocysteine hydrolase that synthesizes homocysteine directly from SAH. Furthermore, phenotype microarrays revealed that the luxS mutation caused numerous metabolic deficiencies, while AI-3 signaling had little effect on metabolism. This study examines how AI-3 production is affected by the luxS mutation and explores the roles of the LuxS/AI-2 system in metabolism and QS.

Heterologous Expression of sahH Reveals That Biofilm Formation Is Autoinducer-2-independent in Streptococcus sanguinis but Is Associated with an Intact Activated Methionine Cycle

“…5, 9, and 12). From experiments in Gram-negative species, it is known that this pathway can produce sufficient amounts of methionine (15,16). Expression of SahH in the S. sanguinis LuxS mutant restored biofilm development back to WT levels without any detectable AI-2 production (Figs.…”

“…Alternatively, an additional pathway for methionine production as described in Gram-negative could exist in Grampositive species. For instance, in E. coli homocysteine can be produced via a pathway starting with oxalacetate (16).…”

“…The functionality of a SahH complemented interrupted AMC has been demonstrated only in Gram-negative bacteria (15,16). Consequently, in our study we examined for the suitability of such heterologous complementation in a Gram-positive bacterium.…”

The two-component system QseEF and the membrane protein QseG link adrenergic and stress sensing to bacterial pathogenesis

“…Primers used in qRT-PCR analysis are listed in Table S2. The qRT-PCR analysis was conducted by using an Applied Biosystems ABI 7500 sequence detection system using a 1-step reaction as previously described (12). The rpoZ gene was used as an internal control.…”

“…EHEC recognizes the AI-3 that is produced by the intestinal microbial flora (4,12), as well as the host hormones Epi and norepinephrine (4). Norepinephrine is produced in the intestine by adrenergic neurons of the enteric nervous system, whereas Epi is a systemic hormone produced in the central nervous system and the adrenal medulla that reaches the intestine through the bloodstream (13,14).…”