Autoinducer-2 functions as both a quorum sensing and metabolic signal in Escherichia coli
Bacteria integrate diverse environmental signals to coordinate behavior, yet the relationship between nutrient sensing and quorum sensing (QS) remains incompletely understood. Autoinducer-2 (AI-2) is unique among QS signals in that its production is tightly linked to central metabolism, raising fundamental questions about the boundary between metabolic and signaling functions. In Escherichia coli, AI-2 coordinates collective behaviors through the lsr operon, whose expression is controlled not only by the AI-2-responsive repressor LsrR but also by the cAMP receptor protein (CRP), placing it at the intersection of carbon sensing and population-level signaling. While inhibition of lsr operon expression by PTS sugars such as glucose was previously established, we demonstrate that non-PTS sugars similarly suppress lsr expression through CRP, further decoupling QS activation from cell density and coupling it to carbon source availability. Systematic analysis of Enterobacteriaceae genomes reveals that CRP binding sites in the lsr promoter region are broadly conserved, indicating that metabolic modulation of AI-2 signaling is an ancestral regulatory feature. Importantly, using a FRET-based biosensor, we show that AI-2 uptake modulates intracellular cAMP levels in a manner resembling non-PTS carbon source transport, suggesting that AI-2 may have originally functioned as a nutrient substrate, with its signaling role emerging subsequently or co-evolving alongside. In support of this hypothesis, we isolated soil- and phyllosphere-associated bacteria capable of utilizing AI-2 as a sole carbon source. Our findings reveal an underappreciated metabolic dimension of AI-2 QS and suggest an evolutionary trajectory in which AI-2 signaling emerged from ancestral carbon utilization pathways. ImportanceQS allows bacteria to coordinate collective behaviors by detecting secreted signaling molecules, yet the evolutionary origins of these systems remain poorly understood. AI-2, one of the most broadly conserved bacterial signals, is derived from central metabolism and processed by machinery in E. coli that strikingly resembles a sugar utilization system. Here, we show that nutrient availability overrides cell density as the primary determinant of AI-2 responsiveness, that this regulatory logic is conserved among Enterobacteriaceae genomes, and that environmental bacteria can grow on AI-2 as a sole carbon source. These findings reframe AI-2 as a signal embedded within, and potentially evolved from, nutrient sensing pathways, with direct implications for understanding how byproducts of cellular metabolism can acquire signaling functions.