Abstract
BackgroundBacterial communication is involved in regulation of cellular mechanisms such as metabolic processes, microbe-host interactions or biofilm formation. In the nitrogen-fixing model endophyte of grasses Azoarcus sp. strain BH72, known cell-cell signaling systems have not been identified; however, the pilA gene encoding the structural protein of type IV pili that are essential for plant colonization appears to be regulated in a population density-dependent manner.Methodology/Principal FindingsOur data suggest that pilAB expression is affected by population density, independent of autoinducers typical for Gram-negative bacteria, likely depending on unknown secreted molecule(s) that can be produced by different bacterial species. We used transcriptomic and proteomic approaches to identify target genes and proteins differentially regulated in conditioned supernatants in comparison to standard growth conditions. Around 8% of the 3992 protein-coding genes of Azoarcus sp. and 18% of the detected proteins were differentially regulated. Regulatory proteins and transcription factors among the regulated proteins indicated a complex hierarchy. Differentially regulated genes and proteins were involved in processes such as type IV pili formation and regulation, metal and nutrient transport, energy metabolism, and unknown functions mediated by hypothetical proteins. Four of the newly discovered target genes were further analyzed and in general they showed regulation patterns similar to pilAB. The expression of one of them was shown to be induced in plant roots.Conclusion/SignificanceThis study is the first global approach to initiate characterization of cell density-dependent gene regulation mediated by soluble molecule(s) in the model endophyte Azoarcus sp. strain BH72. Our data suggest that the putative signaling molecule(s) are also produced by other Proteobacteria and might thus be used for interspecies communication. This study provides the foundation for the development of robust reporter systems for Azoarcus sp. to analyze mechanisms and molecules involved in the population-dependent gene expression in this endophyte in future.
Highlights
Quorum sensing (QS) is a regulatory mechanism operating in response to cell density
Gram-negative bacteria widely use N-acyl-homoserine lactones (AHL) as quorum sensing signal molecules, whereas Gram-positives communicate via peptide-based quorum sensing systems
Similar to quorum sensingdependent mechanisms, expression was elevated upon incubation in conditioned cell-free supernatant, albeit only in a pilS deletion mutant
Summary
Quorum sensing (QS) is a regulatory mechanism operating in response to cell density This cell-to-cell communication system is widespread in bacteria and involves the production and detection of quorum sensing signal molecules, termed autoinducers, followed by transcriptional gene regulation [1,2]. In this way bacteria are capable to communicate with each other and partners in their environment via diverse signal molecules to allow concerted activities of a community. Plant-microbe interactions in several plant pathogens (Erwinia carotovora, Burkholderia pseudomallei), plant-associated bacteria (Pseudomonas putida) or symbionts (Sinorhizobium meliloti) are regulated in a cell density-dependent manner [9,10,11,12]. In the nitrogen-fixing model endophyte of grasses Azoarcus sp. strain BH72, known cell-cell signaling systems have not been identified; the pilA gene encoding the structural protein of type IV pili that are essential for plant colonization appears to be regulated in a population density-dependent manner
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