The Agrobacterium fabrum efflux pump PecM serves as a critical metabolic relief valve to maintain cellular...

Agrobacterium fabrum infects a wide range of plants, resulting in significant economic losses. In the rhizosphere, the bacterium is exposed to several stressors, including compounds secreted from other bacteria, fungi, or plants. A. fabrum encodes the efflux pump PecM for which the substrate is 4-hydroxybenzaldehyde, a compound that is both present in plant exudates and generated by bacteria during degradation of aromatic compounds. As an aromatic aldehyde, 4-hydroxybenzaldehyde is a chemically reactive electrophile that can react with cellular nucleophiles such as thiol groups on proteins and glutathione, leading to disruption of redox homeostasis. To assess the physiological consequences of pecM disruption, we compared the transcriptome and proteome profiles of wild-type and pecM disruption strains. While 30 genes were seen to be differentially expressed in the pecM disruption strain, 118 proteins differentially accumulated. Accumulating proteins included glutathione S-transferase and glutathione-dependent peroxiredoxin, both of which protect from oxidative stress. Consistent with this observation, the pecM disrupted strain was more resistant to oxidative stress. Across the genes and proteins detected in both data sets, there was no overall linear relationship, as reflected in a Pearson correlation of ~0. A marked depletion of bS21C, which is a component of the small ribosomal subunit, was observed; as one of three paralogs, its differential expression suggests the possibility that ribosome heterogeneity is part of adaptation to stress. Taken together, our data suggest that absence of PecM imposes a 4-hydroxybenzaldehyde-induced oxidative or electrophile stress and that PecM serves as a critical metabolic relief valve to maintain cellular homeostasis.

Efflux pumps often receive attention in the context of protecting bacteria from environmental toxins such as antibiotics. However, they are also important for exporting metabolic intermediates that may otherwise disrupt cellular processes. Bacterial metabolism of aromatic compounds can generate 4-hydroxybenzaldehyde, which the bacteria may also encounter in the rhizosphere, and its intracellular accumulation would be expected to disrupt redox homeostasis. Here, we show that disruption of the Agrobacterium fabrum efflux pump PecM, which exports 4-hydroxybenzaldehyde, results in widespread proteomic remodeling, with marked accumulation of proteins associated with oxidative and electrophile stress. These findings emphasize the importance of efflux systems in preventing the accumulation of reactive intermediates, and they expand our understanding of how such efflux systems contribute to metabolic balance and global cellular adaptation in complex environmental niches such as the rhizosphere.