Indole inhibits bacterial quorum sensing signal transmission by interfering with quorum sensing regulator folding

Quorum sensing (QS)-dependent biofilm formation and motility were controlled by AqsR in Acinetobacter oleivorans DR1. QS-controlled phenotypes appeared to be inhibited by indole and the aqsR mutant had the same phenotypes. We demonstrated that the turnover rate of AqsR became more rapid without the N-acylhomoserine lactone (AHL) signal, and that indole could increase the expression of many protease and chaperone proteins. The addition of exogenous indole decreased the expression of two AqsR-targeted genes: AOLE_03905 (putative surface adhesion protein) and AOLE_11355 (L-asparaginase). The overexpression of AqsR in Escherichia coli was impossible with the indole treatment. Surprisingly, our [³⁵S]methionine pulse-labelling data demonstrated that the stability and folding of AqsR protein decreased in the presence of indole without changing aqsR mRNA expression in E. coli. Interestingly, indole resulted in a loss of TraR-dependent traG expression in an Agrobacterium tumefaciens indicator strain. However, when indole was added after incubation with exogenous AHL, indole could not inhibit the TraR-dependent expression of the traG promoter. This indicated that AHL-bound TraR could be protective against indole, but TraR without AHL could not be active in the presence of indole. Here, we provided evidence for the first time showing that the indole effect on QScontrolled bacterial phenotypes is due to inhibited QS regulator folding and not a reduced QS signal.