Agrobacterium uses a unique ligand-binding mode for trapping opines and acquiring a competitive advantage in the niche construction on plant host

By modifying the nuclear genome of its host, the plant pathogen Agrobacterium tumefaciens induces the development of plant tumours in which it proliferates. The transformed plant tissues accumulate uncommon low molecular weight compounds called opines that are growth substrates for A. tumefaciens. In the pathogen-induced niche (the plant tumour), a selective advantage conferred by opine assimilation has been hypothesized, but not experimentally demonstrated. Here, using genetics and structural biology, we deciphered how the pathogen is able to bind opines and use them to efficiently compete in the plant tumour. We report high resolution X-ray structures of the periplasmic binding protein (PBP) NocT unliganded and liganded with the opine nopaline (a condensation product of arginine and α-ketoglurate) and its lactam derivative pyronopaline. NocT exhibited an affinity for pyronopaline (KD of 0.6 µM) greater than that for nopaline (KD of 3.7 µM). Although the binding-mode of the arginine part of nopaline/pyronopaline in NocT resembled that of arginine in other PBPs, affinity measurement by two different techniques showed that NocT did not bind arginine. In contrast, NocT presented specific residues such as M117 to stabilize the bound opines. NocT relatives that exhibit the nopaline/pyronopaline-binding mode were only found in genomes of the genus Agrobacterium. Transcriptomics and reverse genetics revealed that A. tumefaciens uses the same pathway for assimilating nopaline and pyronopaline. Fitness measurements showed that NocT is required for a competitive colonization of the plant tumour by A. tumefaciens. Moreover, even though the Ti-plasmid conjugal transfer was not regulated by nopaline, the competitive advantage gained by the nopaline-assimilating Ti-plasmid donors led to a preferential horizontal propagation of this Ti-plasmid amongst the agrobacteria colonizing the plant-tumour niche. This work provided structural and genetic evidences to support the niche construction paradigm in bacterial pathogens.
Author Summary An ecological niche is defined, in a given environment, by the availability of nutritive resources, which can be specifically assimilated by certain living organisms to promote their proliferation. The bacterial pathogen Agrobacterium tumefaciens is able to engineer an ecological niche in the infected host via the transformation of the plant genome and diversion of the plant metabolism towards production of the opine nutrients. In this work, we quantified the selective advantage conferred to a member of the phytopathogenic species A. tumefaciens which is able to assimilate the opine nopaline. This opine is a condensate of arginine and α-ketoglurate that is produced both under linear and cyclic forms in the plant tumour environment. We further determined at the molecular and atomistic levels how A. tumefaciens is able to sense the nopaline molecules, and which metabolic pathways are activated in response. Overall, this work deciphered some key molecular events in the niche construction of the pathogen A. tumefaciens that is unique among living organisms and used to develop bioengineering tools.