Peribacteroid space acidification: a marker of mature bacteroid functioning in Medicago truncatula nodules

Abstract

Legumes form a symbiotic interaction with Rhizobiaceae bacteria, which differentiate into nitrogen-fixing bacteroids within nodules. Here, we investigated in vivo the pH of the peribacteroid space (PBS) surrounding the bacteroid and pH variation throughout symbiosis. In vivo confocal microscopy investigations, using acidotropic probes, demonstrated the acidic state of the PBS. In planta analysis of nodule senescence induced by distinct biological processes drastically increased PBS pH in the N2-fixing zone (zone III). Therefore, the PBS acidification observed in mature bacteroids can be considered as a marker of bacteroid N2 fixation. Using a pH-sensitive ratiometric probe, PBS pH was measured in vivo during the whole symbiotic process. We showed a progressive acidification of the PBS from the bacteroid release up to the onset of N2 fixation. Genetic and pharmacological approaches were conducted and led to disruption of the PBS acidification. Altogether, our findings shed light on the role of PBS pH of mature bacteroids in nodule functioning, providing new tools to monitor in vivo bacteroid physiology.

Stringent Expression Control of Pathogenic R-body Production in Legume Symbiont Azorhizobium caulinodans

“…Subsequently, internalized bacteria enter acidified food vacuoles, and R bodies are unrolled and penetrate the phagosomal membrane to deliver lethal toxins to the cytoplasm (9, 21). This scenario may also be applicable to interactions of A. caulinodans and S. rostrata , in which the peribacteroid space (microenvironment surrounding bacteroids) is progressively acidified during nodule morphogenesis (22), likely triggering the conformational change of R bodies into the needle-shaped structure that penetrates membranes.…”

Hopanoid lipids: from membranes to plant–bacteria interactions

“…The environment of the PBS is harsh and drastically different from that of free-living rhizobia. Some of this harshness is necessary for efficient nitrogen fixation, including the creation of a low pH environment to provide protons for nitrogenase 122 and concentrated carbon osmolytes for increased bacteroid metabolism 123 . Other stresses result from host control mechanisms, such as antimicrobial nodule-specific cysteine-rich peptides that change the morphology, proliferation and gene expression of bacteroids to maximize their productivity 124–126 .…”

Involvement of papain and legumain proteinase in the senescence process of Medicago truncatula nodules

“…The nitrogen-fixing capacity of bacteroids was determined in vivo by measuring acetylene-reducing activity (ARA; Pierre et al, 2013). Nodulated roots were harvested and incubated at 30°C for 1 h in rubber-capped tubes containing a 10% (v/v) acetylene atmosphere.…”