The Type III Effectome of the Symbiotic Bradyrhizobium vignae Strain ORS3257

Abstract

Many Bradyrhizobium strains are able to establish a Nod factor-independent symbiosis with the leguminous plant Aeschynomene indica by the use of a type III secretion system (T3SS). Recently, an important advance in the understanding of the molecular factors supporting this symbiosis has been achieved by the in silico identification and functional characterization of 27 putative T3SS effectors (T3Es) of Bradyrhizobium vignae ORS3257. In the present study, we experimentally extend this catalog of T3Es by using a multi-omics approach. Transcriptome analysis under non-inducing and inducing conditions in the ORS3257 wild-type strain and the ttsI mutant revealed that the expression of 18 out of the 27 putative effectors previously identified, is under the control of TtsI, the global transcriptional regulator of T3SS and T3Es. Quantitative shotgun proteome analysis of culture supernatant in the wild type and T3SS mutant strains confirmed that 15 of the previously determined candidate T3Es are secreted by the T3SS. Moreover, the combined approaches identified nine additional putative T3Es and one of them was experimentally validated as a novel effector. Our study underscores the power of combined proteome and transcriptome analyses to complement in silico predictions and produce nearly complete effector catalogs. The establishment of the ORS3257 effectome will form the basis for a full appraisal of the symbiotic properties of this strain during its interaction with various host legumes via different processes.

Widespread Bradyrhizobium distribution of diverse Type III effectors that trigger legume nodulation in the absence of Nod factor

“…This indicates that this T3E fulfills its role by deSUMOylation of plant protein targets. Taken together, these data indicate that different classes of rhizobial T3Es able to trigger nodulation and proposed to be called ET-Nods [ 15 ], use different mechanisms to activate the nodulation program in legumes.…”

Signaling in Legume–Rhizobia Symbiosis

“…For example, Sinorhizobium fredii is known to secrete eight Nop proteins (NopA-D, NopL, NopM, NopP, and NopX), which are the extracellular components of the type III secretory system [105][106][107], via T3SS in response to stimulation with genistein secreted by plant roots [108]. In total, about 30 different Nop proteins were discovered over the last two decades [109]. The most of them were predicted in silico based on the results of transcriptomics analysis, whereas only a few could be characterized experimentally at the level of functional proteins.…”

From prediction to function: Current practices and challenges towards the functional characterization of type III effectors

“…Here, the transcriptome of a wild-type strain was compared to a T3E-transcriptional regulator mutant sampled under either T3E-inducing or noninducing conditions whereafter the supernatant culture of these strains was compared by shotgun proteomics. Combined with an in silico approach, 36 putative T3Es were identified in at least one of the approaches ( Busset et al, 2021 ). This method could also work for the RSSC by comparison of the transcriptome and proteome of the wild type and a transcriptional regulator ( hrpB ) mutant strain.…”