Proteomic Analysis of the Soybean Symbiosome Identifies New Symbiotic Proteins*

Abstract

Legumes form a symbiosis with rhizobia in which the plant provides an energy source to the rhizobia bacteria that it uses to fix atmospheric nitrogen. This nitrogen is provided to the legume plant, allowing it to grow without the addition of nitrogen fertilizer. As part of the symbiosis, the bacteria in the infected cells of a new root organ, the nodule, are surrounded by a plant-derived membrane, the symbiosome membrane, which becomes the interface between the symbionts. Fractions containing the symbiosome membrane (SM) and material from the lumen of the symbiosome (peribacteroid space or PBS) were isolated from soybean root nodules and analyzed using nongel proteomic techniques. Bicarbonate stripping and chloroform-methanol extraction of isolated SM were used to reduce complexity of the samples and enrich for hydrophobic integral membrane proteins. One hundred and ninety-seven proteins were identified as components of the SM, with an additional fifteen proteins identified from peripheral membrane and PBS protein fractions. Proteins involved in a range of cellular processes such as metabolism, protein folding and degradation, membrane trafficking, and solute transport were identified. These included a number of proteins previously localized to the SM, such as aquaglyceroporin nodulin 26, sulfate transporters, remorin, and Rab7 homologs. Among the proteome were a number of putative transporters for compounds such as sulfate, calcium, hydrogen ions, peptide/ dicarboxylate, and nitrate, as well as transporters for which the substrate is not easy to predict. Analysis of the promoter activity for six genes encoding putative SM proteins showed nodule specific expression, with five showing expression only in infected cells. Localization of two proteins was confirmed using GFP-fusion experiments. The data have been deposited to the ProteomeXchange with identifier PXD001132. This proteome will provide a rich resource for the study of the legumerhizobium symbiosis. Molecular & Cellular Proteomics

The plasma membrane proteome of Medicago truncatula roots as modified by arbuscular mycorrhizal symbiosis

“…Among them, substantiating similarities in the development of both fungal and bacterial symbioses (Bapaume and Reinhardt 2012), the AM-induced lysine/ histidine transporter, MtAnn1 and flotillin-like 4 (FLOT4) also play role in rhizobial symbiosis (Vincill et al 2005;Breakspear et al 2014). In support of the importance of membrane microdomain-associated proteins in symbiotic relationships, the lysine/histidine transporter and FLOT4 have a continuing presence on the symbiosome membrane (Vincill et al 2005;Clarke et al 2015), and plants silenced for the DRMassociated FLOT4 protein form a reduced number of nodules that do not fix nitrogen efficiently (Haney and Long 2010).…”

Loss‐of‐function of ASPARTIC PEPTIDASE NODULE‐INDUCED 1 (APN1) in Lotus japonicus restricts efficient nitrogen‐fixing symbiosis with specific Mesorhizobium loti strains

“…Similar to AtCDR1, APN1 has a 24-amino acid signal peptide at the N-terminus ( Figure S3), which is predicted to be extracellular-targeting signal. The probable orthologue of Lotus APN1 in soybean (Glycine max), Glyma15 g260700 (Glyma15 g41420.1 in SoyBase v1.0; Figure 3b), is shown to locate in the peribacteroid space by proteomics analysis (Clarke et al, 2015). In addition, the putative APN1 orthologue in M. truncatula, XP013443143 (Figure 3b), is abundant in the proximal infection zone and infected cells but not in uninfected cells (https://mtgea.noble.org/v3/).…”

MtSWEET11, a Nodule-Specific Sucrose Transporter of Medicago truncatula

“…Movement of Suc between compartments of individual cells also requires transporters in the intervening membranes. Many putative Suc transporter genes have been identified in legumes and are expressed in nodules (Wienkoop and Saalbach, 2003;Benedito et al, 2010;Limpens et al, 2013;Clarke et al, 2015), although none of these have been characterized genetically. In this study, we characterized a nodule-specific Suc transporter of M. truncatula, MtSWEET11 at the molecular, biochemical, cellular, and genetic levels.…”