Abstract
In rhizobium strains, the lipid A is modified by the addition of a very long-chain fatty acid (VLCFA) shown to play an important role in rigidification of the outer membrane, thereby facilitating their dual life cycle, outside and inside the plant. In Bradyrhizobium strains, the lipid A is more complex with the presence of at least two VLCFAs, one covalently linked to a hopanoid molecule, but the importance of these modifications is not well-understood. In this study, we identified a cluster of VLCFA genes in the photosynthetic Bradyrhizobium strain ORS278, which nodulates Aeschynomene plants in a Nod factor-independent process. We tried to mutate the different genes of the VLCFA gene cluster to prevent the synthesis of the VLCFAs, but only one mutant in the lpxXL gene encoding an acyltransferase was obtained. Structural analysis of the lipid A showed that LpxXL is involved in the transfer of the C26:25OH VLCFA to the lipid A but not in the one of the C30:29OH VLCFA which harbors the hopanoid molecule. Despite maintaining the second VLCFA, the ability of the mutant to cope with various stresses (low pH, high temperature, high osmolarity, and antimicrobial peptides) and to establish an efficient nitrogen-fixing symbiosis was drastically reduced. In parallel, we investigated whether the BRADO0045 gene, which encodes a putative acyltransferase displaying a weak identity with the apo-lipoprotein N-acyltransferase Lnt, could be involved in the transfer of the C30:29OH VLCFA to the lipid A. Although the mutant exhibited phenotypes similar to the lpxXL mutant, no difference in the lipid A structure was observed from that in the wild-type strain, indicating that this gene is not involved in the modification of lipid A. Our results advance our knowledge of the biosynthesis pathway and the role of VLCFAs-modified lipid A in free-living and symbiotic states of Bradyrhizobium strains.
Highlights
Rhizobia are Gram-negative bacteria with two life styles, one in a free-living state in the soil where they have to cope with changing environmental conditions and the other in symbiosis with plants, inside an organ called a nodule, in which they reduce atmospheric nitrogen to ammonium for the benefit of the host plant
A BLAST search of the ORS278 genome led to the identification of a single gene cluster containing several homologs of genes shown to be involved in very long-chain fatty acid (VLCFA) biosynthesis in the R. leguminosarum bv. viciae 3841 strain (Figure 2) (Bourassa et al, 2017)
It was shown that the lipid A of several Bradyrhizobium strains differs in the presence of at least two VLCFAs, one of which can be linked to a hopanoid molecule (Choma and Komaniecka, 2011; Komaniecka et al, 2014; Silipo et al, 2014)
Summary
Rhizobia are Gram-negative bacteria with two life styles, one in a free-living state in the soil where they have to cope with changing environmental conditions (hydric, acid, saline stresses, nutrient starvation, etc.) and the other in symbiosis with plants, inside an organ called a nodule, in which they reduce atmospheric nitrogen to ammonium for the benefit of the host plant. The biosynthesis of the VLCFA and its addition to the lipid A require a cluster of five genes found in all the bacteria that synthesize a VLCFA-modified lipid A (Ardissone et al, 2011) This region is composed of genes encoding an acyl carrier protein (acpXL), fatty acid elongation proteins (fabF1XL and fabF2XL), a beta-hydroxyacyl-acyl carrier protein (ACP) dehydratase (fabZXL) and an acyltransferase (lpxXL) (Ardissone et al, 2011; Brown et al, 2011)
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