Perception of microbe-associated molecular patterns (MAMPs) through pattern recognition receptors (PRRs) triggers various defense responses in plants. This MAMP-triggered immunity plays a major role in the plant resistance against various pathogens. To clarify the molecular basis of the specific recognition of chitin oligosaccharides by the rice PRR, CEBiP (chitin-elicitor binding protein), as well as the formation and activation of the receptor complex, biochemical, NMR spectroscopic, and computational studies were performed. Deletion and domain-swapping experiments showed that the central lysine motif in the ectodomain of CEBiP is essential for the binding of chitin oligosaccharides. Epitope mapping by NMR spectroscopy indicated the preferential binding of longer-chain chitin oligosaccharides, such as heptamer-octamer, to CEBiP, and also the importance of N-acetyl groups for the binding. Molecular modeling/docking studies clarified the molecular interaction between CEBiP and chitin oligosaccharides and indicated the importance of Ile 122 in the central lysine motif region for ligand binding, a notion supported by site-directed mutagenesis. Based on these results, it was indicated that two CEBiP molecules simultaneously bind to one chitin oligosaccharide from the opposite side, resulting in the dimerization of CEBiP. The model was further supported by the observations that the addition of (GlcNAc) 8 induced dimerization of the ectodomain of CEBiP in vitro, and the dimerization and (GlcNAc) 8 -induced reactive oxygen generation were also inhibited by a unique oligosaccharide, (GlcNβ1,4GlcNAc) 4 , which is supposed to have N-acetyl groups only on one side of the molecule. Based on these observations, we proposed a hypothetical model for the ligand-induced activation of a receptor complex, involving both CEBiP and Oryza sativa chitinelicitor receptor kinase-1.plant immunity | MTI/PTI | chitin signaling | receptor-ligand interaction |

Lipid research represents a frontier for microbiology, as showcased by hopanoid lipids. Hopanoids, which resemble sterols and are found in the membranes of diverse bacteria, have left an extensive molecular fossil record. They were first discovered by petroleum geologists. Today, hopanoid-producing bacteria remain abundant in various ecosystems, such as the rhizosphere. Recently, great progress has been made in our understanding of hopanoid biosynthesis, facilitated in part by technical advances in lipid identification and quantification. A variety of genetically tractable, hopanoid-producing bacteria have been cultured, and tools to manipulate hopanoid biosynthesis and detect hopanoids are improving. However, we still have much to learn regarding how hopanoid production is regulated, how hopanoids act biophysically and biochemically, and how their production affects bacterial interactions with other organisms, such as plants. The study of hopanoids thus offers rich opportunities for discovery.

A better understanding of how bacteria resist stresses encountered during the progression of plant-microbe symbioses will advance our ability to stimulate plant growth. Here, we show that the symbiotic system comprising the nitrogen-fixing bacterium Bradyrhizobium diazoefficiens and the legume Aeschynomene afraspera requires hopanoid production for optimal fitness. While methylated (2Me) hopanoids contribute to growth under plant-cell-like microaerobic and acidic conditions in the free-living state, they are dispensable during symbiosis. In contrast, synthesis of extended (C35) hopanoids is required for growth microaerobically and under various stress conditions (high temperature, low pH, high osmolarity, bile salts, oxidative stress, and antimicrobial peptides) in the free-living state and also during symbiosis. These defects might be due to a less rigid membrane resulting from the absence of free or lipidA-bound C35 hopanoids or the accumulation of the C30 hopanoid diploptene. Our results also show that C35 hopanoids are necessary for symbiosis only with the host Aeschynomene afraspera but not with soybean. This difference is likely related to the presence of cysteine-rich antimicrobial peptides in Aeschynomene nodules that induce drastic modification in bacterial morphology and physiology. The study of hopanoid mutants in plant symbionts thus provides an opportunity to gain insight into host-microbe interactions during later stages of symbiotic progression, as well as the microenvironmental conditions for which hopanoids provide a fitness advantage.

No abstract

The immediate call for translational research in the field of coronavirus disease (COVID-19) pandemic, needs new and unexplored angles to support and contribute to this important worldwide health problem. The aim of this study is to better understand the pathogenic mechanisms underlying COVID-19, deciphering the carbohydrate-mediated interactions of the SARS-CoV-2 spike protein. We studied the carbohydrate-binding receptors that could be important for viral entry and for immune-modulatory responses, and we studied the interactions of the spike protein with the host lung microbiota. Exploring solid-phase immunoassays, we evaluated the interactions between the SARS-CoV-2 spike protein and a library of 12 different human carbohydrate-binding proteins (C-type lectins and Siglecs) involved in binding, triggering and modulation of innate and adaptive immune-responses. We revealed a specific binding of the SARS-CoV-2 spike protein to the receptors DC-SIGN, MGL, Siglec-9 and Siglec-10 that are all expressed on myeloid immune cells. In addition, because the lung microbiota can promote or modulate viral infection, we studied the interactions between the SARS-CoV-2 spike protein and a library of Streptococcus pneumoniae capsular polysaccharides, as well as other bacterial glyco-conjugates. We show specific binding of the spike protein to different S. pneumoniae capsular polysaccharides (serotypes 19F and 23F but not to serotype 14). Moreover we demonstrated a specific binding of SARS-CoV-2 spike protein to the lipopolysaccharide from the opportunistic human pathogen Pseudomonas aeruginosa, one of the leading cause of acute nosocomial infections and pneumonia. Interestingly, we identified rhamnosylated epitopes as one of the discriminating structures in lung microbiota to bind SARS-CoV-2 spike protein. In conclusion, we revealed novel ACE2-independent carbohydrate-mediated interactions with immune modulating lectins expressed on myeloid cells, as well as host lung microbiota glycoconjugates. Our results identified new molecular pathways using host lectins and signalling, that may contribute to viral infection and subsequent immune exacerbation. Moreover we identified specific rhamnosylated epitopes in lung microbiota to bind SARS-CoV-2, providing a hypothetical link between the presence of specific lung microbiota and SARS-CoV-2 infection and severity.

Generated by gram-negative bacteria, lipopolysaccharides (LPSs) are one of the most abundant and potent immunomodulatory substances present in the intestinal lumen. Interaction of agonistic LPS with the host myeloid-differentiation-2/Tolllike receptor 4 (MD-2/TLR4) receptor complex results in nuclear factor kB (NF-kB) activation, followed by the robust induction of pro-inflammatory immune responses. Here we have isolated LPS from a common gut commensal, Bacteroides vulgatus mpk (BVMPK), which provides only weak agonistic activity. This weak agonistic activity leads to the amelioration of inflammatory immune responses in a mouse model for experimental colitis, and it was in sharp contrast to strong agonists and antagonists. In this context, the administration of BVMPK LPS into mice with severe intestinal inflammation re-established intestinal immune homeostasis within only 2 weeks, resulting in the clearance of all symptoms of inflammation. These inflammation-reducing properties of weak agonistic LPS are grounded in the induction of a special type of endotoxin tolerance via the MD-2/TLR4 receptor complex axis in intestinal lamina propria CD11c + cells. Thus, weak agonistic LPS represents a promising agent to treat diseases involving pathological overactivation of the intestinal immune system, e.g., in inflammatory bowel diseases.

The gut microbiota guide the development of the host immune system by setting a systemic threshold for immune activation. Lipopolysaccharides (LPSs) from gut bacteria are able to trigger systemic and local proinflammatory and immunomodulatory responses, and this capability strongly relies on their fine structures. Up to now, only a few LPS structures from gut commensals have been elucidated; therefore, the molecular motifs that may be important for LPS–mammalian cell interactions at the gut level are still obscure. Here, we report on the full structure of the LPS isolated from one of the prominent species of the genus Bacteroides , Bacteroides vulgatus . The LPS turned out to consist of a particular chemical structure based on hypoacylated and mono -phosphorylated lipid A and with a galactofuranose-containing core oligosaccharide and an O-antigen built up of mannose and rhamnose. The evaluation of the immunological properties of this LPS on human in vitro models revealed a very interesting capability to produce anti-inflammatory cytokines and to induce a synergistic action of MD-2/TLR4- and TLR2-mediated signaling pathways.

Transport of lipopolysaccharides (LPS) to the surface of the outer membrane is essential for viability of Gram-negative bacteria. Periplasmic LptC and LptA proteins of the LPS transport system (Lpt) are responsible for LPS transfer between the Lpt inner and outer membrane complexes. Here, using a monomeric E. coli LptA mutant, we first show in vivo that a stable LptA oligomeric form is not strictly essential for bacteria. The LptC-LptA complex was characterized by a combination of SAXS and NMR methods and a low resolution model of the complex was determined. We were then able to observe interaction of LPS with LptC, the monomeric LptA mutant as well as with the LptC-LptA complex. A LptC-LPS complex was built based on NMR data in which the lipid moiety of the LPS is buried at the interface of the two β-jellyrolls of the LptC dimer. The selectivity of LPS for this intermolecular surface and the observation of such cavities at homo- or heteromolecular interfaces in LptC and LptA suggests that intermolecular sites are essential for binding LPS during its transport.