Rhizobial NodL O-Acetyl Transferase and NodS N-Methyl Transferase Functionally Interfere in Production of Modified Nod Factors

ABSTRACT The products of the rhizobial nodulation genes are involved in the biosynthesis of lipochitin oligosaccharides (LCOs), which are host-specific signal molecules required for nodule formation. The presence of an O-acetyl group on C-6 of the nonreducingN-acetylglucosamine residue of LCOs is due to the enzymatic activity of NodL. Here we show that transfer of the nodLgene into four rhizobial species that all normally produce LCOs that are not modified on C-6 of the nonreducing terminal residue results in production of LCOs, the majority of which have an acetyl residue substituted on C-6. Surprisingly, in transconjugant strains ofMesorhizobium loti, Rhizobium etli, and Rhizobium tropici carrying nodL, such acetylation of LCOs prevents the endogenous nodS-dependent transfer of theN-methyl group that is found as a substituent of the acylated nitrogen atom. To study this interference betweennodL and nodS, we have cloned thenodS gene of M. loti and used its product in in vitro experiments in combination with purified NodL protein. It has previously been shown that a chitooligosaccharide N deacetylated on the nonreducing terminus (the so-called NodBC metabolite) is the preferred substrate for NodS as well as for NodL. Here we show that the NodBC metabolite, acetylated by NodL, is not used by the NodS protein as a substrate while the NodL protein can acetylate the NodBC metabolite that has been methylated by NodS.

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Structure, functions and perspectives of practical application of the signal molecules inducing development of rhizobia-legume symbiosis

Ecological genetics ◽

10.17816/ecogen2314-24 ◽

2004 ◽

Vol 2 (3) ◽

pp. 14-24

Author(s):

Aleksandra O Ovtsyna ◽

Igor A Tikhonovich

Keyword(s):

Soil Bacteria ◽

Mutual Recognition ◽

Structural Diversity ◽

Nodule Formation ◽

Signal Molecules ◽

Practical Application ◽

Nod Factors ◽

Legume Symbiosis ◽

Molecular Signals

Soil bacteria rhizobia establish nitrogen-fixing symbiosis with legume plants. Mutual recognition of symbiotic partners and initiation of nodule formation occur via exchange by molecular signals secreted both by plant and bacteria. This review summarizes recent data about structural diversity, genetic control of biosynthesis and functional role of Nod-factors. The possibilities of practical application of flavonoids and Nod-factors in agriculture are discussed

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N-Deacetylation of Sinorhizobium meliloti Nod Factors Increases Their Stability in the Medicago sativa Rhizosphere and Decreases Their Biological Activity

Molecular Plant-Microbe Interactions ◽

10.1094/mpmi.2000.13.1.72 ◽

2000 ◽

Vol 13 (1) ◽

pp. 72-79 ◽

Cited By ~ 20

Author(s):

Christian Staehelin ◽

Michael Schultze ◽

Ken Tokuyasu ◽

Véréna Poinsot ◽

Jean-Claude Promé ◽

...

Keyword(s):

Biological Activity ◽

Medicago Sativa ◽

Sinorhizobium Meliloti ◽

Colletotrichum Lindemuthianum ◽

Signal Molecules ◽

Glycosyl Hydrolases ◽

Nod Factors ◽

Nod Factor ◽

Chemical Structures

Nod factors excreted by rhizobia are signal molecules that consist of a chitin oligomer backbone linked with a fatty acid at the nonreducing end. Modifications of the Nod factor structures influence their stability in the rhizosphere and their biological activity. To test the function of N-acetyl groups in Nod factors, NodSm-IV(C16:2,S) from Sinorhizobium meliloti was enzymatically N-deacetylated in vitro with purified chitin deacetylase from Colletotrichum lindemuthianum. A family of partially and completely deacetylated derivatives was produced and purified. The most abundant chemical structures identified by mass spectrometry were GlcN(C16:2)-GlcNAc-GlcNH2-GlcNAc(OH)(S), GlcN(C16:2)-GlcNAc-GlcNH2-GlcNH2(OH)(S), and GlcN(C16:2)-GlcNH2-GlcNH2-GlcNH2(OH)(S). In contrast to NodSm-IV(C16:2,S), the purified N-deacetylated derivatives were stable in the rhizosphere of Medicago sativa, indicating that the N-acetyl groups make the carbohydrate moiety of Nod factors accessible for glycosyl hydrolases of the host plant. The N-deacetylated derivatives displayed only a low level of activity in inducing root hair deformation. Furthermore, the N-deacetylated molecules were not able to stimulate Nod factor degradation by M. sativa roots, a response elicited by active Nod factors. These data show that N-acetyl groups of Nod factors are required for biological activity.

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Effect of C6-Volatiles on Bioluminescent Plant Pathogens

HortScience ◽

10.21273/hortsci.33.3.557d ◽

1998 ◽

Vol 33 (3) ◽

pp. 557d-557

Author(s):

Jennifer Warr ◽

Fenny Dane ◽

Bob Ebel

Keyword(s):

Biological Activity ◽

Bacterial Growth ◽

Xanthomonas Campestris ◽

Plant Pathogens ◽

Genetically Engineered ◽

The Other ◽

Computer Assisted ◽

Signal Molecules ◽

Low Concentrations

C6 volatile compounds are known to be produced by the plant upon pathogen attack or other stress-related events. The biological activity of many of these substances is poorly understood, but some might produce signal molecules important in host–pathogen interactions. In this research we explored the possibility that lipid-derived C6 volatiles have a direct effect on bacterial plant pathogens. To this purpose we used a unique tool, a bacterium genetically engineered to bioluminesce. Light-producing genes from a fish-associated bacterium were introduced into Xanthomonas campestris pv. campestris, enabling nondestructive detection of bacteria in vitro and in the plant with special computer-assisted camera equipment. The effects of different C6 volatiles (trans-2 hexanal, trans-2 hexen-1-ol and cis-3 hexenol) on growth of bioluminescent Xanthomonas campestris were investigated. Different volatile concentrations were used. Treatment with trans-2 hexanal appeared bactericidal at low concentrations (1% and 10%), while treatments with the other volatiles were not inhibitive to bacterial growth. The implications of these results with respect to practical use of trans-2 hexanal in pathogen susceptible and resistant plants will be discussed.

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Cyclosporine A Suppresses the Malignant Progression of Oral Squamous Cell Carcinoma in vitro

Anti-Cancer Agents in Medicinal Chemistry ◽

10.2174/1871520618666181029170605 ◽

2019 ◽

Vol 19 (2) ◽

pp. 248-255 ◽

Cited By ~ 2

Author(s):

Ling Gao ◽

Jianwei Dong ◽

Nanyang Zhang ◽

Zhanxian Le ◽

Wenhao Ren ◽

...

Keyword(s):

Cell Cycle ◽

Squamous Cell Carcinoma ◽

Oral Squamous Cell Carcinoma ◽

Cell Carcinoma ◽

Squamous Cell ◽

Cyclosporine A ◽

Migration And Invasion ◽

Signal Molecules ◽

Cox 2

Background:The Oral Squamous Cell Carcinoma (OSCC) is one of the most frequent cancer types. Failure of treatment of OSCC is potentially lethal because of local recurrence, regional lymph node metastasis, and distant metastasis. Chemotherapy plays a vital role through suppression of tumorigenesis. Cyclosporine A (CsA), an immunosuppressant drug, has been efficiently used in allograft organ transplant recipients to prevent rejection, and also has been used in a subset of patients with autoimmunity related disorders. The present study aims to investigate novel and effective chemotherapeutic drugs to overcome drug-resistance in the treatment of OSCC.Methods:Cells were incubated in the standard way. Cell viability was assayed using the MTT assay. Cell proliferation was determined using colony formation assay. The cell cycle assay was performed using flow cytometry. Apoptosis was assessed using fluorescence-activated cell sorting after stained by the Annexin V-fluorescein isothiocyanate (FITC). Cell migration and invasion were analyzed using wound healing assay and tranwell. The effect of COX-2, c-Myc, MMP-9, MMP-2, and NFATc1 protein expression was determined using Western blot analysis while NFATc1 mRNA expression was determined by RT-PCR.Results:In vitro studies indicated that CsA inhibited partial OSCC growth by inducing cell cycle arrest, apoptosis, and the migration and invasion of OSCC cells. We also demonstrated that CsA could inhibit the expression of NFATc1 and its downstream genes COX-2, c-Myc, MMP-9, and MMP-2 in OSCC cells. Furthermore, we analyzed the expression of NFATc1 in head and neck cancer through the Oncomine database. The data was consistent with the experimental findings.Conclusion:The present study initially demonstrated that CsA could inhibit the progression of OSCC cells and can mediate the signal molecules of NFATc1 signaling pathway, which has strong relationship with cancer development. That explains us CsA has potential to explore the possibilities as a novel chemotherapeutic drug for the treatment of OSCC.

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Affinity chromatography of Ruta graveolens L. O-methyltransferases. Studies demonstrating the potential of the technique in the mechanistic investigation of O-methyltransferases

Canadian Journal of Biochemistry ◽

10.1139/o79-122 ◽

1979 ◽

Vol 57 (7) ◽

pp. 986-994 ◽

Cited By ~ 12

Author(s):

Satish K. Sharma ◽

Stewart A. Brown

Keyword(s):

Ferulic Acid ◽

Affinity Chromatography ◽

Caffeic Acid ◽

Kinetic Mechanism ◽

Ruta Graveolens ◽

Methyl Transferase ◽

Methyl Group ◽

Acid Ligand ◽

Mechanistic Investigation ◽

Sepharose 4B

Two discrete furanocoumarin (5- and 8-) O-methyltransferases and a caffeic acid 3-O-methyl-transferase from cell cultures of Ruta graveolens L. have been copurified by affinity chromatography on 1,6-diaminohexane agarose (AH-Sepharose 4B) linked with 5-adenosyl-L-homocysteine (SAH). The furanocoumarin O-methyltransferases, which transfer a methyl group from S-adenosyl-L-methionine (SAM) to the 5- or 8-hydroxyls of linear furanocoumarins, were not retarded by 5-(3-carboxypropanamido)-xanthotoxin (CPAX) immobilized to AH-Sepharose 4B, but addition of SAM to the irrigant buffer led to complete retardation of both enzymes on this affinity system. An analogous phenomenon was observed for the caffeic acid O-methyltransferase, with a ferulic acid ligand coupled to the same insoluble support. SAH was as effective as SAM in promoting binding of the furanocoumarin O-methyltransferases to CPAX and caffeic acid 3-O-methyltransferase to immobilized ferulic acid, respectively. The strong and specific adsorption of these enzymes was abolished by exclusion of SAM or SAH from the irrigant buffer. It is concluded that the enzymes bind first to SAM or SAH, and that this binding process in turn induces the binding site for their specific phenolic substrates or their analogs. Based on these findings, a compulsory–ordered kinetic mechanism for the action of these O-methyltransferases is postulated.

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In vitro growth and indoleacetic acid production by Mesorhizobium loti SEMIA806 and SEMIA816 under the influence of copper ions

Microbiology Research ◽

10.4081/mr.2017.7302 ◽

2017 ◽

Vol 8 (2) ◽

Cited By ~ 2

Author(s):

Jéssica Dutra Vieira ◽

Paulo Roberto Diniz Da Silva ◽

Valdir Marcos Stefenon

Keyword(s):

Contaminated Soils ◽

Acid Production ◽

Indoleacetic Acid ◽

Copper Ions ◽

Control Culture ◽

Control Treatment ◽

Symbiotic Interaction ◽

Mesorhizobium Loti ◽

Significant Difference

The indoleacetic acid produced by symbiotic bacteria is an important phytohormone signaling microbe-plant interaction, being therefore essential for rhizoremediation. In this study, the effect of different concentrations of copper ions on the bacterial growth and indoleacetic acid production was investigated in two strains of Mesorhizobium loti in in vitro conditions, aiming to determine critical concentrations of this heavy metal for rhizoremediation of contaminated soils using this bacterium. The experiment consisted on a control culture without copper and three treatments supplemented with 10 mg.L-1, 20 mg.L-1 or 50 mg.L-1 of CuSO4. For both strains, the growth stopped after 48h and no significant difference was observed across treatments. The production of indoleacetic acid by the control treatment without copper was significantly higher in comparison to the copper- containing treatments. Mesorhizobium loti SEMIA806 and SEMIA816 are resistant to up to 50 mg.L-1 of CuSO4 in the culture medium, presenting effective growth. The synthesis of indoleacetic acid was strongly reduced but not excluded by ions copper in the medium. So, it is expected that environmental copper found in the soil up to the concentration of 50 mg.L-1 will not preclude the symbiotic interaction between M. loti and leguminous host plant in rhizoremediation enterprises.

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The Effect of pH Stress on the Survival of Rhizobium Trifolii and Sinorhizobium Meliloti in Vitro

Bulletin of University of Agricultural Sciences and Veterinary Medicine Cluj-Napoca Agriculture ◽

10.15835/buasvmcn-agr:9811 ◽

2013 ◽

Vol 70 (2) ◽

pp. 449-450

Author(s):

Monica NISTE ◽

Roxana VIDICAN ◽

Ioan ROTAR ◽

Rodica POP

Keyword(s):

Sinorhizobium Meliloti ◽

Agar Medium ◽

Rhizobium Trifolii ◽

Ph Values ◽

Optimum Ph ◽

Yeast Extract Mannitol ◽

Rhizobial Species ◽

The Impact ◽

Different Soils

Nitrogen-fixing symbiotic bacteria known as rhizobia can exist in different soils and adapt to different environmental conditions. The aim of this study was to determine the impact of pH on the growth of Rhizobium trifolii and Sinorhizobium meliloti. Rhizobial species were isolated using yeast extract mannitol agar medium) in which the pH values were adjusted to 5.0, 6.0, 8.0 and 9.0 by adding HCl and NaOH. The optimum pH for rhizobia is neutral or slightly alkaline (pH 8) and they are more sensitive to acidity. Sinorhizobium meliloti developed better in an acid medium compared to Rhizobium trifolii.

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Therapeutic Effect of an Antibody-Derived Peptide in a Galleria mellonella Model of Systemic Candidiasis

International Journal of Molecular Sciences ◽

10.3390/ijms222010904 ◽

2021 ◽

Vol 22 (20) ◽

pp. 10904

Author(s):

Emerenziana Ottaviano ◽

Elisa Borghi ◽

Laura Giovati ◽

Monica Falleni ◽

Delfina Tosi ◽

...

Keyword(s):

Galleria Mellonella ◽

Fat Body ◽

Single Injection ◽

Host Immunity ◽

Nodule Formation ◽

Host Immune System ◽

Immune Stimulation ◽

Yeast Infection ◽

Pathogen Clearance

The synthetic peptide T11F (TCRVDHRGLTF), with sequence identical to a fragment of the constant region of human IgM, and most of its alanine-substituted derivatives proved to possess a significant candidacidal activity in vitro. In this study, the therapeutic efficacy of T11F, D5A, the derivative most active in vitro, and F11A, characterized by a different conformation, was investigated in Galleria mellonella larvae infected with Candida albicans. A single injection of F11A and D5A derivatives, in contrast with T11F, led to a significant increase in survival of larvae injected with a lethal inoculum of C. albicans cells, in comparison with infected animals treated with saline. Peptide modulation of host immunity upon C. albicans infection was determined by hemocyte analysis and larval histology, highlighting a different immune stimulation by the studied peptides. F11A, particularly, was the most active in eliciting nodule formation, melanization and fat body activation, leading to a better control of yeast infection. Overall, the obtained data suggest a double role for F11A, able to simultaneously target the fungus and the host immune system, resulting in a more efficient pathogen clearance.

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