A calcium-dependent bacterial surface protein is involved in the attachment of rhizobia to peanut roots

Abstract

Abstract: As part of a project to characterize molecules involved in the crack-entry infection process leading to nodule development, a microscopic assay was used to visualize the attachment of cells of Bradyrhizobium sp. strains SEMIA 6144 and TAL 1000 (labelled by introducing a plasmid expressing constitutively the green fluorescent protein GFP-S65T) to and TAL 1000 (labelled by introducing a plasmid expressing constitutively the green fluorescent protein GFP-S65T) to a microscopic assay was used to visualize the attachment of cells of Bradyrhizobium sp. strains SEMIA 6144 and TAL 1000 (labelled by introducing a plasmid expressing constitutively the green fluorescent protein GFP-S65T) to and TAL 1000 (labelled by introducing a plasmid expressing constitutively the green fluorescent protein GFP-S65T) to As part of a project to characterize molecules involved in the crack-entry infection process leading to nodule development, a microscopic assay was used to visualize the attachment of cells of Bradyrhizobium sp. strains SEMIA 6144 and TAL 1000 (labelled by introducing a plasmid expressing constitutively the green fluorescent protein GFP-S65T) to and TAL 1000 (labelled by introducing a plasmid expressing constitutively the green fluorescent protein GFP-S65T) to Bradyrhizobium sp. strains SEMIA 6144 and TAL 1000 (labelled by introducing a plasmid expressing constitutively the green fluorescent protein GFP-S65T) to Arachis hypogaea L. (peanut). Qualitative and quantitative results revealed that attachment was strongly dependent on the growth phase of the bacteria. Optimal attachment occurred when bacteria were at the late log or early stationary phase. Cell surface proteins from the Bradyrhizobium sp. strains inhibited the attachment when supplied prior to the attachment assay. Root incubation with a 14-kDa protein (eluted from sodium dodecyl sulphate – gel electrophoresis of the cell surface fraction) prior to the attachment assay resulted in a strong decrease of attachment. The adhesin appeared to be a calcium-binding protein, since cells treated with EDTA were found to be able to bind to adhesin-treated peanut roots. Since this protein has properties identical to those reported for rhicadhesin, we propose that this adhesin is also involved in the attachment process of rhizobia to root legumes that are infected by the crack-entry process. attachment assay. Root incubation with a 14-kDa protein (eluted from sodium dodecyl sulphate – gel electrophoresis of the cell surface fraction) prior to the attachment assay resulted in a strong decrease of attachment. The adhesin appeared to be a calcium-binding protein, since cells treated with EDTA were found to be able to bind to adhesin-treated peanut roots. Since this protein has properties identical to those reported for rhicadhesin, we propose that this adhesin is also involved in the attachment process of rhizobia to root legumes that are infected by the crack-entry process. the growth phase of the bacteria. Optimal attachment occurred when bacteria were at the late log or early stationary phase. Cell surface proteins from the Bradyrhizobium sp. strains inhibited the attachment when supplied prior to the attachment assay. Root incubation with a 14-kDa protein (eluted from sodium dodecyl sulphate – gel electrophoresis of the cell surface fraction) prior to the attachment assay resulted in a strong decrease of attachment. The adhesin appeared to be a calcium-binding protein, since cells treated with EDTA were found to be able to bind to adhesin-treated peanut roots. Since this protein has properties identical to those reported for rhicadhesin, we propose that this adhesin is also involved in the attachment process of rhizobia to root legumes that are infected by the crack-entry process. attachment assay. Root incubation with a 14-kDa protein (eluted from sodium dodecyl sulphate – gel electrophoresis of the cell surface fraction) prior to the attachment assay resulted in a strong decrease of attachment. The adhesin appeared to be a calcium-binding protein, since cells treated with EDTA were found to be able to bind to adhesin-treated peanut roots. Since this protein has properties identical to those reported for rhicadhesin, we propose that this adhesin is also involved in the attachment process of rhizobia to root legumes that are infected by the crack-entry process. L. (peanut). Qualitative and quantitative results revealed that attachment was strongly dependent on the growth phase of the bacteria. Optimal attachment occurred when bacteria were at the late log or early stationary phase. Cell surface proteins from the Bradyrhizobium sp. strains inhibited the attachment when supplied prior to the attachment assay. Root incubation with a 14-kDa protein (eluted from sodium dodecyl sulphate – gel electrophoresis of the cell surface fraction) prior to the attachment assay resulted in a strong decrease of attachment. The adhesin appeared to be a calcium-binding protein, since cells treated with EDTA were found to be able to bind to adhesin-treated peanut roots. Since this protein has properties identical to those reported for rhicadhesin, we propose that this adhesin is also involved in the attachment process of rhizobia to root legumes that are infected by the crack-entry process. attachment assay. Root incubation with a 14-kDa protein (eluted from sodium dodecyl sulphate – gel electrophoresis of the cell surface fraction) prior to the attachment assay resulted in a strong decrease of attachment. The adhesin appeared to be a calcium-binding protein, since cells treated with EDTA were found to be able to bind to adhesin-treated peanut roots. Since this protein has properties identical to those reported for rhicadhesin, we propose that this adhesin is also involved in the attachment process of rhizobia to root legumes that are infected by the crack-entry process. Bradyrhizobium sp. strains inhibited the attachment when supplied prior to the attachment assay. Root incubation with a 14-kDa protein (eluted from sodium dodecyl sulphate – gel electrophoresis of the cell surface fraction) prior to the attachment assay resulted in a strong decrease of attachment. The adhesin appeared to be a calcium-binding protein, since cells treated with EDTA were found to be able to bind to adhesin-treated peanut roots. Since this protein has properties identical to those reported for rhicadhesin, we propose that this adhesin is also involved in the attachment process of rhizobia to root legumes that are infected by the crack-entry process.

Dans le cadre d’un projet visant à caractériser les molécules impliquées dans le processus d’infection par entrée par les fissures, menant au développement de nodules, une analyse microscopique a été effectuée afin de visualiser l’attachement de cellules de Bradyrhizobium sp. SEMIA 6144 et TAL 1000 (marquées grâce à l’introduction d’un plasmide exprimant constitutivement la protéine fluorescente verte GFP-S65T) à Arachis hypogaea L. (arachide). Les résultats qualitatifs et quantitatifs on révélé que l’attachement était fortement dépendant de la phase de croissance de la bactérie. L’attachement était optimal lorsque les bactéries étaient dans leurs phases logarithmique tardive ou stationnaire précoce. Les protéines de la surface cellulaire de ces souches ont inhibé l’attachement lorsqu’elles étaient fournies avant l’analyse d’attachement. Une incubation préalable de racines avec une protéine de 14 kDa (éluée d’un gel d’électrophorèse – SDS de la fraction de la surface cellulaire) a entraîné une forte diminution de l’attachement. Cette adhésine semble être une protéine se liant au calcium puisque des cellules traitées au EDTA ont pu se lier à des racines d’arachides traitées à l’adhésine. Comme cette protéine a des caractéristiques identiques à celles de la rhicadhésine, nous proposons que cette adhésine est également impliquée dans le processus d’attachement de rhizobiums aux racines de légumineuses qui sont infectées par le processus d’entrée par les fissures.