RESUMEN
It has been demonstrated that the alpha2 chain of laminin-2 present on the surface of Schwann cells is involved in the process of attachment of Mycobacterium leprae to these cells. Searching for M. leprae laminin-binding molecules, in a previous study we isolated and characterized the cationic proteins histone-like protein (Hlp) and ribosomal proteins S4 and S5 as potential adhesins involved in M. leprae-Schwann cell interaction. Hlp was shown to bind alpha2-laminins and to greatly enhance the attachment of mycobacteria to ST88-14 Schwann cells. In the present study, we investigated the laminin-binding capacity of the ribosomal proteins S4 and S5. The genes coding for these proteins were PCR amplified and their recombinant products were shown to bind alpha2-laminins in overlay assays. However, when tested in ELISA-based assays and in adhesion assays with ST88-14 cells, in contrast to Hlp, S4 and S5 failed to bind laminin and act as adhesins. The laminin-binding property and adhesin capacity of two basic host-derived proteins were also tested, and only histones, but not cytochrome c, were able to increase bacterial attachment to ST88-14 cells. Our data suggest that the alanine/lysine-rich sequences shared by Hlp and eukaryotic H1 histones might be involved in the binding of these cationic proteins to laminin.
Asunto(s)
Proteínas Bacterianas/metabolismo , Laminina/metabolismo , Mycobacterium leprae/química , Proteínas Ribosómicas/metabolismo , Animales , Armadillos , Adhesión Bacteriana/fisiología , Proteínas Bacterianas/genética , Proteínas Bacterianas/aislamiento & purificación , Clonación Molecular , Electroforesis en Gel de Poliacrilamida , Ensayo de Inmunoadsorción Enzimática , Escherichia coli/genética , Histonas/metabolismo , Humanos , Mycobacterium leprae/genética , Reacción en Cadena de la Polimerasa , Unión Proteica/fisiología , Proteínas Ribosómicas/genética , Proteínas Ribosómicas/aislamiento & purificación , Células de Schwann/microbiología , Células de Schwann/fisiologíaRESUMEN
It has been demonstrated that the alpha2 chain of laminin-2 present on the surface of Schwann cells is involved in the process of attachment of Mycobacterium leprae to these cells. Searching for M. leprae laminin-binding molecules, in a previous study we isolated and characterized the cationic proteins histone-like protein (Hlp) and ribosomal proteins S4 and S5 as potential adhesins involved in M. leprae-Schwann cell interaction. Hlp was shown to bind alpha2-laminins and to greatly enhance the attachment of mycobacteria to ST88-14 Schwann cells. In the present study, we investigated the laminin-binding capacity of the ribosomal proteins S4 and S5. The genes coding for these proteins were PCR amplified and their recombinant products were shown to bind alpha2-laminins in overlay assays. However, when tested in ELISA-based assays and in adhesion assays with ST88-14 cells, in contrast to Hlp, S4 and S5 failed to bind laminin and act as adhesins. The laminin-binding property and adhesin capacity of two basic host-derived proteins were also tested, and only histones, but not cytochrome c, were able to increase bacterial attachment to ST88-14 cells. Our data suggest that the alanine/lysine-rich sequences shared by Hlp and eukaryotic H1 histones might be involved in the binding of these cationic proteins to laminin
Asunto(s)
Humanos , Animales , Laminina/metabolismo , Mycobacterium leprae/metabolismo , Proteínas Ribosómicas/metabolismo , Armadillos , Adhesión Celular , Clonación Molecular , Electroforesis en Gel de Poliacrilamida , Ensayo de Inmunoadsorción Enzimática , Escherichia coli/genética , Histonas/metabolismo , Mycobacterium leprae/genética , Reacción en Cadena de la Polimerasa , Unión Proteica/fisiología , Proteínas Ribosómicas/genética , Proteínas Ribosómicas/aislamiento & purificación , Células de Schwann/fisiologíaRESUMEN
It has recently been demonstrated that laminin alpha2 chains present on the surface of Schwann cells are involved in the process of attachment of Mycobacterium leprae to these cells. In this study, a protein in the M. leprae cell wall that was found to be capable of binding alpha2-containing laminins (merosin) was isolated and characterized. The M. leprae laminin-binding protein was identified as a 21-kDa histone-like protein (Hlp), a highly conserved cationic protein present in other species of mycobacteria. The gene that encodes this protein was PCR amplified, cloned, and expressed, and the recombinant protein was shown to bind alpha2-laminins. More significantly, when added exogenously, Hlp was able to greatly enhance the attachment of mycobacteria to ST88-14 human Schwann cells. The capacity to bind alpha2-laminins and to enhance mycobacterial adherence to Schwann cells was also found in other cationic proteins such as host-derived histones. Moreover, mutation in the hlp gene was shown not to affect the capacity of mycobacteria to bind to ST88-14 cells, suggesting that alternative adhesins and/or pathways might be used by mycobacteria during the process of adherence to Schwann cells. The potential role of Hlp as a fortuitous virulence factor contributing to the pathogenesis of M. leprae-mediated nerve damage is discussed.