Identification of the Quorum-Sensing Target DNA Sequence and N -Acyl Homoserine Lactone Responsiveness of the Brucella abortus virB promoter

Author:

Arocena Gastón M.1,Sieira Rodrigo1,Comerci Diego J.12,Ugalde Rodolfo A.1

Affiliation:

1. Instituto de Investigaciones Biotecnológicas, Universidad Nacional de General San Martín, IIB-INTECH-CONICET, San Martín 1650, Buenos Aires, Argentina

2. Comisión Nacional de Energía Atómica, División Agropecuaria, Centro Atómico Ezeiza 1804, Buenos Aires, Argentina

Abstract

ABSTRACT VjbR is a LuxR-type quorum-sensing (QS) regulator that plays an essential role in the virulence of the intracellular facultative pathogen Brucella , the causative agent of brucellosis. It was previously described that VjbR regulates a diverse group of genes, including the virB operon. The latter codes for a type IV secretion system (T4SS) that is central for the pathogenesis of Brucella . Although the regulatory role of VjbR on the virB promoter (P virB ) was extensively studied by different groups, the VjbR-binding site had not been identified so far. Here, we identified the target DNA sequence of VjbR in P virB by DNase I footprinting analyses. Surprisingly, we observed that VjbR specifically recognizes a sequence that is identical to a half-binding site of the QS-related regulator MrtR of Mesorhizobium tianshanense . As shown by DNase I footprinting and electrophoretic mobility shift assays, generation of a palindromic MrtR-like-binding site in P virB increased both the affinity and the stability of the VjbR-DNA complex, which confirmed that the QS regulator of Brucella is highly related to that of M. tianshanense . The addition of N -dodecanoyl homoserine lactone dissociated VjbR from the promoter, which confirmed previous reports that indicated a negative effect of this signal on the VjbR-mediated activation of P virB . Our results provide new molecular evidence for the structure of the virB promoter and reveal unusual features of the QS target DNA sequence of the main regulator of virulence in Brucella .

Publisher

American Society for Microbiology

Subject

Molecular Biology,Microbiology

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