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A Histone-Like Nucleoid Structuring Protein Regulates Several Virulence Traits in Burkholderia multivorans.

ABSTRACT: Burkholderia cepacia complex bacteria comprise opportunistic pathogens causing chronic respiratory infections in cystic fibrosis (CF) patients. These microorganisms produce an exopolysaccharide named cepacian, which is considered a virulence determinant. To find genes implicated in the regulation of cepacian biosynthesis, we characterized an evolved nonmucoid variant (17616nmv) derived from the ancestor, Burkholderia multivorans ATCC 17616, after prolonged stationary phase. Lack of cepacian biosynthesis was correlated with downregulation of the expression of bce genes implicated in its biosynthesis. Furthermore, genome sequencing of the variant identified the transposition of the mobile element IS406 upstream of the coding sequence of an hns-like gene (Bmul_0158) encoding a histone-like nucleoid structuring (H-NS) protein, a known global transcriptional repressor. This insertion sequence (IS) element upregulated the expression of Bmul_0158 by 4-fold. Transcriptome analysis identified the global effects of this mutation on gene expression, with major changes in genes implicated in motility, pilus synthesis, type VI secretion, and chromosome-associated functions. Concomitant with these differences, the nonmucoid variant displays reduced adherence to a CF lung bronchial cell line and reduced surface hydrophobicity and forms smaller cellular aggregates but has an increase in swimming and swarming motilities. Finally, analysis of the GC content of the upstream region of differentially expressed genes led to the identification of various genomic regions, possibly acquired by horizontal gene transfer, which were transcriptionally repressed by the increased expression of the Bmul_0158 gene in the 17616nmv strain. Taken together, the results revealed a significant role for this H-NS protein in the regulation of B. multivorans persistence- and virulence-associated genes. IMPORTANCE Members of the histone-like nucleoid structuring (H-NS) family of proteins, present in many bacteria, are important global regulators of gene expression. Many of the regulated genes were acquired horizontally and include pathogenicity islands and prophages, among others. Additionally, H-NS can play a structural role by bridging and compacting DNA, fulfilling a crucial role in cell physiology. Several virulence phenotypes have been frequently identified in several bacteria as dependent on H-NS activity. Here, we describe an H-NS-like protein of the opportunistic pathogen Burkholderia multivorans, a species commonly infecting the respiratory tract of cystic fibrosis patients. Our results indicate that this protein is involved in regulating virulence traits such as exopolysaccharide biosynthesis, adhesion to biotic surfaces, cellular aggregation, and motility. Furthermore, this H-NS-like protein is one out of eight orthologs present in the B. multivorans ATCC 17616 genome, posing relevant questions to be investigated on how these proteins coordinate the expression of virulence traits.

SUBMITTER: Gomes SC

PROVIDER: S-EPMC8231446 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:The cholera disease bacterium V. cholerae, can adopt planktonic or biofilm lifestyles depending on the intracellular concentration of the second messenger cyclic diguanylic acid (c-di-GMP). Biofilm formation protects Vibrios from stressful conditions and facilitates disease transmission by enhancing infectivity. The histone-like nucleoid structuring protein (H-NS) is a global regulator of genes associated with pathogenicity and responses to environmental stresses. H-NS represses the transcription of genes vpsT, vpsA and vpsL, which are required for the biosynthesis of the biofilm exopolysacchide matrix. Here we demonstrate that the c-di-GMP-binding protein VpsT disrupts H-NS nucleoprotein complexes at the vpsA and vpsL promoters and that this effect is enhanced by c-di-GMP. We used ChIP coupled with Next Generation Sequencing (ChIP-Seq) and transcriptome analysis (RNA-Seq) to identify additional loci repressed by H-NS affecting biofilm formation. This study showed that H-NS directly represses the transcription of genes encoding proteins present in the biofilm matrix such as the rbmA-F cluster, hemolysin and chitinase. Similar to vpsA and vpsL, the promoter region of vpsU, rbmA and rbmF exhibited overlapping H-NS and VpsT binding motifs. Deletion of vpsT increased H-NS occupancy at the vpsU, vpsA, vpsL, rbmA and rbmF promoters. Conversely, artificially increasing the c-di-GMP pool diminished H-NS occupancy at the above promoters. Deletion of vpsT did not affect H-NS occupancy at its own promoter. However, deletion of genes encoding the regulators AphA and VpsR significantly increased H-NS occupancy at the vpsT promoter. In sum, our study shows that c-di-GMP enhances biofilm formation by acting through VpsT to activate an H-NS anti-repression cascade. The Binding profile of V. cholerae H-NS to the genome was determined by ChIP followed by Next Generation Sequencing (ChIP-Seq) using the Illumina HiSeq2000 platform. V. cholerae C7258 cells expressing H-NS-FLAG fusion protein from the hns transcription and translation signals were collected from LB cultures grown to mid-exponential phase (OD600 0.5). An anti-FLAG Immunoprecipitation (IP) and an Input samples were used for the analysis.

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A Histone-Like Nucleoid Structuring Protein Regulates Several Virulence Traits in Burkholderia multivorans.

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