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17 pages, 4740 KB

Open AccessArticle

MucR from Sinorhizobium meliloti: New Insights into Its DNA Targets and Its Ability to Oligomerize

by Martina Slapakova, Domenico Sgambati, Luciano Pirone, Veronica Russo, Gianluca D’Abrosca, Mariangela Valletta, Rosita Russo, Angela Chambery, Gaetano Malgieri, Emilia Maria Pedone, Remus Thei Dame, Paolo Vincenzo Pedone and Ilaria Baglivo

Int. J. Mol. Sci. 2023, 24(19), 14702; https://doi.org/10.3390/ijms241914702 - 29 Sep 2023

Cited by 3 | Viewed by 2164

Abstract

Proteins of the MucR/Ros family play a crucial role in bacterial infection or symbiosis with eukaryotic hosts. MucR from Sinorhizobium meliloti plays a regulatory role in establishing symbiosis with the host plant, both dependent and independent of Quorum Sensing. Here, we report the first characterization of MucR isolated from Sinorhizobium meliloti by mass spectrometry and demonstrate that this protein forms higher-order oligomers in its native condition of expression by SEC-MALS. We show that MucR purified from Sinorhizobium meliloti can bind DNA and recognize the region upstream of the ndvA gene in EMSA, revealing that this gene is a direct target of MucR. Although MucR DNA binding activity was already described, a detailed characterization of Sinorhizobium meliloti DNA targets has never been reported. We, thus, analyze sequences recognized by MucR in the rem gene promoter, showing that this protein recognizes AT-rich sequences and does not require a consensus sequence to bind DNA. Furthermore, we investigate the dependence of MucR DNA binding on the length of DNA targets. Taken together, our studies establish MucR from Sinorhizobium meliloti as a member of a new family of Histone-like Nucleoid Structuring (H-NS) proteins, thus explaining the multifaceted role of this protein in many species of alpha-proteobacteria. Full article

(This article belongs to the Special Issue Protein and DNA Interactions: 2nd Edition)

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17 pages, 2587 KB

Open AccessArticle

YbdO Promotes the Pathogenicity of Escherichia coli K1 by Regulating Capsule Synthesis

by Yu Fan, Hongmin Sun, Wen Yang, Jing Bai, Peng Liu, Min Huang, Xi Guo, Bin Yang and Lu Feng

Int. J. Mol. Sci. 2022, 23(10), 5543; https://doi.org/10.3390/ijms23105543 - 16 May 2022

Cited by 9 | Viewed by 3628

Abstract

Escherichia coli K1 is the most popular neonatal meningitis-causing Gram-negative bacterium. As a key virulence determinant, the K1 capsule enhances the survival of E. coli K1 in human brain microvascular endothelial cells (HBMECs) upon crossing the blood–brain barrier; however, the regulatory mechanisms of capsule synthesis during E. coli K1 invasion of HBMECs remain unclear. Here, we identified YbdO as a transcriptional regulator that promotes E. coli K1 invasion of HBMECs by directly activating K1 capsule gene expression to increase K1 capsule synthesis. We found that ybdO deletion significantly reduced HBMEC invasion by E. coli K1 and meningitis occurrence in mice. Additionally, electrophoretic mobility shift assay and chromatin immunoprecipitation–quantitative polymerase chain reaction analysis indicated that YbdO directly activates kpsMT and neuDBACES expression, which encode products involved in K1 capsule transport and synthesis by directly binding to the kpsM promoter. Furthermore, ybdO transcription was directly repressed by histone-like nucleoid structuring protein (H-NS), and we observed that acidic pH similar to that of early and late endosomes relieves this transcriptional repression. These findings demonstrated the regulatory mechanism of YbdO on K1 capsule synthesis, providing further insights into the evolution of E. coli K1 pathogenesis and host–pathogen interaction. Full article

(This article belongs to the Special Issue Bacterial Adaptive Strategies inside Host Cells)

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7 pages, 1050 KB

Open AccessCommunication

Involvement of the Histone-Like Nucleoid Structuring Protein (H-NS) in Acinetobacter baumannii’s Natural Transformation

by Casin Le, Camila Pimentel, Marisel R. Tuttobene, Tomás Subils, Jenny Escalante, Brent Nishimura, Susana Arriaga, Deja Rodgers, Robert A. Bonomo, Rodrigo Sieira, Marcelo E. Tolmasky and María Soledad Ramírez

Pathogens 2021, 10(9), 1083; https://doi.org/10.3390/pathogens10091083 - 26 Aug 2021

Cited by 6 | Viewed by 3266

Abstract

Most Acinetobacter baumannii strains are naturally competent. Although some information is available about factors that enhance or reduce the frequency of the transformation of this bacterium, the regulatory elements and mechanisms are barely understood. In this article, we describe studies on the role of the histone-like nucleoid structuring protein, H-NS, in the regulation of the expression of genes related to natural competency and the ability to uptake foreign DNA. The expression levels of the natural transformation-related genes pilA, pilT, pilQ, comEA, comEC, comF, and drpA significantly increased in a Δhns derivative of A. baumannii A118. The complementation of the mutant with a recombinant plasmid harboring hns restored the expression levels of six of these genes (pilT remained expressed at high levels) to those of the wild-type strain. The transformation frequency of the A. baumannii A118 Δhns strain was significantly higher than that of the wild-type. Similar, albeit not identical, there were consequences when hns was deleted from the hypervirulent A. baumannii AB5075 strain. In the AB5075 complemented strain, the reduction in gene expression in a few cases was not so pronounced that it reached wild-type levels, and the expression of comEA was enhanced further. In conclusion, the expression of all seven transformation-related genes was enhanced after deleting hns in A. baumannii A118 and AB5075, and these modifications were accompanied by an increase in the cells’ transformability. The results highlight a role of H-NS in A. baumannii’s natural competence. Full article

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10 pages, 1952 KB

Open AccessCommunication

Fluorescence Spectroscopic Analysis of ppGpp Binding to cAMP Receptor Protein and Histone-Like Nucleoid Structuring Protein

by Taner Duysak, Thanh Tuyen Tran, Aqeel Rana Afzal and Che-Hun Jung

Int. J. Mol. Sci. 2021, 22(15), 7871; https://doi.org/10.3390/ijms22157871 - 23 Jul 2021

Cited by 2 | Viewed by 2785

Abstract

The cyclic AMP receptor protein (CRP) is one of the best-known transcription factors, regulating about 400 genes. The histone-like nucleoid structuring protein (H-NS) is one of the nucleoid-forming proteins and is responsible for DNA packaging and gene repression in prokaryotes. In this study, the binding of ppGpp to CRP and H-NS was determined by fluorescence spectroscopy. CRP from Escherichia coli exhibited intrinsic fluorescence at 341 nm when excited at 280 nm. The fluorescence intensity decreased in the presence of ppGpp. The dissociation constant of 35 ± 3 µM suggests that ppGpp binds to CRP with a similar affinity to cAMP. H-NS also shows intrinsic fluorescence at 329 nm. The fluorescence intensity was decreased by various ligands and the calculated dissociation constant for ppGpp was 80 ± 11 µM, which suggests that the binding site was occupied fully by ppGpp under starvation conditions. This study suggests the modulatory effects of ppGpp in gene expression regulated by CRP and H-NS. The method described here may be applicable to many other proteins. Full article

(This article belongs to the Section Biochemistry)

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18 pages, 2562 KB

Open AccessArticle

Metagenomic Analysis of Microbial Community Compositions and Cold-Responsive Stress Genes in Selected Antarctic Lacustrine and Soil Ecosystems

by Hyunmin Koo, Joseph A. Hakim, Casey D. Morrow, Michael R. Crowley, Dale T. Andersen and Asim K. Bej

Life 2018, 8(3), 29; https://doi.org/10.3390/life8030029 - 11 Jul 2018

Cited by 36 | Viewed by 8055

Abstract

This study describes microbial community compositions, and various cold-responsive stress genes, encompassing cold-induced proteins (CIPs) and cold-associated general stress-responsive proteins (CASPs) in selected Antarctic lake water, sediment, and soil metagenomes. Overall, Proteobacteria and Bacteroidetes were the major taxa in all metagenomes. Prochlorococcus and Thiomicrospira were highly abundant in waters, while Myxococcus, Anaeromyxobacter, Haliangium, and Gloeobacter were dominant in the soil and lake sediment metagenomes. Among CIPs, genes necessary for DNA replication, translation initiation, and transcription termination were highly abundant in all metagenomes. However, genes for fatty acid desaturase (FAD) and trehalose synthase (TS) were common in the soil and lake sediment metagenomes. Interestingly, the Lake Untersee water and sediment metagenome samples contained histone-like nucleoid structuring protein (H-NS) and all genes for CIPs. As for the CASPs, high abundances of a wide range of genes for cryo- and osmo-protectants (glutamate, glycine, choline, and betaine) were identified in all metagenomes. However, genes for exopolysaccharide biosynthesis were dominant in Lake Untersee water, sediment, and other soil metagenomes. The results from this study indicate that although diverse microbial communities are present in various metagenomes, they share common cold-responsive stress genes necessary for their survival and sustenance in the extreme Antarctic conditions. Full article

(This article belongs to the Section Microbiology)

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15 pages, 3301 KB

Open AccessReview

H-NS, Its Family Members and Their Regulation of Virulence Genes in Shigella Species

by Michael A. Picker and Helen J. Wing

Genes 2016, 7(12), 112; https://doi.org/10.3390/genes7120112 - 1 Dec 2016

Cited by 24 | Viewed by 8227

Abstract

The histone-like nucleoid structuring protein (H-NS) has played a key role in shaping the evolution of Shigella spp., and provides the backdrop to the regulatory cascade that controls virulence by silencing many genes found on the large virulence plasmid. H-NS and its paralogue StpA are present in all four Shigella spp., but a second H-NS paralogue, Sfh, is found in the Shigella flexneri type strain 2457T, which is routinely used in studies of Shigella pathogenesis. While StpA and Sfh have been proposed to serve as “molecular backups” for H-NS, the apparent redundancy of these proteins is questioned by in vitro studies and work done in Escherichia coli. In this review, we describe the current understanding of the regulatory activities of the H-NS family members, the challenges associated with studying these proteins and their role in the regulation of virulence genes in Shigella. Full article

(This article belongs to the Special Issue Virulence Gene Regulation in Bacteria)

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12 pages, 1657 KB

Open AccessReview

Functional Evolution of Bacterial Histone-Like HU Proteins

by Anne Grove

Curr. Issues Mol. Biol. 2011, 13(1), 1-12; https://doi.org/10.21775/cimb.013.001 - 20 May 2010

Cited by 10 | Viewed by 1603

Abstract

Bacterial histone-like HU proteins are critical to maintenance of the nucleoid structure. In addition, they participate in all DNA-dependent functions, including replication, repair, recombination and gene regulation. In these capacities, their function is typically architectural, inducing a specific DNA topology that promotes assembly of higher-order nucleo-protein structures. Although HU proteins are highly conserved, individual homologs have been shown to exhibit a wide range of different DNA binding specificities and affinities. The existence of such distinct specificities indicates functional evolution and predicts distinct in vivo roles. Emerging evidence suggests that HU proteins discriminate between DNA target sites based on intrinsic flexure, and that two primary features of protein binding contribute to target site selection: The extent to which protein-mediated DNA kinks are stabilized and a network of surface salt-bridges that modulate interaction between DNA flanking the kinks and the body of the protein. These features confer target site selection for a specific HU homolog, they suggest the ability of HU to induce different DNA structural deformations depending on substrate, and they explain the distinct binding properties characteristic of HU homologs. Further divergence is evidenced by the existence of HU homologs with an additional lysine-rich domain also found in eukaryotic histone H1. Full article

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