Global Transcriptional Analysis of Virus-Host Interactions between Phage ϕ29 and Bacillus subtilis

ABSTRACTThe study of phage-host relationships is essential to understanding the dynamic of microbial systems. Here, we analyze genome-wide interactions ofBacillus subtilisand its lytic phage ϕ29 during the early stage of infection. Simultaneous high-resolution analysis of virus and host transcriptomes by deep RNA sequencing allowed us to identify differentially expressed bacterial genes. Phage ϕ29 induces significant transcriptional changes in about 0.9% (38/4,242) and 1.8% (76/4,242) of the host protein-coding genes after 8 and 16 min of infection, respectively. Gene ontology enrichment analysis clustered upregulated genes into several functional categories, such as nucleic acid metabolism (including DNA replication) and protein metabolism (including translation). Surprisingly, most of the transcriptional repressed genes were involved in the utilization of specific carbon sources such as ribose and inositol, and many contained promoter binding-sites for the catabolite control protein A (CcpA). Another interesting finding is the presence of previously uncharacterized antisense transcripts complementary to the well-known phage ϕ29 messenger RNAs that adds an additional layer to the viral transcriptome complexity.IMPORTANCEThe specific virus-host interactions that allow phages to redirect cellular machineries and energy resources to support the viral progeny production are poorly understood. This study provides, for the first time, an insight into the genome-wide transcriptional response of the Gram-positive modelBacillus subtilisto phage ϕ29 infection.

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BSGatlas: a unified Bacillus subtilis genome and transcriptome annotation atlas with enhanced information access

Microbial Genomics ◽

10.1099/mgen.0.000524 ◽

2021 ◽

Vol 7 (2) ◽

Author(s):

Adrian Sven Geissler ◽

Christian Anthon ◽

Ferhat Alkan ◽

Enrique González-Tortuero ◽

Line Dahl Poulsen ◽

...

Keyword(s):

Bacillus Subtilis ◽

Type Species ◽

Large Scale ◽

Information Access ◽

Model Systems ◽

Transcription Start ◽

Content Type ◽

Link Type ◽

Genome Wide ◽

Transcript Map

A large part of our current understanding of gene regulation in Gram-positive bacteria is based on Bacillus subtilis , as it is one of the most well studied bacterial model systems. The rapid growth in data concerning its molecular and genomic biology is distributed across multiple annotation resources. Consequently, the interpretation of data from further B. subtilis experiments becomes increasingly challenging in both low- and large-scale analyses. Additionally, B. subtilis annotation of structured RNA and non-coding RNA (ncRNA), as well as the operon structure, is still lagging behind the annotation of the coding sequences. To address these challenges, we created the B. subtilis genome atlas, BSGatlas, which integrates and unifies multiple existing annotation resources. Compared to any of the individual resources, the BSGatlas contains twice as many ncRNAs, while improving the positional annotation for 70 % of the ncRNAs. Furthermore, we combined known transcription start and termination sites with lists of known co-transcribed gene sets to create a comprehensive transcript map. The combination with transcription start/termination site annotations resulted in 717 new sets of co-transcribed genes and 5335 untranslated regions (UTRs). In comparison to existing resources, the number of 5′ and 3′ UTRs increased nearly fivefold, and the number of internal UTRs doubled. The transcript map is organized in 2266 operons, which provides transcriptional annotation for 92 % of all genes in the genome compared to the at most 82 % by previous resources. We predicted an off-target-aware genome-wide library of CRISPR–Cas9 guide RNAs, which we also linked to polycistronic operons. We provide the BSGatlas in multiple forms: as a website (https://rth.dk/resources/bsgatlas/), an annotation hub for display in the UCSC genome browser, supplementary tables and standardized GFF3 format, which can be used in large scale -omics studies. By complementing existing resources, the BSGatlas supports analyses of the B. subtilis genome and its molecular biology with respect to not only non-coding genes but also genome-wide transcriptional relationships of all genes.

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Effect of Soybean Coumestrol on Bradyrhizobium japonicum Nodulation Ability, Biofilm Formation, and Transcriptional Profile

Applied and Environmental Microbiology ◽

10.1128/aem.07336-11 ◽

2012 ◽

Vol 78 (8) ◽

pp. 2896-2903 ◽

Cited By ~ 29

Author(s):

Hae-In Lee ◽

Jin-Hwan Lee ◽

Ki-Hun Park ◽

Dipen Sangurdekar ◽

Woo-Suk Chang

Keyword(s):

Bradyrhizobium Japonicum ◽

Biofilm Formation ◽

Transcriptional Analysis ◽

Plant Metabolites ◽

Content Type ◽

Secondary Plant Metabolites ◽

Reverse Transcription Pcr ◽

Genome Wide ◽

A Genome ◽

Soybean Nodulation

ABSTRACTFlavonoids, secondary plant metabolites which mainly have a polyphenolic structure, play an important role in plant-microbe communications for nitrogen-fixing symbiosis. Among 10 polyphenolic compounds isolated from soybean roots in our previous study, coumestrol showed the highest antioxidant activity. In this study, its effect on the soybean nodulation was tested. The soybean symbiontBradyrhizobium japonicumUSDA110 pretreated with 20 μM coumestrol enhanced soybean nodulation by increasing the number of nodules 1.7-fold compared to the control. We also tested the effect of coumestrol onB. japonicumbiofilm formation. At a concentration of 2 μM, coumestrol caused a higher degree of biofilm formation than two major soybean isoflavonoids, genistein and daidzein, although no biofilm formation was observed at a concentration of 20 μM each compound. A genome-wide transcriptional analysis was performed to obtain a comprehensive snapshot of theB. japonicumresponse to coumestrol. When the bacterium was incubated in 20 μM coumestrol for 24 h, a total of 371 genes (139 upregulated and 232 downregulated) were differentially expressed at a 2-fold cutoff with aqvalue of less than 5%. No commonnodgene induction was found in the microarray data. However, quantitative reverse transcription-PCR (qRT-PCR) data showed that incubation for 12 h resulted in a moderate induction (ca. 2-fold) ofnodD1andnodABC, indicating that soybean coumestrol is a weak inducer of commonnodgenes. In addition, disruption ofnfeD(bll4952) affected the soybean nodulation by an approximate 30% reduction in the average number of nodules.

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Genome-Wide Characterization of the Fur Regulatory Network Reveals a Link between Catechol Degradation and Bacillibactin Metabolism in Bacillus subtilis

mBio ◽

10.1128/mbio.01451-18 ◽

2018 ◽

Vol 9 (5) ◽

Cited By ~ 4

Author(s):

Hualiang Pi ◽

John D. Helmann

Keyword(s):

Bacillus Subtilis ◽

Binding Sites ◽

Regulatory Network ◽

Iron Acquisition ◽

Sufficient Conditions ◽

Enteric Bacteria ◽

Degradation Pathway ◽

Content Type ◽

Genome Wide

ABSTRACT The ferric uptake regulator (Fur) is the global iron biosensor in many bacteria. Fur functions as an iron-dependent transcriptional repressor for most of its regulated genes. There are a few examples where holo-Fur activates transcription, either directly or indirectly. Recent studies suggest that apo-Fur might also act as a positive regulator and that, besides iron metabolism, the Fur regulon might encompass other biological processes such as DNA synthesis, energy metabolism, and biofilm formation. Here, we obtained a genomic view of the Fur regulatory network in Bacillus subtilis using chromatin immunoprecipitation sequencing (ChIP-seq). Besides the known Fur target sites, 70 putative DNA binding sites were identified, and the vast majority had higher occupancy under iron-sufficient conditions. Among the new sites detected, a Fur binding site in the promoter region of the catDE operon is of particular interest. This operon, encoding catechol 2,3-dioxygenase, is critical for catechol degradation and is under negative regulation of CatR and YodB. These three repressors (Fur, CatR, and YodB) function cooperatively to regulate the transcription of catDE, with Fur functioning as a sensor of iron limitation and CatR as the major sensor of catechol stress. Genetic analysis suggests that CatDE is involved in metabolism of the catecholate siderophore bacillibactin, particularly when bacillibactin is constitutively produced and accumulates intracellularly, potentially generating endogenous toxic catechol derivatives. This study documents a role for catechol degradation in bacillibactin metabolism and provides evidence that catechol 2,3-dioxygenase can detoxify endogenously produced catechol substrates in addition to its more widely studied role in biodegradation of environmental aromatic compounds and pollutants. IMPORTANCE Many bacteria synthesize high-affinity iron chelators (siderophores). Siderophore-mediated iron acquisition is an efficient and widely utilized strategy for bacteria to meet their cellular iron requirements. One prominent class of siderophores uses catecholate groups to chelate iron. B. subtilis bacillibactin, structurally similar to enterobactin (made by enteric bacteria), is a triscatecholate siderophore that is hydrolyzed to monomeric units after import to release iron. However, the ultimate fates of these catechol compounds and their potential toxicities have not been defined previously. We performed genome-wide identification of Fur binding sites in vivo and uncovered a connection between catechol degradation and bacillibactin metabolism in B. subtilis. Besides its role in the detoxification of environmental catechols, the catechol 2,3-dioxygenase encoded by catDE also protects cells from intoxication by endogenous bacillibactin-derived catechol metabolites under iron-limited conditions. These findings shed light on the degradation pathway and precursor recycling of the catecholate siderophores.

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Ehrlichia chaffeensis TRP120 Moonlights as a HECT E3 Ligase Involved in Self- and Host Ubiquitination To Influence Protein Interactions and Stability for Intracellular Survival

Infection and Immunity ◽

10.1128/iai.00290-17 ◽

2017 ◽

Vol 85 (9) ◽

Cited By ~ 7

Author(s):

Bing Zhu ◽

Seema Das ◽

Shubhajit Mitra ◽

Tierra R. Farris ◽

Jere W. McBride

Keyword(s):

Ubiquitin Ligase ◽

Protein Interactions ◽

Ectopic Expression ◽

E3 Ligase ◽

Intracellular Survival ◽

Host Protein ◽

Ehrlichia Chaffeensis ◽

Content Type ◽

Host Interactions

ABSTRACT Ehrlichia chaffeensis secretes tandem repeat protein (TRP) effectors that are involved in a diverse array of host cell interactions, some of which directly activate cell signaling pathways and reprogram host gene transcription to promote survival in the mononuclear phagocyte. However, the molecular details of these effector-host interactions and roles in pathobiology are incompletely understood. In this study, we determined that the E. chaffeensis effector TRP120 is posttranslationally modified by ubiquitin (Ub) and that ubiquitination occurs through intrinsic and host-mediated HECT ligase activity. A functional HECT E3 ligase domain with a conserved catalytic site was identified in the C-terminal region of TRP120, and TRP120 autoubiquitination occurred in vitro in the presence of host UbcH5b/c E2 enzymes. TRP120 ubiquitination sites were mapped using a high-density microfluidic peptide array and confirmed by ectopic expression of TRP120 lysine mutants in cells. Moreover, we determined that the HECT E3 ubiquitin ligase, Nedd4L, interacts with TRP120 during infection and also mediates TRP120 ubiquitination. Nedd4L knockdown resulted in the reduction of TRP120-Ub, decreased ehrlichial infection, and reduced recruitment of a known TRP120-interacting host protein, PCGF5, to ehrlichial inclusions. TRP120-mediated PCGF5 polyubiquitination was associated with a reduction in PCGF5 levels. Inhibition of ubiquitination with small molecules also significantly decreased ehrlichial infection, indicating that the Ub pathway is critical for ehrlichial intracellular replication and survival. The current study identified a novel E. chaffeensis ubiquitin ligase and revealed an important role for the ubiquitin pathway in effector-host interactions and pathogen-mediated host protein stability in order to promote intracellular survival.

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Genome-Wide Transcriptional Analysis of the Phosphate Starvation Stimulon of Bacillus subtilis

Journal of Bacteriology ◽

10.1128/jb.187.23.8063-8080.2005 ◽

2005 ◽

Vol 187 (23) ◽

pp. 8063-8080 ◽

Cited By ~ 45

Author(s):

Nicholas E. E. Allenby ◽

Nicola O'Connor ◽

Zoltán Prágai ◽

Alan C. Ward ◽

Anil Wipat ◽

...

Keyword(s):

Bacillus Subtilis ◽

Phosphate Starvation ◽

Transcriptional Analysis ◽

Specific Response ◽

Starvation Stress ◽

Nonspecific Resistance ◽

New Members ◽

Genome Wide ◽

A Genome ◽

Cellular Components

ABSTRACT Bacillus subtilis responds to phosphate starvation stress by inducing the PhoP and SigB regulons. While the PhoP regulon provides a specific response to phosphate starvation stress, maximizing the acquisition of phosphate (Pi) from the environment and reducing the cellular requirement for this essential nutrient, the SigB regulon provides nonspecific resistance to stress by protecting essential cellular components, such as DNA and membranes. We have characterized the phosphate starvation stress response of B. subtilis at a genome-wide level using DNA macroarrays. A combination of outlier and cluster analyses identified putative new members of the PhoP regulon, namely, yfkN (2′,3′ cyclic nucleotide 2′-phosphodiesterase), yurI (RNase), yjdB (unknown), and vpr (extracellular serine protease). YurI is thought to be responsible for the nonspecific degradation of RNA, while the activity of YfkN on various nucleotide phosphates suggests that it could act on substrates liberated by YurI, which produces 3′ or 5′ phosphoribonucleotides. The putative new PhoP regulon members are either known or predicted to be secreted and are likely to be important for the recovery of inorganic phosphate from a variety of organic sources of phosphate in the environment.

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ThespsGene Products Affect the Germination, Hydrophobicity, and Protein Adsorption of Bacillus subtilis Spores

Applied and Environmental Microbiology ◽

10.1128/aem.02893-14 ◽

2014 ◽

Vol 80 (23) ◽

pp. 7293-7302 ◽

Cited By ~ 17

Author(s):

Giuseppina Cangiano ◽

Teja Sirec ◽

Cristina Panarella ◽

Rachele Isticato ◽

Loredana Baccigalupi ◽

...

Keyword(s):

Bacillus Subtilis ◽

Experimental Validation ◽

Surface Display ◽

Deep Understanding ◽

Transcriptional Analysis ◽

Functional Study ◽

Spore Surface ◽

Wild Type ◽

Content Type ◽

Hydrophobic Force

ABSTRACTThe multilayered surface of theBacillus subtilisspore is composed of proteins and glycans. While over 70 different proteins have been identified as surface components, carbohydrates associated with the spore surface have not been characterized in detail yet. Bioinformatic data suggest that the 11 products of thespsoperon are involved in the synthesis of polysaccharides present on the spore surface, but an experimental validation is available only for the four distal genes of the operon. Here, we report a transcriptional analysis of thespsoperon and a functional study performed by constructing and analyzing two null mutants lacking either all or only the promoter-proximal gene of the operon. Our results show that bothspsmutant spores apparently have normal coat and crust but have a small germination defect and are more hydrophobic than wild-type spores. We also show that spores lacking all Sps proteins are highly adhesive and form extensive clumps. In addition,spsmutant spores have an increased efficiency in adsorbing a heterologous enzyme, suggesting that hydrophobic force is a major determinant of spore adsorption and indicating that a deep understanding of the surface properties of the spore is essential for its full development as a surface display platform.

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Examination of the Genome-Wide Transcriptional Response of Escherichia coli O157:H7 to Cinnamaldehyde Exposure

Applied and Environmental Microbiology ◽

10.1128/aem.02767-12 ◽

2012 ◽

Vol 79 (3) ◽

pp. 942-950 ◽

Cited By ~ 33

Author(s):

Jeyachchandran Visvalingam ◽

Juan David Hernandez-Doria ◽

Richard A. Holley

Keyword(s):

Gene Expression ◽

Escherichia Coli ◽

Hplc Analysis ◽

Expression Profiles ◽

Transcriptional Analysis ◽

Content Type ◽

E Coli ◽

Rp Hplc ◽

Genome Wide ◽

Cinnamic Alcohol

ABSTRACTCinnamaldehyde is a natural antimicrobial that has been found to be effective against many food-borne pathogens, includingEscherichia coliO157:H7. Although its antimicrobial effects have been well investigated, limited information is available on its effects at the molecular level. Sublethal treatment at 200 mg/liter cinnamaldehyde inhibited growth ofE. coliO157:H7 at 37°C and for ≤2 h caused cell elongation, but from 2 to 4 h growth resumed and cells reverted to normal length. To understand this transient behavior, genome-wide transcriptional analysis ofE. coliO157:H7 was performed at 2 and 4 h of exposure to cinnamaldehyde in conjunction with reverse-phase high-performance liquid chromatography (RP-HPLC) analysis for cinnamaldehyde and other cinnamic compounds. Drastically different gene expression profiles were obtained at 2 and 4 h. RP-HPLC analysis showed that cinnamaldehyde was structurally stable for at least 2 h. At 2 h of exposure, cinnamaldehyde induced expression of many oxidative stress-related genes and repressed expression of DNA, protein, O-antigen, and fimbrial synthetic genes. At 4 h, many cinnamaldehyde-induced repressive effects onE. coliO157:H7 gene expression were reversed, and cells became more motile and grew at a slightly higher rate. Data indicated that by 4 h,E. coliO157:H7 was able to convert cinnamaldehyde into the less toxic cinnamic alcohol using dehydrogenase/reductase enzymes (YqhD and DkgA). This is the first study to characterize the ability ofE. coliO157:H7 to convert cinnamaldehyde into cinnamic alcohol which, in turn, showed that the antimicrobial activity of cinnamaldehyde is mainly attributable to its carbonyl aldehyde group.

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