scholarly journals Virus Evolution toward Limited Dependence on Nonessential Functions of the Host: the Case of Bacteriophage SPP1

2014 ◽  
Vol 89 (5) ◽  
pp. 2875-2883 ◽  
Author(s):  
Virginija Cvirkaite-Krupovic ◽  
Rut Carballido-López ◽  
Paulo Tavares
Keyword(s):  
Bacillus Subtilis ◽  
Host Cell ◽  
Exponential Growth ◽  
Receptor Protein ◽  
Exponential Growth Phase ◽  
Exact Nature ◽  
Lytic Bacteriophage ◽  
Content Type ◽  
Cell Functions ◽  
Bacteriophage Infection

ABSTRACTAll viruses are obligate intracellular parasites and depend on certain host cell functions for multiplication. However, the extent of such dependence and the exact nature of the functions provided by the host cell remain poorly understood. Here, we investigated if nonessentialBacillus subtilisgenes are necessary for multiplication of bacteriophage SPP1. Screening of a collection of 2,514 single-gene knockouts of nonessentialB. subtilisgenes yielded only a few genes necessary for efficient SPP1 propagation. Among these were genes belonging to theyukoperon, which codes for the Esat-6-like secretion system, including the SPP1 receptor protein YueB. In addition, we found that SPP1 multiplication was negatively affected by the absence of two other genes,putBandefp. The geneefpencodes elongation factor P, which enhances ribosome activity by alleviating translational stalling during the synthesis of polyproline-containing proteins. PutB is an enzyme involved in the proline degradation pathway that is required for infection in the post-exponential growth phase ofB. subtilis, when the bacterium undergoes a complex genetic reprogramming. TheputBknockout shortens significantly the window of opportunity for SPP1 infection during the host cell life cycle. This window is a critical parameter for competitive phage multiplication in the soil environment, whereB. subtilisrarely meets conditions for exponential growth. Our results in combination with those reported for other virus-host systems suggest that bacterial viruses have evolved toward limited dependence on nonessential host functions.IMPORTANCEA successful viral infection largely depends on the ability of the virus to hijack cellular machineries and to redirect the flow of building blocks and energy resources toward viral progeny production. However, the specific virus-host interactions underlying this fundamental transformation are poorly understood. Here, we report on the first systematic analysis of virus-host cross talk during bacteriophage infection in Gram-positive bacteria. We show that lytic bacteriophage SPP1 is remarkably independent of nonessential genes of its host,Bacillus subtilis, with only a few cellular genes being necessary for efficient phage propagation. We hypothesize that such limited dependence of the virus on its host results from a constant “evolutionary arms race” and might be much more widespread than currently thought.

scholarly journals The Presence of Conjugative Plasmid pLS20 Affects Global Transcription of Its Bacillus subtilis Host and Confers Beneficial Stress Resistance to Cells

2013 ◽  
Vol 80 (4) ◽  
pp. 1349-1358 ◽  
Author(s):  
Thomas C. Rösch ◽  
Wladislaw Golman ◽  
Laura Hucklesby ◽  
Jose E. Gonzalez-Pastor ◽  
Peter L. Graumann
Keyword(s):  
Bacillus Subtilis ◽  
Stationary Phase ◽  
Host Cell ◽  
Stress Resistance ◽  
Transcriptional Profiling ◽  
Conjugative Plasmid ◽  
Receptor Protein ◽  
Content Type ◽  
Phage Receptor ◽  
In Cells

ABSTRACTConjugation activity of plasmid pLS20 fromBacillus subtilissubsp.nattois induced when cells are diluted into fresh medium and diminishes as cells enter into stationary-phase growth. Transcriptional profiling shows that during mid-exponential growth, more than 5% of the host genes are affected in the presence of the plasmid, in contrast to the minor changes seen in freshly diluted and stationary-phase cells. Changes occurred in many metabolic pathways, although pLS20 does not confer any detectable burden on its host cell, as well as in membrane and cell wall-associated processes, in the large motility operon, and in several other cellular processes. In agreement with these changes, we found considerable alterations in motility and enzyme activity and increased resistance against several different forms of stress in cells containing the plasmid, revealing that the presence of pLS20 has a broad impact on the physiology of its host cell and increases its stress resistance in multiple aspects. Additionally, we found that the lack of chromosomal geneyueB, known to encode a phage receptor protein, which is upregulated in cells containing pLS20, strongly reduced conjugation efficiency, revealing that pLS20 not only increases fitness of its host but also employs host proteins for efficient transfer into a new cell.

scholarly journals Interplay of CodY and ScoC in the Regulation of Major Extracellular Protease Genes of Bacillus subtilis

2016 ◽  
Vol 198 (6) ◽  
pp. 907-920 ◽  
Author(s):  
Giulia Barbieri ◽  
Alessandra M. Albertini ◽  
Eugenio Ferrari ◽  
Abraham L. Sonenshein ◽  
Boris R. Belitsky
Keyword(s):  
Gene Expression ◽  
Bacillus Subtilis ◽  
Growth Phase ◽  
Exponential Growth ◽  
Degradation Products ◽  
Transcriptional Regulator ◽  
Complex Medium ◽  
Exponential Growth Phase ◽  
Content Type ◽  
In Cells

ABSTRACTAprE and NprE are two major extracellular proteases inBacillus subtiliswhose expression is directly regulated by several pleiotropic transcriptional factors, including AbrB, DegU, ScoC, and SinR. In cells growing in a rich, complex medium, theaprEandnprEgenes are strongly expressed only during the post-exponential growth phase; mutations in genes encoding the known regulators affect the level of post-exponential-phase gene expression but do not permit high-level expression during the exponential growth phase. Using DNA-binding assays and expression and mutational analyses, we have shown that the genes for both exoproteases are also under strong, direct, negative control by the global transcriptional regulator CodY. However, because CodY also repressesscoC, little or no derepression ofaprEandnprEwas seen in acodYnull mutant due to overexpression ofscoC. Thus, CodY is also an indirect positive regulator of these genes by limiting the synthesis of a second repressor. In addition, in cells growing under conditions that activate CodY, ascoCnull mutation had little effect onaprEornprEexpression; full effects ofscoCorcodYnull mutations could be seen only in the absence of the other regulator. However, even thecodY scoCdouble mutant did not show high levels ofaprEandnprEgene expression during exponential growth phase in a rich, complex medium. Only a third mutation, inabrB, allowed such expression. Thus, three repressors can contribute to reducing exoprotease gene expression during growth in the presence of excess nutrients.IMPORTANCEThe majorBacillus subtilisexoproteases, AprE and NprE, are important metabolic enzymes whose genes are subject to complex regulation by multiple transcription factors. We show here that expression of theaprEandnprEgenes is also controlled, both directly and indirectly, by CodY, a global transcriptional regulator that responds to the intracellular pools of amino acids. Direct CodY-mediated repression explains a long-standing puzzle, that is, why exoproteases are not produced when cells are growing exponentially in a medium containing abundant quantities of proteins or their degradation products. Indirect regulation ofaprEandnprEthrough CodY-mediated repression of thescoCgene, encoding another pleiotropic repressor, serves to maintain a significant level of repression of exoprotease genes when CodY loses activity.

scholarly journals Bacteriophage SP01 Gene Product 56 Inhibits Bacillus subtilis Cell Division by Interacting with FtsL and Disrupting Pbp2B and FtsW Recruitment

2020 ◽  
Vol 203 (2) ◽  
pp. e00463-20
Author(s):  
Amit Bhambhani ◽  
Isabella Iadicicco ◽  
Jules Lee ◽  
Syed Ahmed ◽  
Max Belfatto ◽  
...  
Keyword(s):  
Cell Wall ◽  
Bacillus Subtilis ◽  
Cell Division ◽  
Gene Product ◽  
Fluorescent Protein ◽  
Lytic Bacteriophage ◽  
Cell Wall Synthesis ◽  
Content Type ◽  
Ftsz Ring ◽  
Green Fluorescent

ABSTRACTPrevious work identified gene product 56 (gp56), encoded by the lytic bacteriophage SP01, as being responsible for inhibition of Bacillus subtilis cell division during its infection. Assembly of the essential tubulin-like protein FtsZ into a ring-shaped structure at the nascent site of cytokinesis determines the timing and position of division in most bacteria. This FtsZ ring serves as a scaffold for recruitment of other proteins into a mature division-competent structure permitting membrane constriction and septal cell wall synthesis. Here, we show that expression of the predicted 9.3-kDa gp56 of SP01 inhibits later stages of B. subtilis cell division without altering FtsZ ring assembly. Green fluorescent protein-tagged gp56 localizes to the membrane at the site of division. While its localization does not interfere with recruitment of early division proteins, gp56 interferes with the recruitment of late division proteins, including Pbp2b and FtsW. Imaging of cells with specific division components deleted or depleted and two-hybrid analyses suggest that gp56 localization and activity depend on its interaction with FtsL. Together, these data support a model in which gp56 interacts with a central part of the division machinery to disrupt late recruitment of the division proteins involved in septal cell wall synthesis.IMPORTANCE Studies over the past decades have identified bacteriophage-encoded factors that interfere with host cell shape or cytokinesis during viral infection. The phage factors causing cell filamentation that have been investigated to date all act by targeting FtsZ, the conserved prokaryotic tubulin homolog that composes the cytokinetic ring in most bacteria and some groups of archaea. However, the mechanisms of several phage factors that inhibit cytokinesis, including gp56 of bacteriophage SP01 of Bacillus subtilis, remain unexplored. Here, we show that, unlike other published examples of phage inhibition of cytokinesis, gp56 blocks B. subtilis cell division without targeting FtsZ. Rather, it utilizes the assembled FtsZ cytokinetic ring to localize to the division machinery and to block recruitment of proteins needed for septal cell wall synthesis.

scholarly journals Bacteriophage infection of Escherichia coli leads to the formation of membrane vesicles via both explosive cell lysis and membrane blebbing

Microbiology ◽  
2021 ◽  
Vol 167 (4) ◽  
Author(s):  
Pappu K. Mandal ◽  
Giulia Ballerin ◽  
Laura M. Nolan ◽  
Nicola K. Petty ◽  
Cynthia B. Whitchurch
Keyword(s):  
Escherichia Coli ◽  
Outer Membrane ◽  
Type Species ◽  
Bacteriophage T4 ◽  
Membrane Vesicles ◽  
Cell Lysis ◽  
Lytic Bacteriophage ◽  
Content Type ◽  
Link Type ◽  
Bacteriophage Infection

Membrane vesicles (MVs) are membrane-bound spherical nanostructures that prevail in all three domains of life. In Gram-negative bacteria, MVs are thought to be produced through blebbing of the outer membrane and are often referred to as outer membrane vesicles (OMVs). We have recently described another mechanism of MV formation in Pseudomonas aeruginosa that involves explosive cell-lysis events, which shatters cellular membranes into fragments that rapidly anneal into MVs. Interestingly, MVs are often observed within preparations of lytic bacteriophage, however the source of these MVs and their association with bacteriophage infection has not been explored. In this study we aimed to determine if MV formation is associated with lytic bacteriophage infection. Live super-resolution microscopy demonstrated that explosive cell lysis of Escherichia coli cells infected with either bacteriophage T4 or T7, resulted in the formation of MVs derived from shattered membrane fragments. Infection by either bacteriophage was also associated with the formation of membrane blebs on intact bacteria. TEM revealed multiple classes of MVs within phage lysates, consistent with multiple mechanisms of MV formation. These findings suggest that bacteriophage infection may be a major contributor to the abundance of bacterial MVs in nature.

scholarly journals Neighbors Working Together: a Toxoplasma Rhoptry Protein That Facilitates Dense Granule Protein Translocation into the Host Cell

mSphere ◽  
2019 ◽  
Vol 4 (4) ◽  
Author(s):  
Gustavo Arrizabalaga
Keyword(s):  
Host Cell ◽  
Protein Translocation ◽  
Parasitophorous Vacuole ◽  
Opportunistic Pathogen ◽  
Dense Granule ◽  
Effector Proteins ◽  
Content Type ◽  
Dense Granules ◽  
Cell Functions ◽  
Host Signaling

ABSTRACT The opportunistic pathogen Toxoplasma gondii is highly adept at manipulating host cell functions. While inside a host cell, Toxoplasma divides within a parasitophorous vacuole from which it secretes numerous effector proteins from its dense granules. Many of these so-called GRA proteins are translocated from the parsitophorous vacuole into the host cell where they directly disrupt host signaling pathways. The machinery that drives the translocation of GRA proteins across the parasitophorous vacuole membrane is being elucidated through both genetic and biochemical approaches. A new mSphere research article (M. W. Panas, A. Ferrel, A. Naor, E. Tenborg, et al., mSphere 4:e00276-19, 2019, https://doi.org/10.1128/mSphere.00276-19) describes how the kinase ROP17, which is secreted from the parasite’s rhoptries into the host cell during invasion, regulates the translocation of GRA effectors.

Induction of autolysis in Bacillus subtilis by ochratoxin A

1978 ◽  
Vol 24 (5) ◽  
pp. 563-568 ◽  
Author(s):  
U. Singer ◽  
R. Röschenthaler
Keyword(s):  
Cell Wall ◽  
Protein Synthesis ◽  
Bacillus Subtilis ◽  
Ochratoxin A ◽  
Optical Density ◽  
Growth Phase ◽  
Exponential Growth ◽  
Rna Synthesis ◽  
Exponential Growth Phase ◽  
Cell Wall Synthesis

Ochratoxin A (OTA) added during the exponential growth phase at a concentration higher than 12 μg/ml caused autolysis of Bacillus subtilis. Optical density of cultures decreased, and at higher concentrations the cultures became sterile. Optimum OTA-induced lysis was about pH 5. At concentrations below 10 μg/ml, protein synthesis was inhibited more strongly than RNA synthesis. Cell wall synthesis was also strongly inhibited. A fraction extracted from the lysates had the property of a lysis inhibitor. The relevance of this fraction in respect to autolysis is discussed.

scholarly journals Net Production and Consumption of Fluorescent Colored Dissolved Organic Matter by Natural Bacterial Assemblages Growing on Marine Phytoplankton Exudates

2011 ◽  
Vol 77 (21) ◽  
pp. 7490-7498 ◽  
Author(s):  
Cristina Romera-Castillo ◽  
Hugo Sarmento ◽  
Xosé Antón Álvarez-Salgado ◽  
Josep M. Gasol ◽  
Celia Marrasé
Keyword(s):  
Organic Matter ◽  
Dissolved Organic Matter ◽  
Growth Phase ◽  
Exponential Growth ◽  
Prorocentrum Minimum ◽  
Marine Phytoplankton ◽  
Exponential Growth Phase ◽  
Phytoplankton Species ◽  
Colored Dissolved Organic Matter ◽  
Content Type

ABSTRACTAn understanding of the distribution of colored dissolved organic matter (CDOM) in the oceans and its role in the global carbon cycle requires a better knowledge of the colored materials produced and consumed by marine phytoplankton and bacteria. In this work, we examined the net uptake and release of CDOM by a natural bacterial community growing on DOM derived from four phytoplankton species cultured under axenic conditions. Fluorescent humic-like substances exuded by phytoplankton (excitation/emission [Ex/Em] wavelength, 310 nm/392 nm; Coble's peak M) were utilized by bacteria in different proportions depending on the phytoplankton species of origin. Furthermore, bacteria produced humic-like substances that fluoresce at an Ex/Em wavelength of 340 nm/440 nm (Coble's peak C). Differences were also observed in the Ex/Em wavelengths of the protein-like materials (Coble's peak T) produced by phytoplankton and bacteria. The induced fluorescent emission of CDOM produced by prokaryotes was an order of magnitude higher than that of CDOM produced by eukaryotes. We have also examined the final compositions of the bacterial communities growing on the exudates, which differed markedly depending on the phytoplankton species of origin.AlteromonasandRoseobacterwere dominant during all the incubations onChaetocerossp. andProrocentrum minimumexudates, respectively.Alteromonaswas the dominant group growing onSkeletonema costatumexudates during the exponential growth phase, but it was replaced byRoseobacterafterwards. OnMicromonas pusillaexudates,Roseobacterwas replaced byBacteroidetesafter the exponential growth phase. Our work shows that fluorescence excitation-emission matrices of CDOM can be a helpful tool for the identification of microbial sources of DOM in the marine environment, but further studies are necessary to explore the association of particular bacterial groups with specific fluorophores.

scholarly journals CcpA-Independent Regulation of Expression of the Mg2+-Citrate Transporter Gene citM by Arginine Metabolism in Bacillus subtilis

2003 ◽  
Vol 185 (3) ◽  
pp. 854-859 ◽  
Author(s):  
Jessica B. Warner ◽  
Christian Magni ◽  
Juke S. Lolkema
Keyword(s):  
Bacillus Subtilis ◽  
Growth Phase ◽  
Exponential Growth ◽  
Stationary Growth Phase ◽  
Luria Bertani ◽  
Transition Phase ◽  
Exponential Growth Phase ◽  
Uptake System ◽  
Regulation Of Expression ◽  
Citrate Transporter

ABSTRACT Transcriptional regulation of the Mg2+-citrate transporter, CitM, the main citrate uptake system of Bacillus subtilis, was studied during growth in rich medium. Citrate in the growth medium was required for induction under all growth conditions. In Luria-Bertani medium containing citrate, citM expression was completely repressed during the exponential growth phase, marginally expressed in the transition phase, and highly expressed in the stationary growth phase. The repression was relieved when the cells were grown in spent Luria-Bertani medium. The addition of a mixture of 18 amino acids restored repression. l-Arginine in the mixture appeared to be solely responsible for the repression, and ornithine appeared to be an equally potent repressor of citM expression. Studies of mutant strains deficient in RocR and SigL, proteins required for the expression of the enzymes of the arginase pathway, confirmed that uptake into the cell and, most likely, conversion of arginine to ornithine were required for repression. Arginine-mediated repression was independent of a functional CcpA, the global regulator protein in carbon catabolite repression (CCR). Nevertheless, CCR-mediated repression was the major mechanism controlling the expression during exponential growth, while the newly described, CcpA-independent arginine-mediated repression was specifically apparent during the transition phase of growth.

scholarly journals Infection by Toxoplasma gondii Specifically Induces Host c-Myc and the Genes This Pivotal Transcription Factor Regulates

2014 ◽  
Vol 13 (4) ◽  
pp. 483-493 ◽  
Author(s):  
Magdalena Franco ◽  
Anjali J. Shastri ◽  
John C. Boothroyd
Keyword(s):  
Transcription Factor ◽  
Toxoplasma Gondii ◽  
Host Cell ◽  
Host Response ◽  
Neospora Caninum ◽  
Apicomplexan Parasite ◽  
Intracellular Growth ◽  
Terminal Protein ◽  
Content Type ◽  
Cell Functions

ABSTRACT Toxoplasma gondii infection has previously been described to cause dramatic changes in the host transcriptome by manipulating key regulators, including STATs, NF-κB, and microRNAs. Here, we report that Toxoplasma tachyzoites also mediate rapid and sustained induction of another pivotal regulator of host cell transcription, c-Myc. This induction is seen in cells infected with all three canonical types of Toxoplasma but not the closely related apicomplexan parasite Neospora caninum . Coinfection of cells with both Toxoplasma and Neospora still results in an increase in the level of host c-Myc, showing that c-Myc is actively upregulated by Toxoplasma infection (rather than repressed by Neospora ). We further demonstrate that this upregulation may be mediated through c-Jun N-terminal protein kinase (JNK) and is unlikely to be a nonspecific host response, as heat-killed Toxoplasma parasites do not induce this increase and neither do nonviable parasites inside the host cell. Finally, we show that the induced c-Myc is active and that transcripts dependent on its function are upregulated, as predicted. Hence, c-Myc represents an additional way in which Toxoplasma tachyzoites have evolved to specifically alter host cell functions during intracellular growth.

scholarly journals Orientia tsutsugamushi Strain Ikeda Ankyrin Repeat-Containing Proteins Recruit SCF1 Ubiquitin Ligase Machinery via Poxvirus-Like F-Box Motifs

2015 ◽  
Vol 197 (19) ◽  
pp. 3097-3109 ◽  
Author(s):  
Andrea R. Beyer ◽  
Lauren VieBrock ◽  
Kyle G. Rodino ◽  
Daniel P. Miller ◽  
Brittney K. Tegels ◽  
...  
Keyword(s):  
Host Cell ◽  
Ubiquitin Ligase ◽  
Scrub Typhus ◽  
Ankyrin Repeat ◽  
Intracellular Bacterium ◽  
Host Cells ◽  
Orientia Tsutsugamushi ◽  
Ligand Interaction ◽  
Content Type ◽  
Cell Functions

ABSTRACTA rising theme among intracellular microbes is the delivery of ankyrin repeat-containing effectors (Anks) that interact with target proteins to co-opt host cell functions.Orientia tsutsugamushi, an obligate intracellular bacterium and the etiologic agent of scrub typhus, encodes one of the largest Ank repertoires of any sequenced microorganism. They have been previously identified as type 1 secretion system substrates. Here,in silicoand manual sequence analyses revealed that a large proportion ofO. tsutsugamushistrain Ikeda Anks bear a eukaryotic/poxvirus-like F-box motif, which is known to recruit host cell SCF1 ubiquitin ligase machinery. We assessed the Anks for the ability to serve as F-box proteins. Coimmunoprecipitation assays demonstrated that F-box-containing Anks interact with overexpressed and/or endogenous SCF1 components. When coexpressed with FLAG-Ank4_01 or FLAG-Ank9, a glutathioneS-transferase (GST)-tagged version of the SCF1 component SKP1 localized to subcellular sites of FLAG-Ank accumulation. The abilities of recombinant Anks to interact and colocalize with SKP1 were F-box dependent. GST-SKP1 precipitatedO. tsutsugamushi-derived Ank9 from infected host cells, verifying both that the pathogen expresses Ank9 during infection and the protein's capability to bind SKP1. AligningO. tsutsugamushi, poxviral, and eukaryotic F-box sequences delineated three F-box residues that are highly conserved and likely to be functionally important. Substitution of these residues ablated the ability of GFP-Ank9 to interact with GST-SKP1. These results demonstrate thatO. tsutsugamushistrain Ikeda Anks can co-opt host cell polyubiquitination machinery, provide the first evidence that anO. tsutsugamushiAnk does so during infection, and advance overall understanding of microbial F-box proteins.IMPORTANCEAnkyrin repeat-containing proteins (Anks) are important virulence factors of intracellular bacteria that mediate protein-protein interactions with host cell targets.Orientia tsutsugamushi, which causes a debilitating infection called scrub typhus in one of the most densely populated regions of the world, encodes one of the largest Ank armamentariums of any sequenced bacterium. This study demonstrates thatO. tsutsugamushistrain Ikeda Anks also bear F-box motifs that interact with host cell polyubiquitination machinery. By proving that anOrientia-derived Ank interacts with SKP1 in infected cells, this evidences the first bona fideOrientiaeffector and the first example of an endogenous F-box-containing Ank–mammalian-host ligand interaction for any intracellular bacterium. Also, importantly, this work identifies key residues that are essential for microbial F-box function.

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