scholarly journals The Hybrid Sensor Kinase RscS Integrates Positive and Negative Signals To Modulate Biofilm Formation in Vibrio fischeri

2008 ◽  
Vol 190 (13) ◽  
pp. 4437-4446 ◽  
Author(s):  
Kati Geszvain ◽  
Karen L. Visick
Keyword(s):  
Biofilm Formation ◽  
Vibrio Fischeri ◽  
Transmembrane Helix ◽  
Bioinformatic Analysis ◽  
Colony Morphology ◽  
Sensor Kinase ◽  
Pas Domain ◽  
Transmembrane Helices ◽  
Pellicle Formation ◽  
Carboxy Terminal

ABSTRACT Overexpression of the Vibrio fischeri sensor kinase RscS induces expression of the syp (symbiosis polysaccharide) gene cluster and promotes biofilm phenotypes such as wrinkled colony morphology, pellicle formation, and surface adherence. RscS is predicted to be a hybrid sensor kinase with a histidine kinase/ATPase (HATPase) domain, a receiver (Rec) domain, and a histidine phosphotransferase (Hpt) domain. Bioinformatic analysis also revealed the following three potential signal detection domains within RscS: two transmembrane helices forming a transmembrane region (TMR), a large periplasmic (PP) domain, and a cytoplasmic PAS domain. In this work, we genetically dissected the contributions of these domains to RscS function. Substitutions within the carboxy-terminal domain supported identification of RscS as a hybrid sensor kinase; disruption of both the HATPase and Rec domains eliminated induction of syp transcription, wrinkled colony morphology, pellicle formation, and surface adherence, while disruption of Hpt resulted in decreased activity. The PAS domain was also critical for RscS activity; substitutions in PAS resulted in a loss of activity. Generation of a cytoplasmic, N-terminal deletion derivative of RscS resulted in a partial loss of activity, suggesting a role for localization to the membrane and/or sequences within the TMR and PP domain. Finally, substitutions within the first transmembrane helix of the TMR and deletions within the PP domain both resulted in increased activity. Thus, RscS integrates both inhibitory and stimulatory signals from the environment to regulate biofilm formation by V. fischeri.

scholarly journals The Putative Hybrid Sensor Kinase SypF Coordinates Biofilm Formation in Vibrio fischeri by Acting Upstream of Two Response Regulators, SypG and VpsR

2008 ◽  
Vol 190 (14) ◽  
pp. 4941-4950 ◽  
Author(s):  
Cynthia L. Darnell ◽  
Elizabeth A. Hussa ◽  
Karen L. Visick
Keyword(s):  
Biofilm Formation ◽  
Vibrio Fischeri ◽  
Response Regulator ◽  
Cell Aggregation ◽  
Putative Hybrid ◽  
Sensor Kinase ◽  
Gene Encoding ◽  
Subsequent Work ◽  
Pellicle Formation ◽  
Dependent Cell

ABSTRACT Colonization of the Hawaiian squid Euprymna scolopes by the marine bacterium Vibrio fischeri requires the symbiosis polysaccharide (syp) gene cluster, which contributes to symbiotic initiation by promoting biofilm formation on the surface of the symbiotic organ. We previously described roles for the syp-encoded response regulator SypG and an unlinked gene encoding the sensor kinase RscS in controlling syp transcription and inducing syp-dependent cell-cell aggregation phenotypes. Here, we report the involvement of an additional syp-encoded regulator, the putative sensor kinase SypF, in promoting biofilm formation. Through the isolation of an increased activity allele, sypF1, we determined that SypF can function to induce syp transcription as well as a variety of biofilm phenotypes, including wrinkled colony formation, adherence to glass, and pellicle formation. SypF1-mediated transcription of the syp cluster was entirely dependent on SypG. However, the biofilm phenotypes were reduced, not eliminated, in the sypG mutant. These phenotypes were also reduced in a mutant deleted for sypE, another syp-encoded response regulator. However, SypF1 still induced phenotypes in a sypG sypE double mutant, suggesting that SypF1 might activate another regulator(s). Our subsequent work revealed that the residual SypF1-induced biofilm formation depended on VpsR, a putative response regulator, and cellulose biosynthesis. These data support a model in which a network of regulators and at least two polysaccharide loci contribute to biofilm formation in V. fischeri.

scholarly journals Arabinose Induces Pellicle Formation by Vibrio fischeri

2013 ◽  
Vol 79 (6) ◽  
pp. 2069-2080 ◽  
Author(s):  
Karen L. Visick ◽  
Kevin P. Quirke ◽  
Sheila M. McEwen
Keyword(s):  
Biofilm Formation ◽  
Sole Carbon Source ◽  
Marine Bacterium ◽  
Vibrio Fischeri ◽  
Luria Bertani ◽  
Phenol Red ◽  
Liquid Interfaces ◽  
Acid Base ◽  
Pellicle Formation ◽  
Content Type

ABSTRACTBiofilms are multicellular communities of bacteria attached to a surface and embedded in a protective matrix. In many cases, the signals that induce biofilm formation are unknown. Here, we report that biofilm formation by the marine bacteriumVibrio fischerican be induced by the addition of arabinose to LBS (Luria-Bertani-salt), a tryptone-based medium. Growth of cells in the presence of 0.2% arabinose, but not other sugars, induced the production of a pellicle at the air/liquid interfaces of static cultures.V. fischerifailed to grow on arabinose as the sole carbon source, suggesting that pellicle production did not occur as a result of increased growth, but experiments using the acid/base indicator phenol red suggested thatV. fischerimay partially metabolize arabinose. Pellicle production was independent of thesyppolysaccharide locus but was altered upon disruption of thebcscellulose locus. Through a screen for mutants defective for pellicle production, we found that loss of motility disrupted the formation of the arabinose-induced pellicle. Among the ∼20 mutants that retained motility were strains with insertions in a putativemshpilus locus and a strain with a defect inyidK, which is involved in galactose catabolism. Mutants with themshgene disrupted grew poorly in the presence of arabinose, while theyidKmutant appeared to be “blind” to the presence of arabinose. Finally, arabinose impaired symbiotic colonization byV. fischeri. This work thus identifies a novel signal and new pathways involved in control of biofilm formation byV. fischeri.

scholarly journals LitR Is a Repressor of syp Genes and Has a Temperature-Sensitive Regulatory Effect on Biofilm Formation and Colony Morphology in Vibrio (Aliivibrio) salmonicida

2014 ◽  
Vol 80 (17) ◽  
pp. 5530-5541 ◽  
Author(s):  
Hilde Hansen ◽  
Ane Mohn Bjelland ◽  
Maria Ronessen ◽  
Espen Robertsen ◽  
Nils Peder Willassen
Keyword(s):  
Biofilm Formation ◽  
Cold Water ◽  
Vibrio Fischeri ◽  
Seawater Temperature ◽  
Colony Morphology ◽  
Effective Vaccine ◽  
Temperature Sensitive ◽  
Regulatory Effect ◽  
Content Type ◽  
Fish Model

ABSTRACTVibrio(Aliivibrio)salmonicidais the etiological agent of cold water vibriosis, a disease in farmed Atlantic salmon (Salmo salar) that is kept under control due to an effective vaccine. A seawater temperature below 12°C is normally required for disease development. Quorum sensing (QS) is a cell density-regulated communication system that bacteria use to coordinate activities involved in colonization and pathogenesis, and we have previously shown that inactivation of the QS master regulator LitR attenuates theV. salmonicidastrain LFI1238 in a fish model. We show here that strain LFI1238 and a panel of naturally occurringV. salmonicidastrains are poor biofilm producers. Inactivation oflitRin the LFI1238 strain enhances medium- and temperature-dependent adhesion, rugose colony morphology, and biofilm formation. Chemical treatment and electron microscopy of the biofilm identified an extracellular matrix consisting mainly of a fibrous network, proteins, and polysaccharides. Further, by microarray analysis of planktonic and biofilm cells, we identified a number of genes regulated by LitR and, among these, were homologues of theVibrio fischerisymbiosis polysaccharide (syp) genes. Thesypgenes were regulated by LitR in both planktonic and biofilm lifestyle analyses. Disruption ofsypgenes in theV. salmonicidaΔlitRmutant alleviated adhesion, rugose colony morphology, and biofilm formation. Hence, LitR is a repressor ofsyptranscription that is necessary for expression of the phenotypes examined. The regulatory effect of LitR on colony morphology and biofilm formation is temperature sensitive and weak or absent at temperatures above the bacterium's upper threshold for pathogenicity.

scholarly journals Burkholderia cenocepacia ShvR-Regulated Genes That Influence Colony Morphology, Biofilm Formation, and Virulence

2011 ◽  
Vol 79 (8) ◽  
pp. 2984-2997 ◽  
Author(s):  
Sujatha Subramoni ◽  
David T. Nguyen ◽  
Pamela A. Sokol
Keyword(s):  
Biofilm Formation ◽  
Citrate Synthase ◽  
Tricarboxylic Acid Cycle ◽  
Functional Characterization ◽  
Burkholderia Cenocepacia ◽  
Colony Morphology ◽  
Appreciable Effect ◽  
Pellicle Formation ◽  
Content Type ◽  
Branched Chain

ABSTRACTBurkholderia cenocepaciais an opportunistic pathogen that primarily infects cystic fibrosis (CF) patients. Previously, we reported that ShvR, a LysR regulator, influences colony morphology, virulence, and biofilm formation and regulates the expression of an adjacent 24-kb genomic region encoding 24 genes. In this study, we report the functional characterization of selected genes in this region. A Tn5mutant with shiny colony morphology was identified with a polar mutation in BCAS0208, predicted to encode an acyl-coenzyme A dehydrogenase. Mutagenesis of BCAS0208 and complementation analyses revealed that BCAS0208 is required for rough colony morphology, biofilm formation, and virulence on alfalfa seedlings. It was not possible to complement with BCAS0208 containing a mutation in the catalytic site. BCAS0201, encoding a putative flavin adenine dinucleotide (FAD)-dependent oxidoreductase, and BCAS0207, encoding a putative citrate synthase, do not influence colony morphology but are required for optimum levels of biofilm formation and virulence. Both BCAS0208 and BCAS0201 contribute to pellicle formation, although individual mutations in each of these genes had no appreciable effect on pellicle formation. A mutant with a polar insertion in BCAS0208 was significantly less virulent in a rat model of chronic lung infection as well as in the alfalfa model. Genes in this region were shown to influence utilization of branched-chain fatty acids, tricarboxylic acid cycle substrates,l-arabinose, and branched-chain amino acids. Together, our data show that the ShvR-regulated genes BCAS0208 to BCAS0201 are required for the rough colony morphotype, biofilm and pellicle formation, and virulence inB. cenocepacia.

scholarly journals Discovery of Calcium as a Biofilm-Promoting Signal for Vibrio fischeri Reveals New Phenotypes and Underlying Regulatory Complexity

2018 ◽  
Vol 200 (15) ◽  
Author(s):  
Alice H. Tischler ◽  
Louise Lie ◽  
Cecilia M. Thompson ◽  
Karen L. Visick
Keyword(s):  
Biofilm Formation ◽  
Vibrio Fischeri ◽  
Response Regulator ◽  
Negative Regulator ◽  
Regulatory Control ◽  
Cellulose Synthesis ◽  
Sensor Kinase ◽  
Content Type ◽  
First Time ◽  
Symbiosis Polysaccharide

ABSTRACT Vibrio fischeri uses biofilm formation to promote symbiotic colonization of its squid host, Euprymna scolopes. Control over biofilm formation is exerted at the level of transcription of the symbiosis polysaccharide (syp) locus by a complex set of two-component regulators. Biofilm formation can be induced by overproduction of the sensor kinase RscS, which requires the activities of the hybrid sensor kinase SypF and the response regulator SypG and is negatively regulated by the sensor kinase BinK. Here, we identify calcium as a signal that promotes biofilm formation by biofilm-competent strains under conditions in which biofilms are not typically observed (growth with shaking). This was true for RscS-overproducing cells as well as for strains in which only the negative regulator binK was deleted. The latter results provided, for the first time, an opportunity to induce and evaluate biofilm formation without regulator overexpression. Using these conditions, we determined that calcium induces both syp-dependent and bacterial cellulose synthesis (bcs)-dependent biofilms at the level of transcription of these loci. The calcium-induced biofilms were dependent on SypF, but SypF's Hpt domain was sufficient for biofilm formation. These data suggested the involvement of another sensor kinase(s) and led to the discovery that both RscS and a previously uncharacterized sensor kinase, HahK, functioned in this pathway. Together, the data presented here reveal both a new signal and biofilm phenotype produced by V. fischeri cells, the coordinate production of two polysaccharides involved in distinct biofilm behaviors, and a new regulator that contributes to control over these processes. IMPORTANCE Biofilms, or communities of surface-attached microorganisms adherent via a matrix that typically includes polysaccharides, are highly resistant to environmental stresses and are thus problematic in the clinic and important to study. Vibrio fischeri forms biofilms to colonize its symbiotic host, making this organism useful for studying biofilms. Biofilm formation depends on the syp polysaccharide locus and its regulators. Here, we identify a signal, calcium, that induces both SYP-PS and cellulose-dependent biofilms. We also identify a new syp regulator, the sensor kinase HahK, and discover a mutant phenotype for the sensor kinase RscS. This work thus reveals a specific biofilm-inducing signal that coordinately controls two polysaccharides, identifies a new regulator, and clarifies the regulatory control over biofilm formation by V. fischeri.

scholarly journals Extracellular DNA of slow growers of mycobacteria and its contribution to biofilm formation and drug tolerance

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Aleksandr Ilinov ◽  
Akihito Nishiyama ◽  
Hiroki Namba ◽  
Yukari Fukushima ◽  
Hayato Takihara ◽  
...  
Keyword(s):  
Biofilm Formation ◽  
Drug Tolerance ◽  
Extracellular Dna ◽  
Exponential Growth Phase ◽  
Human Pathogens ◽  
Pellicle Formation ◽  
Phage Dna ◽  
Physiological Impact ◽  
Slow Growing ◽  
Growth And Reproduction

AbstractDNA is basically an intracellular molecule that stores genetic information and carries instructions for growth and reproduction in all cellular organisms. However, in some bacteria, DNA has additional roles outside the cells as extracellular DNA (eDNA), which is an essential component of biofilm formation and hence antibiotic tolerance. Mycobacteria include life-threating human pathogens, most of which are slow growers. However, little is known about the nature of pathogenic mycobacteria’s eDNA. Here we found that eDNA is present in slow-growing mycobacterial pathogens, such as Mycobacterium tuberculosis, M. intracellulare, and M. avium at exponential growth phase. In contrast, eDNA is little in all tested rapid-growing mycobacteria. The physiological impact of disrupted eDNA on slow-growing mycobacteria include reduced pellicle formation, floating biofilm, and enhanced susceptibility to isoniazid and amikacin. Isolation and sequencing of eDNA revealed that it is identical to the genomic DNA in M. tuberculosis and M. intracellulare. In contrast, accumulation of phage DNA in eDNA of M. avium, suggests that the DNA released differs among mycobacterial species. Our data show important functions of eDNA necessary for biofilm formation and drug tolerance in slow-growing mycobacteria.

scholarly journals Point Mutations in Transmembrane Helices 2 and 3 of ExbB and TolQ Affect Their Activities in Escherichia coli K-12

2004 ◽  
Vol 186 (13) ◽  
pp. 4402-4406 ◽  
Author(s):  
Volkmar Braun ◽  
Christina Herrmann
Keyword(s):  
Escherichia Coli ◽  
Amino Acid ◽  
Transmembrane Helix ◽  
Point Mutations ◽  
Transmembrane Helices ◽  
The Third ◽  
Form Part ◽  
K 12

ABSTRACT Replacement of glutamate 176, the only charged amino acid in the third transmembrane helix of ExbB, with alanine (E176A) abolished ExbB activity in all determined ExbB-dependent functions of Escherichia coli. Combination of the mutations T148A in the second transmembrane helix and T181A in the third transmembrane helix, proposed to form part of a proton pathway through ExbB, also resulted in inactive ExbB. E176 and T148 are strictly conserved in ExbB and TolQ proteins, and T181 is almost strictly conserved in ExbB, TolQ, and MotA.

scholarly journals FOLDING ELASTIC TRANSMEMBRANE HELICES TO FIT IN A LOW-RESOLUTION IMAGE BY ELECTRON MICROSCOPY

2011 ◽  
Vol 09 (supp01) ◽  
pp. 37-50 ◽  
Author(s):  
YUTAKA UENO ◽  
KAZUNORI KAWASAKI ◽  
OSAMU SAITO ◽  
MASAFUMI ARAI ◽  
MAKIKO SUWA
Keyword(s):  
Membrane Proteins ◽  
Structure Prediction ◽  
Molecular Model ◽  
Imaging Techniques ◽  
Transmembrane Helix ◽  
Prediction Method ◽  
Molecular Shape ◽  
Coarse Grained ◽  
Transmembrane Helices ◽  
Low Resolution

Structure prediction of membrane proteins could be constrained and thereby improved by introducing data of the observed molecular shape. We studied a coarse-grained molecular model that relied on residue-based dummy atoms to fold the transmembrane helices of a protein in the observed molecular shape. Based on the inter-residue potential, the α-helices were folded to contact each other in a simulated annealing protocol to search optimized conformation. Fitting the model into a three-dimensional volume was tested for proteins with known structures and resulted in a fairly reasonable arrangement of helices. In addition, the constraint to the packing transmembrane helix with the two-dimensional region was tested and found to work as a very similar folding guide. The obtained models nicely represented α-helices with the desired slight bend. Our structure prediction method for membrane proteins well demonstrated reasonable folding results using a low-resolution structural constraint introduced from recent cell-surface imaging techniques.

Crystal structure of an Lrs14-like archaeal biofilm regulator fromSulfolobus acidocaldarius

2018 ◽  
Vol 74 (11) ◽  
pp. 1105-1114
Author(s):  
Marian S. Vogt ◽  
Simon L. Völpel ◽  
Sonja-Verena Albers ◽  
Lars-Oliver Essen ◽  
Ankan Banerjee
Keyword(s):  
Crystal Structure ◽  
Biofilm Formation ◽  
Coiled Coil ◽  
Transcriptional Regulators ◽  
Bioinformatic Analysis ◽  
Considerable Degree ◽  
Sulfolobus Acidocaldarius ◽  
Asymmetric Unit ◽  
Winged Helix ◽  
Structural Differences

The small winged helix–turn–helix (wHTH) proteins of the Lrs14 family are major transcriptional regulators and act as archaeal biofilm regulators (AbfRs) in the crenarchaeoteSulfolobus acidocaldarius. Here, the first crystal structure of an AbfR ortholog, AbfR2, the deletion of which is known to impair biofilm formation, is presented. Like most other wHTH orthologs, AbfR2 is dimeric in solution as well as in its 2.45 Å resolution crystal structure. Given the presence of three independent AbfR2 dimers in the asymmetric unit, the crystal structure shows a considerable degree of conformational variation within the dimer, the antiparallel orientations of which are stabilized by coiled-coil interaction between H4 helices. Conserved anchor interactions between helices H0 and H4 of AbfR2 further contribute to dimer stabilization. The combined structural and bioinformatic analysis reveals cluster-specific structural differences between different members of the Lrs14 protein family.

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