Volatile Anesthetics Affect Nutrient Availability in Yeast

Genetics ◽  
2002 ◽  
Vol 161 (2) ◽  
pp. 563-574
Author(s):  
Laura K Palmer ◽  
Darren Wolfe ◽  
Jessica L Keeley ◽  
Ralph L Keil
Keyword(s):  
Amino Acids ◽  
Nutrient Availability ◽  
Wild Type Strain ◽  
Volatile Anesthetic ◽  
Volatile Anesthetics ◽  
Wild Type ◽  
Anesthetic Exposure ◽  
Sites Of Action ◽  
Insight Into ◽  
Lines Of Evidence

Abstract Volatile anesthetics affect all cells and tissues tested, but their mechanisms and sites of action remain unknown. To gain insight into the cellular activities of anesthetics, we have isolated genes that, when overexpressed, render Saccharomyces cerevisiae resistant to the volatile anesthetic isoflurane. One of these genes, WAK3/TAT1, encodes a permease that transports amino acids including leucine and tryptophan, for which our wild-type strain is auxotrophic. This suggests that availability of amino acids may play a key role in anesthetic response. Multiple lines of evidence support this proposal: (i) Deletion or overexpression of permeases that transport leucine and/or tryptophan alters anesthetic response; (ii) prototrophic strains are anesthetic resistant; (iii) altered concentrations of leucine and tryptophan in the medium affect anesthetic response; and (iv) uptake of leucine and tryptophan is inhibited during anesthetic exposure. Not all amino acids are critical for this response since we find that overexpression of the lysine permease does not affect anesthetic sensitivity. These findings are consistent with models in which anesthetics have a physiologically important effect on availability of specific amino acids by altering function of their permeases. In addition, we show that there is a relationship between nutrient availability and ubiquitin metabolism in this response.

scholarly journals Influence of Chemotaxis and Swimming Patterns on the Virulence of the Coral PathogenVibrio coralliilyticus

2018 ◽  
Vol 200 (15) ◽  
Author(s):  
Blake Ushijima ◽  
Claudia C. Häse
Keyword(s):  
Type Strain ◽  
Wild Type Strain ◽  
Bacterial Colonization ◽  
Valuable Insight ◽  
Wild Type ◽  
Coral Mucus ◽  
Content Type ◽  
Swimming Pattern ◽  
Vibrio Coralliilyticus ◽  
Insight Into

ABSTRACTChemotaxis, the directed movement toward or away from a chemical signal, can be essential to bacterial pathogens for locating hosts or avoiding hostile environments. The coral pathogenVibrio coralliilyticuschemotaxes toward coral mucus; however, chemotaxis has not been experimentally demonstrated to be important for virulence. To further examine this, in-frame mutations were constructed in genes predicted to be important forV. coralliilyticuschemotaxis. MostVibriogenomes contain multiple homologs of various chemotaxis-related genes, and two paralogs of each forcheB,cheR, andcheAwere identified. Based on single mutant analyses, the paralogscheB2,cheR2, andcheA1were essential for chemotaxis in laboratory assays. As predicted, the ΔcheA1and ΔcheR2strains had a smooth-swimming pattern, while the ΔcheB2strain displayed a zigzag pattern when observed under light microscopy. However, these mutants, unlike the parent strain, were unable to chemotax toward the known attractants coral mucus, dimethylsulfoniopropionate, andN-acetyl-d-glucosamine. The ΔcheB2strain and an aflagellate ΔfliG1strain were avirulent to coral, while the ΔcheA1and ΔcheR2strains were hypervirulent (90 to 100% infection within 14 h on average) compared to the wild-type strain (66% infection within 36 h on average). Additionally, the ΔcheA1and ΔcheR2strains appeared to better colonize coral fragments than the wild-type strain. These results suggest that although chemotaxis may be involved with infection (the ΔcheB2strain was avirulent), a smooth-swimming phenotype is important for bacterial colonization and infection. This study provides valuable insight into understandingV. coralliilyticuspathogenesis and how this pathogen may be transmitted between hosts.IMPORTANCECorals are responsible for creating the immense structures that are essential to reef ecosystems; unfortunately, pathogens like the bacteriumVibrio coralliilyticuscan cause fatal infections of reef-building coral species. However, compared to related human pathogens, the mechanisms by whichV. coralliilyticusinitiates infections and locates new coral hosts are poorly understood. This study investigated the effects of chemotaxis, the directional swimming in response to chemical signals, and bacterial swimming patterns on infection of the coralMontipora capitata. Infection experiments with different mutant strains suggested that a smooth-swimming pattern resulted in hypervirulence. These results demonstrate that the role of chemotaxis in coral infection may not be as straightforward as previously hypothesized and provide valuable insight intoV. coralliilyticuspathogenesis.

Comparison of the PR mutant with the wild-type strain of Proteus mirabilis brings insight into peroxide resistance factors and regulation of catalase expression

2001 ◽  
Vol 47 (2) ◽  
pp. 130-138 ◽  
Author(s):  
Pierre Andreoletti ◽  
Bruno Franzetti ◽  
Laurent Nussaume ◽  
Jean-Pierre Andrieu ◽  
Jean Gagnon ◽  
...  
Keyword(s):  
Proteus Mirabilis ◽  
Type Strain ◽  
Wild Type Strain ◽  
Wild Type ◽  
Resistance Factors ◽  
Insight Into

scholarly journals The Stringent Response Is Required for Amino Acid and Nitrate Utilization, Nod Factor Regulation, Nodulation, and Nitrogen Fixation in Rhizobium etli

2005 ◽  
Vol 187 (15) ◽  
pp. 5075-5083 ◽  
Author(s):  
Arturo Calderón-Flores ◽  
Gisela Du Pont ◽  
Alejandro Huerta-Saquero ◽  
Horacio Merchant-Larios ◽  
Luis Servín-González ◽  
...  
Keyword(s):  
Amino Acids ◽  
Type Strain ◽  
Wild Type Strain ◽  
Stringent Response ◽  
Insertion Mutant ◽  
Rhizobium Etli ◽  
Wild Type ◽  
Nod Factor ◽  
Truncated Protein ◽  
Carbon And Nitrogen

ABSTRACT A Rhizobium etli Tn5 insertion mutant, LM01, was selected for its inability to use glutamine as the sole carbon and nitrogen source. The Tn5 insertion in LM01 was localized to the rsh gene, which encodes a member of the RelA/SpoT family of proteins. The LM01 mutant was affected in the ability to use amino acids and nitrate as nitrogen sources and was unable to accumulate (p)ppGpp when grown under carbon and nitrogen starvation, as opposed to the wild-type strain, which accumulated (p)ppGpp under these conditions. The R. etli rsh gene was found to restore (p)ppGpp accumulation to a ΔrelA ΔspoT mutant of Escherichia coli. The R. etli Rsh protein consists of 744 amino acids, and the Tn5 insertion in LM01 results in the synthesis of a truncated protein of 329 amino acids; complementation experiments indicate that this truncated protein is still capable of (p)ppGpp hydrolysis. A second rsh mutant of R. etli, strain AC1, was constructed by inserting an Ω element at the beginning of the rsh gene, resulting in a null allele. Both AC1 and LM01 were affected in Nod factor production, which was constitutive in both strains, and in nodulation; nodules produced by the rsh mutants in Phaseolus vulgaris were smaller than those produced by the wild-type strain and did not fix nitrogen. In addition, electron microscopy revealed that the mutant bacteroids lacked poly-β-hydroxybutyrate granules. These results indicate a central role for the stringent response in symbiosis.

scholarly journals Campylobacter jejuni FlpA Binds Fibronectin and Is Required for Maximal Host Cell Adherence

2009 ◽  
Vol 192 (1) ◽  
pp. 68-76 ◽  
Author(s):  
Michael E. Konkel ◽  
Charles L. Larson ◽  
Rebecca C. Flanagan
Keyword(s):  
Amino Acids ◽  
Epithelial Cells ◽  
Campylobacter Jejuni ◽  
Type Strain ◽  
Wild Type Strain ◽  
Major Constituent ◽  
Enzyme Linked Immunosorbent Assay ◽  
Type I ◽  
Wild Type ◽  
Type Iii

ABSTRACT Campylobacter jejuni is one of the most frequent bacterial causes of food-borne gastrointestinal disease in developed countries. Previous work indicates that the binding of C. jejuni to human intestinal cells is crucial for host colonization and disease. Fibronectin (Fn), a major constituent of the extracellular matrix, is a ∼250-kDa glycoprotein present at regions of cell-to-cell contact in the intestinal epithelium. Fn is composed of three types of repeating units: type I (∼45 amino acids), type II (∼60 amino acids), and type III (∼90 amino acids). The deduced amino acid sequence of C. jejuni flpA (Cj1279c) contains at least three Fn type III domains. Based on the presence of the Fn type III domains, we hypothesized that FlpA contributes to the binding of C. jejuni to human INT 407 epithelial cells and Fn. We assessed the contribution of FlpA in C. jejuni binding to host cells by in vitro adherence assays with a C. jejuni wild-type strain and a C. jejuni flpA mutant and binding of purified FlpA protein to Fn by enzyme-linked immunosorbent assay (ELISA). Adherence assays revealed the binding of the C. jejuni flpA mutant to INT 407 epithelial cells was significantly reduced compared with that for a wild-type strain. In addition, rabbit polyclonal serum generated against FlpA blocked C. jejuni adherence to INT 407 cells in a concentration-dependent manner. Binding of FlpA to Fn was found to be dose dependent and saturable by ELISA, demonstrating the specificity of the interaction. Based on these data, we conclude that FlpA mediates C. jejuni attachment to host epithelial cells via Fn binding.

scholarly journals Increased Volatile Anesthetic Requirement in Short-sleeping Drosophila  Mutants

2009 ◽  
Vol 110 (2) ◽  
pp. 313-316 ◽  
Author(s):  
Bernd Weber ◽  
Christian Schaper ◽  
Daniel Bushey ◽  
Marko Rohlfs ◽  
Markus Steinfath ◽  
...  
Keyword(s):  
Gene Mutation ◽  
Single Gene ◽  
Volatile Anesthetic ◽  
Volatile Anesthetics ◽  
Wild Type ◽  
Response Curves ◽  
Sleep Behavior ◽  
Single Gene Mutation ◽  
Anesthetic Requirement ◽  
Drosophila Mutants

Background Anesthesia and sleep share physiologic and behavioral similarities. The anesthetic requirement of the recently identified Drosophila mutant minisleeper and other Drosophila mutants was investigated. Methods Sleep and wakefulness were determined by measuring activity of individual wild-type and mutant flies. Based on the response of the flies at different concentrations of the volatile anesthetics isoflurane and sevoflurane, concentration-response curves were generated and EC50 values were calculated. Results The average amount of daily sleep in wild-type Drosophila (n = 64) was 965 +/- 15 min, and 1,022 +/- 29 in Na[har](P > 0.05; n = 32) (mean +/- SEM, all P compared to wild-type and other shaker alleles). Sh flies slept 584 +/- 13 min (n = 64, P < 0.01), Sh flies 412 +/- 22 min (n = 32, P < 0.01), and Sh flies 782 +/- 25 min (n = 32, P < 0.01). The EC50 values for isoflurane were 0.706 (95% CI 0.649 to 0.764, n = 661) and for sevoflurane 1.298 (1.180 to 1.416, n = 522) in wild-type Drosophila; 1.599 (1.527 to 1.671, n = 308) and 2.329 (2.177 to 2.482, n = 282) in Sh, 1.306 (1.212 to 1.400, n = 393) and 2.013 (1.868 to 2.158, n = 550) in Sh, 0.957 (0.860 to 1.054, n = 297) and 1.619 (1.508 to 1.731, n = 386) in Sh, and 0.6154 (0.581 to 0.649, n = 360; P < 0.05) and 0.9339 (0.823 to 1.041, n = 274) in Na[har], respectively (all P < 0.01). Conclusions A single-gene mutation in Drosophila that causes an extreme reduction in daily sleep is responsible for a significant increase in the requirement of volatile anesthetics. This suggests that a single gene mutation affects both sleep behavior and anesthesia and sedation.

scholarly journals Discrete Change in Volatile Anesthetic Sensitivity in Mice with Inactivated Tandem Pore Potassium Ion Channel TRESK

2010 ◽  
Vol 113 (6) ◽  
pp. 1326-1337 ◽  
Author(s):  
Yun Jeong Chae ◽  
Jianan Zhang ◽  
Paul Au ◽  
Marta Sabbadini ◽  
Guo-Xi Xie ◽  
...  
Keyword(s):  
Ion Channel ◽  
Knockout Mice ◽  
Minimum Alveolar Concentration ◽  
Volatile Anesthetic ◽  
Volatile Anesthetics ◽  
Potassium Ion ◽  
Potassium Ion Channel ◽  
Wild Type ◽  
Hot Plate Test ◽  
Neurobehavioral Function

Background We investigated the role of tandem pore potassium ion channel (K2P) TRESK in neurobehavioral function and volatile anesthetic sensitivity in genetically modified mice. Methods Exon III of the mouse TRESK gene locus was deleted by homologous recombination using a targeting vector. The genotype of bred mice (wild type, knockout, or heterozygote) was determined using polymerase chain reaction. Morphologic and behavioral evaluations of TRESK knockout mice were compared with wild-type littermates. Sensitivity of bred mice to isoflurane, halothane, sevoflurane, and desflurane were studied by determining the minimum alveolar concentration preventing movement to tail clamping in 50% of each genotype. Results With the exception of decreased number of inactive periods and increased thermal pain sensitivity (20% decrease in latency with hot plate test), TRESK knockout mice had healthy development and behavior. TRESK knockout mice showed a statistically significant 8% increase in isoflurane minimum alveolar concentration compared with wild-type littermates. Sensitivity to other volatile anesthetics was not significantly different. Spontaneous mortality of TRESK knockout mice after initial anesthesia testing was nearly threefold higher than that of wild-type littermates. Conclusions TRESK alone is not critical for baseline central nervous system function but may contribute to the action of volatile anesthetics. The inhomogeneous change in anesthetic sensitivity corroborates findings in other K2P knockout mice and supports the theory that the mechanism of volatile anesthetic action involves multiple targets. Although it was not shown in this study, a compensatory effect by other K2P channels may also contribute to these observations.

scholarly journals Epigenetic Control of Chromosome Breakage at the 5′ End of Paramecium tetraurelia Gene A

2004 ◽  
Vol 3 (5) ◽  
pp. 1136-1146 ◽  
Author(s):  
Laurence Amar ◽  
Karine Dubrana
Keyword(s):  
Mutant Strain ◽  
Type Strain ◽  
Wild Type Strain ◽  
Chromosome Breakage ◽  
Paramecium Tetraurelia ◽  
Wild Type ◽  
Dna Rearrangements ◽  
Rna Molecules ◽  
Extensive Analysis ◽  
Insight Into

ABSTRACT Macronuclei and micronuclei of ciliates have related genomes, with macronuclei developing from zygotic micronuclei through programmed DNA rearrangements. While Paramecium tetraurelia wild-type strain 51 and mutant strain d48 have the same micronuclear genome, qualitative differences between their macronuclear genomes have been described, demonstrating that programmed DNA rearrangements could be epigenetically controlled in ciliates. Macronuclear chromosomes end downstream of gene A (A51 Mac ends) and at the 5′ end of gene A (Ad48 Mac ends) in strains 51 and d48, respectively. To gain further insight into the process of chromosome end formation, we performed an extensive analysis of locus A rearrangement in strains d48 and 51, in strain d12, which harbors a gene A deletion, and in interstrain cross progeny. We show that (i) allele Ad12 harbors a deletion of >16 kb, (ii) A51 Mac ends distribute over four rather than three DNA regions, (iii) strains d48 and 51 display only quantitative differences (rare Ad48 and A51 Mac ends do form in strains 51 and d48, respectively), (iv) the level of A51 Mac ends is severalfold enhanced in d12- and d48-derived progeny, and (v) this level inversely correlates with the level of Ad48 Mac ends in the d48 parent. Together, these data lead to a model in which the formation of Ad48 Mac ends is epigenetically controlled by a d48 factor(s). We propose that the d48 factor(s) may be derived from RNA molecules transcribed from the Ad48 Mac ends and encompassing the truncated A gene and telomeric repeats.

Cloning and overexpression of the xylose isomerase gene from Burkholderia sacchari and production of polyhydroxybutyrate from xylose

2009 ◽  
Vol 55 (8) ◽  
pp. 1012-1015 ◽  
Author(s):  
Mateus Schreiner Garcez Lopes ◽  
José Gregório Cabrera Gomez ◽  
Luiziana Ferreira Silva
Keyword(s):  
Amino Acids ◽  
Type Strain ◽  
Wild Type Strain ◽  
Xylose Isomerase ◽  
Homologous Gene ◽  
Broad Host Range ◽  
Xylose Utilization ◽  
Wild Type ◽  
Broad Host Range Plasmid ◽  
In The Wild

A different organization for the xyl operon was found in different genomes of Burkholderia and Pseudomomas species. Degenerated primers were designed based on Burkholderia genomes and used to amplify the xylose isomerase gene (xylA) from Burkholderia sacchari IPT101. The gene encoded a protein of 329 amino acids, which showed the highest similarity (90%) to the homologous gene of Burkholderia dolosa . It was cloned in the broad host range plasmid pBBR1MCS-2, which partially restored growth and polyhydroxybutyrate production capability in xylose to a B. sacchari xyl– mutant. When xylA was overexpressed in the wild-type strain, it was not able to increase growth and polyhydroxybutyrate production, suggesting that XylA activity is not limiting for xylose utilization in B. sacchari.

Genetic Differences Affecting the Potency of Stereoisomers of Isoflurane

1996 ◽  
Vol 85 (2) ◽  
pp. 385-392 ◽  
Author(s):  
Phil G. Morgan ◽  
Marianne F. Usiak ◽  
Margaret M. Sedensky
Keyword(s):  
Volatile Anesthetic ◽  
Genetic Differences ◽  
Wild Type ◽  
C Elegans ◽  
Mutant Strains ◽  
Anesthetic Action ◽  
Type C ◽  
Sites Of Action ◽  
Standard Techniques

Background In previous studies, researchers demonstrated the ability of a variety of organisms and in vitro sites of anesthetic action to distinguish between stereoisomers of isoflurane or halothane. However, it was not shown whether organisms with differing sensitivities to stereoisomers of one volatile anesthetic are able to distinguish between stereoisomers of another. In this study, the responses of mutants of Caenorbabditis elegans to stereoisomers of isoflurane were determined for comparison to previous results in halothane. Methods Mutant strains of C. elegans were isolated and grown by standard techniques. The EC50s (the effective concentrations of anesthetia at which 50% of the animals are immobilized for 10 s) of stereoisomers of isoflurane and the racemate were determined in wild type and mutant strains of C. elegans. Results Wild type C. elegans and strains with high EC50S of the racemate were more sensitive to the (+) isomer of isoflurane by approximately 30%. The racemate showed a EC50s similar to the less potent isomer, the (-) form. In the strains with low EC50s, one strain showed no ability to differentiate between the stereoisomers, whereas two showed a 60% difference between the (+) and (-) forms. Conclusions The ability to distinguish between stereoisomers of isoflurane is associated with genetic loci separate from those that distinguish between stereoisomers of halothane. These results are consistent with multiple sites of action for these anesthetics.

scholarly journals Inactivation of Lgt Allows Systematic Characterization of Lipoproteins from Listeria monocytogenes

2006 ◽  
Vol 189 (2) ◽  
pp. 313-324 ◽  
Author(s):  
Maja Baumgärtner ◽  
Uwe Kärst ◽  
Birgit Gerstel ◽  
Martin Loessner ◽  
Jürgen Wehland ◽  
...  
Keyword(s):  
Listeria Monocytogenes ◽  
Wild Type Strain ◽  
Culture Supernatant ◽  
Eukaryotic Cell ◽  
Signal Peptidase ◽  
Wild Type ◽  
Intracellular Growth ◽  
Extracellular Proteome ◽  
Insight Into

ABSTRACT Lipoprotein anchoring in bacteria is mediated by the prolipoprotein diacylglyceryl transferase (Lgt), which catalyzes the transfer of a diacylglyceryl moiety to the prospective N-terminal cysteine of the mature lipoprotein. Deletion of the lgt gene in the gram-positive pathogen Listeria monocytogenes (i) impairs intracellular growth of the bacterium in different eukaryotic cell lines and (ii) leads to increased release of lipoproteins into the culture supernatant. Comparative extracellular proteome analyses of the EGDe wild-type strain and the Δlgt mutant provided systematic insight into the relative expression of lipoproteins. Twenty-six of the 68 predicted lipoproteins were specifically released into the extracellular proteome of the Δlgt strain, and this proved that deletion of lgt is an excellent approach for experimental verification of listerial lipoproteins. Consequently, we generated Δlgt ΔprfA double mutants to detect lipoproteins belonging to the main virulence regulon that is controlled by PrfA. Overall, we identified three lipoproteins whose extracellular levels are regulated and one lipoprotein that is posttranslationally modified depending on PrfA. It is noteworthy that in contrast to previous studies of Escherichia coli, we unambiguously demonstrated that lipidation by Lgt is not a prerequisite for activity of the lipoprotein-specific signal peptidase II (Lsp) in Listeria.

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