Open Access. Powered by Scholars. Published by Universities.®
- File Type
Articles 1 - 17 of 17
Full-Text Articles in Life Sciences
Phosphorylase A Activity As An Indicator Of Neutrophil Activation Of Chemotactic Peptide, Joan Slonczewski, Mary W. Wilde, Sally H. Zigmond
Phosphorylase A Activity As An Indicator Of Neutrophil Activation Of Chemotactic Peptide, Joan Slonczewski, Mary W. Wilde, Sally H. Zigmond
Joan Slonczewski
ABSTRACT The activity of glycogen phosphorylase, an enzyme that is activated by both cAMP and calcium, was used as an indicator of the state of the cytoplasm after chemotactic stimulation of polymorphonuclear leukocytes (neutrophils). The activity of the enzyme showed a clear dependence on cytoplasmic calcium. Addition of the calcium ionophore A23187 caused a 4-5-fold increase in activity of phosphorylase a. In the absence of external Ca 2+, A23187 caused only brief transient activation of phosphorylase; probably reflecting release of sequestered intracellular Ca 2÷. Addition of the chemotactic peptide N-formylnorleucylleucylphenylalanine (FNLLP) caused a transient 2-3-fold activation of the enzyme. The …
Cytoplasmic Ph Measurement And Homeostasis In Bacteria And Archaea., Joan Slonczewski
Cytoplasmic Ph Measurement And Homeostasis In Bacteria And Archaea., Joan Slonczewski
Joan Slonczewski
Of all the molecular determinants for growth, the hydronium and hydroxide ions are found naturally in the widest concentration range, from acid mine drainage below pH 0 to soda lakes above pH 13. Most bacteria and archaea have mechanisms that maintain their internal, cytoplasmic pH within a narrower range than the pH outside the cell, termed "pH homeostasis." Some mechanisms of pH homeostasis are specific to particular species or groups of microorganisms while some common principles apply across the pH spectrum. The measurement of internal pH of microbes presents challenges, which are addressed by a range of techniques under varying …
Rapid Acid Treatment Of Escherichia Coli: Transcriptomic Response And Recovery, Joan Slonczewski, Geetha Kannan, Jessica C . Wilks, Devon M. Fitzgerald, Sandra S. Bondurant, Brian D. Jones
Rapid Acid Treatment Of Escherichia Coli: Transcriptomic Response And Recovery, Joan Slonczewski, Geetha Kannan, Jessica C . Wilks, Devon M. Fitzgerald, Sandra S. Bondurant, Brian D. Jones
Joan Slonczewski
Background: Many E. coli genes show pH-dependent expression during logarithmic growth in acid (pH 5–6) or in base (pH 8–9). The effect of rapid pH change, however, has rarely been tested. Rapid acid treatment could distinguish between genes responding to external pH, and genes responding to cytoplasmic acidification, which occurs transiently following rapid external acidification. It could reveal previously unknown acid-stress genes whose effects are transient, as well as show which acid-stress genes have a delayed response. Results: Microarray hybridization was employed to observe the global gene expression of E. coli K-12 W3110 following rapid acidification of the external medium, …
Ph Of The Cytoplasm And Periplasm Of Escherichia Coli: Rapid Measurement By Green Fluorescent Protein Fluorimetry, Joan Slonczewski, Jessica C. Wilks
Ph Of The Cytoplasm And Periplasm Of Escherichia Coli: Rapid Measurement By Green Fluorescent Protein Fluorimetry, Joan Slonczewski, Jessica C. Wilks
Joan Slonczewski
Cytoplasmic pH and periplasmic pH of Escherichia coli cells in suspension were observed with 4-s time resolution using fluorimetry of TorA-green fluorescent protein mutant 3* (TorA-GFPmut3*) and TetR-yellow fluorescent protein. Fluorescence intensity was correlated with pH using cell suspensions containing 20 mM benzoate, which equalizes the cytoplasmic pH with the external pH. When the external pH was lowered from pH 7.5 to 5.5, the cytoplasmic pH fell within 10 to 20 s to pH 5.6 to 6.5. Rapid recovery occurred until about 30 s after HCl addition and was followed by slower recovery over the next 5 min. As a …
Acid-Adapted Strains Of Escherichia Coli K-12 Obtained By Experimental Evolution, Joan Slonczewski, Mark M. Harden, Amanda He, Michelle W. Clark, Kaitlin Creamer, Issam Hamdallah, Keith A. Martinez Ii, Robert L. Kresslein, Sean P. Bush
Acid-Adapted Strains Of Escherichia Coli K-12 Obtained By Experimental Evolution, Joan Slonczewski, Mark M. Harden, Amanda He, Michelle W. Clark, Kaitlin Creamer, Issam Hamdallah, Keith A. Martinez Ii, Robert L. Kresslein, Sean P. Bush
Joan Slonczewski
Enteric bacteria encounter a wide range of pHs throughout the human intestinal tract. We conducted experimental evolution of Escherichia coli K-12 to isolate clones with increased fitness during growth under acidic conditions (pH 4.5 to 4.8). Twenty-four independent populations of E. coli K-12 W3110 were evolved in LBK medium (10 g/liter tryptone, 5 g/liter yeast extract, 7.45 g/liter KCl) buffered with homopiperazine-N,N=-bis-2-(ethanosulfonic acid) and malate at pH 4.8. At generation 730, the pH was decreased to 4.6 with HCl. By 2,000 generations, all populations had achieved higher endpoint growth than the ancestor at pH 4.6 but not at pH 7.0. …
Cytoplasmic Ph Response To Acid Stress In Individual Cells Of Escherichia Coli And Bacillus Subtilis Observed By Fluorescence Ratio Imaging Microscopy, Joan Slonczewski, Keith A. Martinez Ii, Ryan D. Kitko, J. Patrick Mershon, Haley E. Adcox, Kotiba A. Malek, Melanie B. Berkmen
Cytoplasmic Ph Response To Acid Stress In Individual Cells Of Escherichia Coli And Bacillus Subtilis Observed By Fluorescence Ratio Imaging Microscopy, Joan Slonczewski, Keith A. Martinez Ii, Ryan D. Kitko, J. Patrick Mershon, Haley E. Adcox, Kotiba A. Malek, Melanie B. Berkmen
Joan Slonczewski
The ability of Escherichia coli and Bacillus subtilis to regulate their cytoplasmic pH is well studied in cell suspensions but is poorly understood in individual adherent cells and biofilms. We observed the cytoplasmic pH of individual cells using ratiometric pHluorin. A standard curve equating the fluorescence ratio with pH was obtained by perfusion at a range of external pH 5.0 to 9.0, with uncouplers that collapse the transmembrane pH difference. Adherent cells were acid stressed by switching the perfusion medium from pH 7.5 to pH 5.5. The E. coli cytoplasmic pH fell to a value that varied among individual cells …
Cytoplasmic Acidification And The Benzoate Transcriptome In Bacillus Subtilis, Joan Slonczewski, Ryan D. Kitko, Rebecca L. Cleeton, Erin I. Armentrout, Grace E. Lee, Ken Noguchi, Melanie B. Berkmen, Brian D. Jones
Cytoplasmic Acidification And The Benzoate Transcriptome In Bacillus Subtilis, Joan Slonczewski, Ryan D. Kitko, Rebecca L. Cleeton, Erin I. Armentrout, Grace E. Lee, Ken Noguchi, Melanie B. Berkmen, Brian D. Jones
Joan Slonczewski
Background: Bacillus subtilis encounters a wide range of environmental pH. The bacteria maintain cytoplasmic pH within a narrow range. Response to acid stress is a poorly understood function of external pH and of permeant acids that conduct protons into the cytoplasm. Methods and Principal Findings: Cytoplasmic acidification and the benzoate transcriptome were observed in Bacillus subtilis. Cytoplasmic pH was measured with 4-s time resolution using GFPmut3b fluorimetry. Rapid external acidification (pH 7.5 to 6.0) acidified the B. subtilis cytoplasm, followed by partial recovery. Benzoate addition up to 60 mM at external pH 7 depressed cytoplasmic pH but left a transmembrane …
Ph Regulates Genes For Flagellar Motility, Catabolism, And Oxidative Stress In Escherichia Coli K-12†, Joan Slonczewski
Ph Regulates Genes For Flagellar Motility, Catabolism, And Oxidative Stress In Escherichia Coli K-12†, Joan Slonczewski
Joan Slonczewski
Gene expression profiles of Escherichia coli K-12 W3110 were compared as a function of steady-state external pH. Cultures were grown to an optical density at 600 nm of 0.3 in potassium-modified Luria-Bertani medium buffered at pH 5.0, 7.0, and 8.7. For each of the three pH conditions, cDNA from RNA of five independent cultures was hybridized to Affymetrix E. coli arrays. Normalized expression indices were calculated for each gene and intergenic region (IG). Differential expression among the three pH classes was observed for 763 genes and 353 IGs. Hierarchical clustering yielded six well-defined clusters of pH profiles, designated Acid High …
Generalization Of Dna Microarray Dispersion Properties: Microarray Equivalent Of T-Distribution, Joan Slonczewski
Generalization Of Dna Microarray Dispersion Properties: Microarray Equivalent Of T-Distribution, Joan Slonczewski
Joan Slonczewski
Background: DNA microarrays are a powerful technology that can provide a wealth of gene expression data for disease studies, drug development, and a wide scope of other investigations. Because of the large volume and inherent variability of DNA microarray data, many new statistical methods have been developed for evaluating the significance of the observed differences in gene expression. However, until now little attention has been given to the characterization of dispersion of DNA microarray data. Results: Here we examine the expression data obtained from 682 Affymetrix GeneChips® with 22 different types and we demonstrate that the Gaussian (normal) frequency distribution …
A Requirement Of Tolc And Mdr Efflux Pumps For Acid Adaptation And Gadab Induction In Escherichia Coli, Joan Slonczewski, Kari N. W. Deininger, Akina Horikawa, Ryan D. Kitko, Ryoko Tatsumi, Judah L. Rosner, Masaaki Wachi
A Requirement Of Tolc And Mdr Efflux Pumps For Acid Adaptation And Gadab Induction In Escherichia Coli, Joan Slonczewski, Kari N. W. Deininger, Akina Horikawa, Ryan D. Kitko, Ryoko Tatsumi, Judah L. Rosner, Masaaki Wachi
Joan Slonczewski
Background: The TolC outer membrane channel is a key component of several multidrug resistance (MDR) efflux pumps driven by H+ transport in Escherichia coli. While tolC expression is under the regulation of the EvgA-Gad acid resistance regulon, the role of TolC in growth at low pH and extreme-acid survival is unknown. Methods and Principal Findings: TolC was required for extreme-acid survival (pH 2) of strain W3110 grown aerobically to stationary phase. A tolC deletion decreased extreme-acid survival (acid resistance) of aerated pH 7.0-grown cells by 105 -fold and of pH 5.5-grown cells by 10-fold. The requirement was specific for acid …
Escherichia Coli K-12 Survives Anaerobic Exposure At Ph 2 Without Rpos, Gad, Or Hydrogenases, But Shows Sensitivity To Autoclaved Broth Products, Joan Slonczewski, Daniel P. Riggins, Maria J. Narvaez, Keith A. Martinez, Mark M. Harden
Escherichia Coli K-12 Survives Anaerobic Exposure At Ph 2 Without Rpos, Gad, Or Hydrogenases, But Shows Sensitivity To Autoclaved Broth Products, Joan Slonczewski, Daniel P. Riggins, Maria J. Narvaez, Keith A. Martinez, Mark M. Harden
Joan Slonczewski
Escherichia coli and other enteric bacteria survive exposure to extreme acid (pH 2 or lower) in gastric fluid. Aerated cultures survive via regulons expressing glutamate decarboxylase (Gad, activated by RpoS), cyclopropane fatty acid synthase (Cfa) and others. But extreme-acid survival is rarely tested under low oxygen, a condition found in the stomach and the intestinal tract. We observed survival of E. coli K-12 W3110 at pH 1.2–pH 2.0, conducting all manipulations (overnight culture at pH 5.5, extreme-acid exposure, dilution and plating) in a glove box excluding oxygen (10% H2, 5% CO2, balance N2). With dissolved O2 concentrations maintained below 6 …
Hydrogenase-3 Contributes To Anaerobic Acid Resistance Of Escherichia Coli, Joan Slonczewski, Ken Noguchi, Daniel P. Riggins, Khalid C. Eldahan, Ryan D. Kitko
Hydrogenase-3 Contributes To Anaerobic Acid Resistance Of Escherichia Coli, Joan Slonczewski, Ken Noguchi, Daniel P. Riggins, Khalid C. Eldahan, Ryan D. Kitko
Joan Slonczewski
Background: Hydrogen production by fermenting bacteria such as Escherichia coli offers a potential source of hydrogen biofuel. Because H2 production involves consumption of 2H+ , hydrogenase expression is likely to involve pH response and regulation. Hydrogenase consumption of protons in E. coli has been implicated in acid resistance, the ability to survive exposure to acid levels (pH 2–2.5) that are three pH units lower than the pH limit of growth (pH 5–6). Enhanced survival in acid enables a larger infective inoculum to pass through the stomach and colonize the intestine. Most acid resistance mechanisms have been defined using aerobic cultures, …
Osmolytes Contribute To Ph Homeostasis Of Escherichia Coli, Joan Slonczewski, Ryan D. Kitko, Jessica C. Wilks, Gian M. Garduque
Osmolytes Contribute To Ph Homeostasis Of Escherichia Coli, Joan Slonczewski, Ryan D. Kitko, Jessica C. Wilks, Gian M. Garduque
Joan Slonczewski
Background: Cytoplasmic pH homeostasis in Escherichia coli includes numerous mechanisms involving pH-dependent catabolism and ion fluxes. An important contributor is transmembrane K+ flux, but the actual basis of K+ compensation for pH stress remains unclear. Osmoprotection could mediate the pH protection afforded by K+ and other osmolytes. Methods and Principal Findings: The cytoplasmic pH of E. coli K-12 strains was measured by GFPmut3 fluorimetry. The wild-type strain Frag1 was exposed to rapid external acidification by HCl addition. Recovery of cytoplasmic pH was enhanced equally by supplementation with NaCl, KCl, proline, or sucrose. A triple mutant strain TK2420 defective for the …
Oxygen Limitation Modulates Ph Regulation Of Catabolism And Hydrogenases, Multidrug Transporters, And Envelope Composition In Escherichia Coli K-12, Joan Slonczewski, Everett T. Hayes, Jessica C. Wilks, Piero Sanfilippo, Elizabeth Yohannes, Daniel P. Tate, Brian D. Jones, Michael D. Radmacher, Sandra S. Bondurant
Oxygen Limitation Modulates Ph Regulation Of Catabolism And Hydrogenases, Multidrug Transporters, And Envelope Composition In Escherichia Coli K-12, Joan Slonczewski, Everett T. Hayes, Jessica C. Wilks, Piero Sanfilippo, Elizabeth Yohannes, Daniel P. Tate, Brian D. Jones, Michael D. Radmacher, Sandra S. Bondurant
Joan Slonczewski
Background: In Escherichia coli, pH regulates genes for amino-acid and sugar catabolism, electron transport, oxidative stress, periplasmic and envelope proteins. Many pH-dependent genes are co-regulated by anaerobiosis, but the overall intersection of pH stress and oxygen limitation has not been investigated. Results: The pH dependence of gene expression was analyzed in oxygen-limited cultures of E. coli K-12 strain W3110. E. coli K-12 strain W3110 was cultured in closed tubes containing LBK broth buffered at pH 5.7, pH 7.0, and pH 8.5. Affymetrix array hybridization revealed pH-dependent expression of 1,384 genes and 610 intergenic regions. A core group of 251 genes …
Ph-Dependent Expression Of Periplasmic Proteins And Amino Acid Catabolism In Escherichia Coli, Joan Slonczewski, Lauren M. Stancik, Dawn M. Stancik, Brian Schmidt, D. Michael Barnhart, Yuliya N. Yoncheva
Ph-Dependent Expression Of Periplasmic Proteins And Amino Acid Catabolism In Escherichia Coli, Joan Slonczewski, Lauren M. Stancik, Dawn M. Stancik, Brian Schmidt, D. Michael Barnhart, Yuliya N. Yoncheva
Joan Slonczewski
Escherichia coli grows over a wide range of pHs (pH 4.4 to 9.2), and its own metabolism shifts the external pH toward either extreme, depending on available nutrients and electron acceptors. Responses to pH values across the growth range were examined through two-dimensional electrophoresis (2-D gels) of the proteome and through lac gene fusions. Strain W3110 was grown to early log phase in complex broth buffered at pH 4.9, 6.0, 8.0, or 9.1. 2-D gel analysis revealed the pH dependence of 19 proteins not previously known to be pH dependent. At low pH, several acetate-induced proteins were elevated (LuxS, Tpx, …
Ph-Dependent Catabolic Protein Expression During Anaerobic Growth Of Escherichia Coli K-12, Joan Slonczewski, Elizabeth Yohannes, D. Michael Barnhart
Ph-Dependent Catabolic Protein Expression During Anaerobic Growth Of Escherichia Coli K-12, Joan Slonczewski, Elizabeth Yohannes, D. Michael Barnhart
Joan Slonczewski
During aerobic growth of Escherichia coli, expression of catabolic enzymes and envelope and periplasmic proteins is regulated by pH. Additional modes of pH regulation were revealed under anaerobiosis. E. coli K-12 strain W3110 was cultured anaerobically in broth medium buffered at pH 5.5 or 8.5 for protein identification on proteomic two-dimensional gels. A total of 32 proteins from anaerobic cultures show pH-dependent expression, and only four of these proteins (DsbA, TnaA, GatY, and HdeA) showed pH regulation in aerated cultures. The levels of 19 proteins were elevated at the high pH; these proteins included metabolic enzymes (DhaKLM, GapA, TnaA, HisC, …
Polyamine Stress At High Ph In Escherichia Coli K-12, Joan Slonczewski, Elizabeth Yohannes, Amy E. Thurber, Jessica C. Wilks, Daniel P. Tate
Polyamine Stress At High Ph In Escherichia Coli K-12, Joan Slonczewski, Elizabeth Yohannes, Amy E. Thurber, Jessica C. Wilks, Daniel P. Tate
Joan Slonczewski
Background: Polyamines such as spermine and spermidine are required for growth of Escherichia coli; they interact with nucleic acids, and they bind to ribosomes. Polyamines block porins and decrease membrane permeability, activities that may protect cells in acid. At high concentrations, however, polyamines impair growth. They impair growth more severely at high pH, probably due to their increased uptake as membrane-permeant weak bases. The role of pH is critical in understanding polyamine stress. Results: The effect of polyamines was tested on survival of Escherichia coli K-12 W3110 in extreme acid or base (pH conditions outside the growth range). At pH …