Pseudomonas aeruginosa lectins I and II and their interaction with human airway cilia

2005 ◽  
Vol 119 (8) ◽  
pp. 595-599 ◽  
Author(s):  
Marco Mewe ◽  
Denis Tielker ◽  
Robert Schönberg ◽  
Melitta Schachner ◽  
Karl-Erich Jaeger ◽  
...  
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Joint Action ◽  
Mammalian Cells ◽  
Time Dependent ◽  
Carbohydrate Binding ◽  
Dependent Manner ◽  
Similar Time ◽  
Airway Cilia ◽  
Kinetics Of ◽  
Ciliary Beat

The bacterium Pseudomonas aeruginosa (PA) produces two carbohydrate binding lectins, designated PA lectin-I and lectin-II (PA-IL, PA-IIL). Both lectins are used by the bacterium to adhere to the glycocalyx of mammalian cells. In addition, the lectins immobilize ciliary beat. The kinetics of ciliary beat inhibition by each individual lectin have been analysed; however, their joint action on cilia has not been reported. Here we demonstrate that PA-IL and PA-IIL inhibit ciliary beat in a similar time-dependent manner. If applied simultaneously, ciliary beat inhibition after five hours of incubation was weaker than if each lectin was applied separately. Thus it can be hypothesized that the lectins compete for the same binding site(s) of the glycocalyx. Sugar inhibition experiments demonstrate that D-galactose and L-fucose inhibit both lectins, although clear preferences of D-galactose for PA-IL and of L-fucose for PA-IIL exist. These interactions have to be kept in mind when designing sugar-based therapies.

scholarly journals Determinants of mRNA stability in Dictyostelium discoideum amoebae: differences in poly(A) tail length, ribosome loading, and mRNA size cannot account for the heterogeneity of mRNA decay rates.

1988 ◽  
Vol 8 (5) ◽  
pp. 1957-1969 ◽  
Author(s):  
R A Shapiro ◽  
D Herrick ◽  
R E Manrow ◽  
D Blinder ◽  
A Jacobson
Keyword(s):  
Steady State ◽  
Dictyostelium Discoideum ◽  
Mrna Decay ◽  
Decay Rates ◽  
Time Dependent ◽  
Translational Efficiency ◽  
Cdna Clones ◽  
Dependent Manner ◽  
Significant Difference ◽  
Kinetics Of

As an approach to understanding the structures and mechanisms which determine mRNA decay rates, we have cloned and begun to characterize cDNAs which encode mRNAs representative of the stability extremes in the poly(A)+ RNA population of Dictyostelium discoideum amoebae. The cDNA clones were identified in a screening procedure which was based on the occurrence of poly(A) shortening during mRNA aging. mRNA half-lives were determined by hybridization of poly(A)+ RNA, isolated from cells labeled in a 32PO4 pulse-chase, to dots of excess cloned DNA. Individual mRNAs decayed with unique first-order decay rates ranging from 0.9 to 9.6 h, indicating that the complex decay kinetics of total poly(A)+ RNA in D. discoideum amoebae reflect the sum of the decay rates of individual mRNAs. Using specific probes derived from these cDNA clones, we have compared the sizes, extents of ribosome loading, and poly(A) tail lengths of stable, moderately stable, and unstable mRNAs. We found (i) no correlation between mRNA size and decay rate; (ii) no significant difference in the number of ribosomes per unit length of stable versus unstable mRNAs, and (iii) a general inverse relationship between mRNA decay rates and poly(A) tail lengths. Collectively, these observations indicate that mRNA decay in D. discoideum amoebae cannot be explained in terms of random nucleolytic events. The possibility that specific 3'-structural determinants can confer mRNA instability is suggested by a comparison of the labeling and turnover kinetics of different actin mRNAs. A correlation was observed between the steady-state percentage of a given mRNA found in polysomes and its degree of instability; i.e., unstable mRNAs were more efficiently recruited into polysomes than stable mRNAs. Since stable mRNAs are, on average, "older" than unstable mRNAs, this correlation may reflect a translational role for mRNA modifications that change in a time-dependent manner. Our previous studies have demonstrated both a time-dependent shortening and a possible translational role for the 3' poly(A) tracts of mRNA. We suggest, therefore, that the observed differences in the translational efficiency of stable and unstable mRNAs may, in part, be attributable to differences in steady-state poly(A) tail lengths.

scholarly journals The Pseudomonas aeruginosa product pyochelin interferes with Trypanosoma cruzi infection and multiplication in vitro

2020 ◽  
Vol 114 (7) ◽  
pp. 492-498 ◽  
Author(s):  
Gabriele Sass ◽  
Laura C Miller Conrad ◽  
Terrence-Thang H Nguyen ◽  
David A Stevens
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Trypanosoma Cruzi ◽  
Mammalian Cells ◽  
Iron Acquisition ◽  
Vero Cells ◽  
Dependent Manner ◽  
Wild Type ◽  
Current Standard ◽  
Cruzi Infection

Abstract Background Bacteria are sources of numerous molecules used in treatment of infectious diseases. We investigated effects of molecules produced by 26 Pseudomonas aeruginosa strains against infection of mammalian cell cultures with Trypanosoma cruzi, the aetiological agent of Chagas disease. Methods Vero cells were infected with T. cruzi in the presence of wild-type P. aeruginosa supernatants or supernatants of mutants with defects in the production of various virulence, quorum sensing and iron acquisition factors. Quantification of T. cruzi infection (percentage of infected cells) and multiplication (number of amastigotes per infected cell) was performed and cell viability was determined. Results Wild-type P. aeruginosa products negatively affected T. cruzi infection and multiplication in a dose-dependent manner, without evident toxicity for mammalian cells. PvdD/pchE mutation (loss of the P. aeruginosa siderophores pyoverdine and pyochelin) had the greatest impact on anti–T. cruzi activity. Negative effects on T. cruzi infection by pure pyochelin, but not pyoverdine, or other P. aeruginosa exoproducts studied, were quantitatively similar to the effects of benznidazole, the current standard therapy against T. cruzi. Conclusions The P. aeruginosa product pyochelin showed promising activity against T. cruzi and might become a new lead molecule for therapy development.

scholarly journals The Metabolic Chemical Reporter Ac46AzGal Could Incorporate Intracellular Protein Modification in the Form of UDP-6AzGlc Mediated by OGT and Enzymes in the Leloir Pathway

2021 ◽  
Vol 9 ◽  
Author(s):  
Jiajia Wang ◽  
Biao Dou ◽  
Lu Zheng ◽  
Wei Cao ◽  
Peiyu Dong ◽  
...  
Keyword(s):  
Mammalian Cells ◽  
Protein Modification ◽  
Time Dependent ◽  
Dependent Manner ◽  
Intracellular Protein ◽  
Galactose Metabolism ◽  
Naturally Occurring ◽  
Leloir Pathway ◽  
Chemical Reporter ◽  
Complex Glycans

Galactose is a naturally occurring monosaccharide used to build complex glycans that has not been targeted for labeling as a metabolic reporter. Here, we characterize the cellular modification of proteins by using Ac46AzGal in a dose- and time-dependent manner. It is noted that a vast majority of this labeling of Ac46AzGal occurs intracellularly in a range of mammalian cells. We also provided evidence that this labeling is dependent on not only the enzymes of OGT responsible for O-GlcNAcylation but also the enzymes of GALT and GALE in the Leloir pathway. Notably, we discover that Ac46AzGal is not the direct substrate of OGT, and the labeling results may attribute to UDP-6AzGlc after epimerization of UDP-6AzGal via GALE. Together, these discoveries support the conclusion that Ac46AzGal as an analogue of galactose could metabolically label intracellular O-glycosylation modification, raising the possibility of characterization with impaired functions of the galactose metabolism in the Leloir pathway under certain conditions, such as galactosemias.

scholarly journals Cytotoxic activity of Bacillus thuringiensis Cry proteins on mammalian cells transfected with cadherin-like Cry receptor gene of Bombyx mori (silkworm)

2003 ◽  
Vol 369 (3) ◽  
pp. 697-703 ◽  
Author(s):  
Yoko TSUDA ◽  
Fumiki NAKATANI ◽  
Keiko HASHIMOTO ◽  
Satoshi IKAWA ◽  
Chikako MATSUURA ◽  
...  
Keyword(s):  
Bacillus Thuringiensis ◽  
Bombyx Mori ◽  
Mammalian Cells ◽  
Receptor Gene ◽  
Time Dependent ◽  
Hek293 Cells ◽  
Dependent Manner ◽  
Specific Receptor ◽  
Cry Proteins ◽  
Variant Alleles

Cry1Aa, an insecticidal protein produced by Bacillus thuringiensis, has been shown to bind to cadherin-like protein, BtR175, in Bombyx mori (silkworm) midgut. We previously reported three variant alleles of BtR175 (BtR175a, b and c). When transiently expressed in COS7 cells, all the three BtR175 variants bound to Cry1Aa. We stably expressed BtR175b in HEK293 cells. These BtR175b-expressing cells swelled and died in the presence of activated Cry1Aa in a dose- and time-dependent manner, showing that BtR175b itself can impart Cry1Aa-susceptibility to mammalian cells. These cells were more susceptible to Cry1Aa than to Cry1Ab and Cry1Ac. Since dispersed B. mori midgut cells were reported to be highly susceptible to Cry1Ac, this result suggested that other Cry1Ac-specific receptor(s) were simultaneously working with BtR175 in the midgut cells. Advantages are also discussed of applying these transfected mammalian cells to toxicity assays of mutant Cry proteins.

scholarly journals Transcriptional Induction of the Pseudomonas aeruginosa Type III Secretion System by Low Ca2+ and Host Cell Contact Proceeds through Two Distinct Signaling Pathways

2006 ◽  
Vol 74 (6) ◽  
pp. 3334-3341 ◽  
Author(s):  
Nandini Dasgupta ◽  
Alix Ashare ◽  
Gary W. Hunninghake ◽  
Timothy L. Yahr
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Host Cell ◽  
Type Iii Secretion ◽  
Mammalian Cells ◽  
Type Iii Secretion System ◽  
Growth Conditions ◽  
Secretion System ◽  
Host Cells ◽  
Dependent Manner ◽  
Type Iii

ABSTRACT The opportunistic pathogen Pseudomonas aeruginosa utilizes a type III secretion system (T3SS) to intoxicate eukaryotic host cells. Transcription of the T3SS is induced under calcium-limited growth conditions or following intimate contact of P. aeruginosa with host cells. In the present study, we demonstrate that expression of the T3SS is controlled by two distinct regulatory mechanisms and that these mechanisms are differentially activated in a host cell-dependent manner. The first mechanism is dependent upon ExsC, a regulatory protein that couples transcription of the T3SS to the activity of the type III secretion machinery. ExsC is essential for induction of the T3SS under low-calcium-growth conditions and for T3SS-dependent cytotoxicity towards social amoebae, insect cells, and erythrocytes. The second regulatory mechanism functions independently of ExsC and is sufficient to elicit T3SS-dependent cytotoxicity towards certain types of mammalian cells. Although this second pathway (ExsC independent) is sufficient, an exsC mutant demonstrates a lag in the induction of cytotoxicity towards Chinese hamster ovary cells and is attenuated for virulence in a mouse pneumonia model. We propose that the ExsC-dependent pathway is required for full cytotoxicity towards all host cell types tested whereas the ExsC-independent pathway may represent an adaptation that allows P. aeruginosa to increase expression of the T3SS in response to specific types of mammalian cells.

scholarly journals Pharmacokinetics/Pharmacodynamics of Colistin and Imipenem on Mucoid and Nonmucoid Pseudomonas aeruginosa Biofilms

2011 ◽  
Vol 55 (9) ◽  
pp. 4469-4474 ◽  
Author(s):  
Wang Hengzhuang ◽  
Hong Wu ◽  
Oana Ciofu ◽  
Zhijun Song ◽  
Niels Høiby
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Time Course ◽  
Time Dependent ◽  
Content Type ◽  
Planktonic Bacteria ◽  
Kinetics Of ◽  
Higher Doses

ABSTRACTThe time course of activity of colistin and imipenem against mucoid and nonmucoidPseudomonas aeruginosagrowing in a biofilm showed that compared with those for planktonic bacteria, the kinetics of colistin and imipenem retained the concentration- and time-dependent killing, respectively, but higher doses of antibiotics and longer dosing periods were required for biofilm eradication. Biofilms of mucoidP. aeruginosawere more difficult to eradicate than nonmucoid biofilms.

scholarly journals Determinants of mRNA stability in Dictyostelium discoideum amoebae: differences in poly(A) tail length, ribosome loading, and mRNA size cannot account for the heterogeneity of mRNA decay rates

1988 ◽  
Vol 8 (5) ◽  
pp. 1957-1969
Author(s):  
R A Shapiro ◽  
D Herrick ◽  
R E Manrow ◽  
D Blinder ◽  
A Jacobson
Keyword(s):  
Steady State ◽  
Dictyostelium Discoideum ◽  
Mrna Decay ◽  
Decay Rates ◽  
Time Dependent ◽  
Translational Efficiency ◽  
Cdna Clones ◽  
Dependent Manner ◽  
Significant Difference ◽  
Kinetics Of

As an approach to understanding the structures and mechanisms which determine mRNA decay rates, we have cloned and begun to characterize cDNAs which encode mRNAs representative of the stability extremes in the poly(A)+ RNA population of Dictyostelium discoideum amoebae. The cDNA clones were identified in a screening procedure which was based on the occurrence of poly(A) shortening during mRNA aging. mRNA half-lives were determined by hybridization of poly(A)+ RNA, isolated from cells labeled in a 32PO4 pulse-chase, to dots of excess cloned DNA. Individual mRNAs decayed with unique first-order decay rates ranging from 0.9 to 9.6 h, indicating that the complex decay kinetics of total poly(A)+ RNA in D. discoideum amoebae reflect the sum of the decay rates of individual mRNAs. Using specific probes derived from these cDNA clones, we have compared the sizes, extents of ribosome loading, and poly(A) tail lengths of stable, moderately stable, and unstable mRNAs. We found (i) no correlation between mRNA size and decay rate; (ii) no significant difference in the number of ribosomes per unit length of stable versus unstable mRNAs, and (iii) a general inverse relationship between mRNA decay rates and poly(A) tail lengths. Collectively, these observations indicate that mRNA decay in D. discoideum amoebae cannot be explained in terms of random nucleolytic events. The possibility that specific 3'-structural determinants can confer mRNA instability is suggested by a comparison of the labeling and turnover kinetics of different actin mRNAs. A correlation was observed between the steady-state percentage of a given mRNA found in polysomes and its degree of instability; i.e., unstable mRNAs were more efficiently recruited into polysomes than stable mRNAs. Since stable mRNAs are, on average, "older" than unstable mRNAs, this correlation may reflect a translational role for mRNA modifications that change in a time-dependent manner. Our previous studies have demonstrated both a time-dependent shortening and a possible translational role for the 3' poly(A) tracts of mRNA. We suggest, therefore, that the observed differences in the translational efficiency of stable and unstable mRNAs may, in part, be attributable to differences in steady-state poly(A) tail lengths.

Frequency-dependent effects of variousIKrblockers on cardiac action potential duration in a human atrial model

2007 ◽  
Vol 293 (1) ◽  
pp. H660-H669 ◽  
Author(s):  
Kenji Tsujimae ◽  
Shingo Suzuki ◽  
Shingo Murakami ◽  
Yoshihisa Kurachi
Keyword(s):  
Action Potential ◽  
Torsade De Pointes ◽  
Time Dependent ◽  
Type I ◽  
Dependent Manner ◽  
Frequency Dependent ◽  
Modified Model ◽  
Slow Development ◽  
Current Activation ◽  
Kinetics Of

Rapidly activating K+current ( IKr) blockers prolong action potential (AP) duration (APD) in a reverse-frequency-dependent manner and may induce arrhythmias, including torsade de pointes in the ventricle. The IKrblocker dofetilide has been approved for treatment of atrial arrhythmias, including fibrillation. There are, however, a limited number of studies on the action of IKrblockers on atrial AP. When we tested a mathematical model of the human atrial AP (M Courtemanche, RJ Ramirez, S Nattel. Am J Physiol Heart Circ Physiol 275: H301–H321, 1998) to examine the effects of dofetilide-type IKrblockade, this model could not reproduce the reverse-frequency-dependent nature of IKrblockade on atrial APD. We modified the model by introducing a slowly activating K+current activation parameter. As the slow time constant was increased, dofetilide-type blockade induced more prominent reverse-frequency-dependent APD prolongation. Using the modified model, we also examined the effects of two more types of IKrblockade similar to those of quinidine and vesnarinone. Voltage- and time-dependent block of IKrthrough the onset of inhibition by quinidine is much faster than by vesnarinone. When we incorporated the kinetics of the effects of these drugs on IKrinto the model, we found that quinidine-type blockade caused a reverse-frequency-dependent prolongation of APD that was similar to the effect of dofetilide-type blockade, whereas vesnarinone-type blockade did not. This finding coincides with experimental observations. The lack of the reverse frequency dependence in vesnarinone-type blockade was accounted for by the slow development of IKrblockade at depolarized potentials. These results suggest that the voltage- and time-dependent nature of IKrblockade by drugs may be critical for the phenotype of the drug effect on atrial AP.

Influence of Brain Gangliosides on the Formation and Properties of Supported Lipid Bilayers for Binding Kinetics Studies

2018 ◽  
Author(s):  
Luke Jordan ◽  
Nathan Wittenberg
Keyword(s):  
Lipid Bilayers ◽  
Binding Kinetics ◽  
Supported Lipid Bilayers ◽  
Dependent Manner ◽  
Concentration Dependent Manner ◽  
Supported Lipid Bilayer ◽  
Head Groups ◽  
Lipid Diffusion ◽  
Kinetics Of ◽  
Brain Gangliosides

This is a comprehensive study of the effects of the four major brain gangliosides (GM1, GD1b, GD1a, and GT1b) on the adsorption and rupture of phospholipid vesicles on SiO2 surfaces for the formation of supported lipid bilayer (SLB) membranes. Using quartz crystal microbalance with dissipation monitoring (QCM-D) we show that gangliosides GD1a and GT1b significantly slow the SLB formation process, whereas GM1 and GD1b have smaller effects. This is likely due to the net ganglioside charge as well as the positions of acidic sugar groups on ganglioside glycan head groups. Data is included that shows calcium can accelerate the formation of ganglioside-rich SLBs. Using fluorescence recovery after photobleaching (FRAP) we also show that the presence of gangliosides significantly reduces lipid diffusion coefficients in SLBs in a concentration-dependent manner. Finally, using QCM-D and GD1a-rich SLB membranes we measure the binding kinetics of an anti-GD1a antibody that has similarities to a monoclonal antibody that is a hallmark of a variant of Guillain-Barre syndrome.

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