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Thoughts On Quorum Sensing And Fungal Dimorphism, Kenneth W. Nickerson, Audrey L. Atkin, Jessica C. Hargarten, Ruvini U. Pathirana, Sahar Hasim Jan 2012

Thoughts On Quorum Sensing And Fungal Dimorphism, Kenneth W. Nickerson, Audrey L. Atkin, Jessica C. Hargarten, Ruvini U. Pathirana, Sahar Hasim

Papers in Microbiology

Farnesol has been best studied for its role in regulating fungal dimorphism. However, farnesol is also a lipid and in this review we analyze data relevant to farnesol’s function and synthesis from the perspective of farnesol and bacterial endotoxins acting as membrane active compounds. This analysis implicates the possible roles of: (1) endotoxins in the regulation of farnesol production by C. albicans; (2) farnesol in the interactions between C. albicans and the host during disseminated infections; and (3) ubiquinones in the mechanisms for unusually high resistance to farnesol by some C. albicans cell types. Finally we discuss the implications ...


Exogenous Farnesol Interferes With The Normal Progression Of Cytokine Expression During Candidiasis In A Mouse Model, Dhammika H. M. L. P. Navarathna, Kenneth W. Nickerson, Gerald E. Duhamel, Thomas R. Jerrels, Thomas M. Petro Aug 2007

Exogenous Farnesol Interferes With The Normal Progression Of Cytokine Expression During Candidiasis In A Mouse Model, Dhammika H. M. L. P. Navarathna, Kenneth W. Nickerson, Gerald E. Duhamel, Thomas R. Jerrels, Thomas M. Petro

Papers in Microbiology

Candida albicans, a dimorphic fungus composed of yeast and mycelial forms, is the most common human fungal pathogen. Th1 cytokines such as interleukin-2 (IL-2), gamma interferon (IFN-γ), and tumor necrosis factor alpha (TNF-α), which are induced by macrophage IL-12, are critical to resistance against systemic candidiasis, while Th2 cytokines such as IL-4 and IL-5 are less critical. Farnesol is a quorum-sensing molecule produced by C. albicans that controls the formation of mycelia but is also a virulence factor. To determine whether farnesol enhances the virulence of C. albicans by modulating the production of Th1 and Th2 cytokines, mice were pretreated ...


Effect Of Farnesol On A Mouse Model Of Systemic Candidiasis, Determined By Use Of A Dpp3 Knockout Mutant Of Candida Albicans, Dhammika H.M.L.P. Navarathna, Jacob M. Hornby, Navasona Krishnan, Anne M. Parkhurst, Gerald E. Duhamel, Kenneth W. Nickerson Apr 2007

Effect Of Farnesol On A Mouse Model Of Systemic Candidiasis, Determined By Use Of A Dpp3 Knockout Mutant Of Candida Albicans, Dhammika H.M.L.P. Navarathna, Jacob M. Hornby, Navasona Krishnan, Anne M. Parkhurst, Gerald E. Duhamel, Kenneth W. Nickerson

Papers in Microbiology

This work extends our previous observation that the fungus Candida albicans secretes micromolar levels of farnesol and that accumulation of farnesol in vitro prevents the yeast-to-mycelium conversion in a quorumsensing manner. What does farnesol do in vivo? The purpose of this study was to determine the role of farnesol during infection with a well-established mouse model of systemic candidiasis with C. albicans A72 administered by tail vein injection. This question was addressed by altering both endogenous and exogenous farnesol. For endogenous farnesol, we created a knockout mutation in DPP3, the gene encoding a phosphatase which converts farnesyl pyrophosphate to farnesol ...


In Vivo And In Vitro Anaerobic Mating In Candida Albicans, Raluca Dumitru, Dhammika H.M.L.P. Navarathna, Camile P. Semighini, Christian G. Elowsky, Razvan Dumitru, Daniel Dignard, Malcom Whiteway, Audrey L. Atkin, Kenneth W. Nickerson Mar 2007

In Vivo And In Vitro Anaerobic Mating In Candida Albicans, Raluca Dumitru, Dhammika H.M.L.P. Navarathna, Camile P. Semighini, Christian G. Elowsky, Razvan Dumitru, Daniel Dignard, Malcom Whiteway, Audrey L. Atkin, Kenneth W. Nickerson

Papers in Microbiology

Candida albicans cells of opposite mating types are thought to conjugate during infection in mammalian hosts, but paradoxically, the mating-competent opaque state is not stable at mammalian body temperatures. We found that anaerobic conditions stabilize the opaque state at 37°C, block production of farnesol, and permit in vitro mating at 37°C at efficiencies of up to 84%. Aerobically, farnesol prevents mating because it kills the opaque cells necessary for mating, and as a corollary, farnesol production is turned off in opaque cells. These in vitro observations suggest that naturally anaerobic sites, such as the efficiently colonized gastrointestinal (GI ...


Pichia Pastoris Fermentation Optimization: Energy State And Testing A Growth-Associated Model , Bradley A. Plantz, Jayanta Sinha, Lorelie Villarete, Kenneth W. Nickerson, Vicki Schlegel Oct 2006

Pichia Pastoris Fermentation Optimization: Energy State And Testing A Growth-Associated Model , Bradley A. Plantz, Jayanta Sinha, Lorelie Villarete, Kenneth W. Nickerson, Vicki Schlegel

Papers in Microbiology

A growth-associated model was applied to the production of recombinant ovine interferon-τ (rOvIFN-τ) with Pichia pastoris for the purpose of manufacturing pre clinical and clinical active material. This model predicts that product yields will be the greatest when the specific growth of the culture is maintained at a steady and optimal rate. However, rOvIFN-τ yields did not meet the expected linear model but most closely corresponded to a polynomial relationship. After transitioning from glycerol to methanol, product accumulated for 31–45 h, and then the yield decreased. This production shift, which has been termed decoupling, was clearly related to time ...


Quorum Sensing In Dimorphic Fungi: Farnesol And Beyond, Kenneth W. Nickerson, Audrey L. Atkin, Jacob M. Hornby Jun 2006

Quorum Sensing In Dimorphic Fungi: Farnesol And Beyond, Kenneth W. Nickerson, Audrey L. Atkin, Jacob M. Hornby

Papers in Microbiology

Production of farnesol by Candida albicans is the first quorum- sensing system discovered in a eukaryote (29). In C. albicans, accumulated farnesol affects both dimorphism (29, 50) and biofilm formation (62). Fungal dimorphism is defined (64) as an environmentally controlled reversible interconversion of morphology, particularly yeast and mycelial morphologies. Interest in this shift derives from the dimorphic character of many fungi that are pathogenic toward plants and animals (64). Numerous chemical and environmental parameters can shift the yeast-mycelium dimorphism, including temperature, pH, glucose levels, nitrogen source, carbon dioxide levels, transition metals, chelating agents, and inoculum size or initial cell density ...


Survey Of Extreme Solvent Tolerance In Gram-Positive Cocci: Membrane Fatty Acid Changes In Staphylococcus Haemolyticus Grown In Toluene, Lindsey E. Nielsen, Dana R. Kadavy, Soumitra Rajagopal, Rhae A. Drijber, Kenneth W. Nickerson Sep 2005

Survey Of Extreme Solvent Tolerance In Gram-Positive Cocci: Membrane Fatty Acid Changes In Staphylococcus Haemolyticus Grown In Toluene, Lindsey E. Nielsen, Dana R. Kadavy, Soumitra Rajagopal, Rhae A. Drijber, Kenneth W. Nickerson

Papers in Microbiology

We exploited the unique ecological niche of oil fly larval guts to isolate a strain of Staphylococcus haemolyticus which may be the most solvent-tolerant gram-positive bacterium yet described. This organism is able to tolerate 100% toluene, benzene, and p-xylene on plate overlays and saturating levels of these solvents in monophasic liquid cultures. A comparison of membrane fatty acids by gas chromatography after growth in liquid media with and without toluene showed that in cells continuously exposed to solvent the proportion of anteiso fatty acids increased from 25.8 to 33.7% while the proportion of 20:0 straight-chain fatty ...


Farnesol Concentrations Required To Block Germ Tube Formation In Candida Albicans In The Presence And Absence Of Serum, Daniel D. Mosel, Raluca Dumitru, Jacob M. Hornby, Audrey L. Atkin, Kenneth W. Nickerson Aug 2005

Farnesol Concentrations Required To Block Germ Tube Formation In Candida Albicans In The Presence And Absence Of Serum, Daniel D. Mosel, Raluca Dumitru, Jacob M. Hornby, Audrey L. Atkin, Kenneth W. Nickerson

Papers in Microbiology

Concentrations of (E,E)-farnesol needed to inhibit germ tube formation were determined for Candida albicans strains A72 and SC5314 by using six different conditions known to trigger germination. For defined media, 1 to 2 μM farnesol was sufficient. However, with serum at 2 to 20%, up to 250 μM farnesol was required. Farnesol blocked germ tube formation but did not block elongation of existing germ tubes.


Surface Display Of Recombinant Proteins On Bacillus Thuringiensis Spores, Cheng Du, Wing C. Chan, Timothy W. Mckeithan, Kenneth W. Nickerson Jun 2005

Surface Display Of Recombinant Proteins On Bacillus Thuringiensis Spores, Cheng Du, Wing C. Chan, Timothy W. Mckeithan, Kenneth W. Nickerson

Papers in Microbiology

The insecticidal protoxin from Bacillus thuringiensis has been shown to be a major component of the spore coat. We have developed a novel surface display system using B. thuringiensis spores in which the N-terminal portion of the protoxin is replaced with a heterologous protein. The expression vector with a sporulationspecific promoter was successfully used to display green fluorescent protein and a single-chain antibody (scFv) gene that encodes anti-4-ethoxymethylene-2-phenyl-2-oxazolin-5-one (anti-phOx) antibody. The spores that carry the anti-phOx antibody can bind to phOx specifically.


Defined Anaerobic Growth Medium For Studying Candida Albicans Basic Biology And Resistance To Eight Antifungal Drugs, Raluca Dumitru, Jacob M. Hornby, Kenneth W. Nickerson Jul 2004

Defined Anaerobic Growth Medium For Studying Candida Albicans Basic Biology And Resistance To Eight Antifungal Drugs, Raluca Dumitru, Jacob M. Hornby, Kenneth W. Nickerson

Papers in Microbiology

The polymorphic fungus Candida albicans is one of the most versatile opportunistic pathogens in humans. Many organs of the human body are potential targets for infection by this pathogen, but infection is commonly localized in the gastrointestinal tract, an environment providing anaerobic growth conditions. We describe a chemically defined anaerobic growth medium for four strains of Candida albicans (A72, SC5314, MEN, and 10261). It is a defined liquid glucose-phosphate-proline growth medium supplemented with oleic acid, nicotinic acid, and ammonium chloride. The cells did not require or respond to added ergosterol. Oleic acid and nicotinic acid are growth factors which are ...


Enhanced Production Of Farnesol By Candida Albicans Treated With Four Azoles, Jacob M. Hornby, Kenneth Nickerson Jun 2004

Enhanced Production Of Farnesol By Candida Albicans Treated With Four Azoles, Jacob M. Hornby, Kenneth Nickerson

Papers in Microbiology

The dimorphic fungus Candida albicans excretes farnesol, which is produced enzymatically from the sterol biosynthetic intermediate farnesyl pyrophosphate. Inhibition of C. albicans by four azole antifungals, fluconazole, ketoconazole, miconazole, and clotrimazole, caused elevated farnesol production (10- to 45-fold). Furthermore, farnesol production occurs in both laboratory strains and clinical isolates (J. M. Hornby et al., Appl. Environ. Microbiol. 67:2982-2992, 2001) of C. albicans.


Inoculum Size Effect In Dimorphic Fungi: Extracellular Control Of Yeast-Mycelium Dimorphism In Ceratocystis Ulmi, Jacob M. Hornby, Sarah M. Jacobitz-Kizzier, Donna J. Mcneel, Ellen C. Jensen, David S. Treves, Kenneth W. Nickerson Mar 2004

Inoculum Size Effect In Dimorphic Fungi: Extracellular Control Of Yeast-Mycelium Dimorphism In Ceratocystis Ulmi, Jacob M. Hornby, Sarah M. Jacobitz-Kizzier, Donna J. Mcneel, Ellen C. Jensen, David S. Treves, Kenneth W. Nickerson

Papers in Microbiology

We studied the inoculum size effect in Ceratocystis ulmi, the dimorphic fungus that causes Dutch elm disease. In a defined glucose-proline-salts medium, cells develop as budding yeasts when inoculated at >106 spores per ml and as mycelia when inoculated at type, age of the spores, temperature, pH, oxygen availability, trace metals, sulfur source, phosphorous source, or the concentration of glucose or proline. Similarly, it was not influenced by added adenosine, reducing agents, methyl donors, amino sugars, fatty acids, or carbon dioxide. Instead, growing cells excreted an unknown quorum-sensing factor that caused a morphological shift from mycelia to budding yeasts. This ...


Phenotype Of The Triplo-Lethal Locus Of Drosophila Melanogaster And Its Suppression By Hyperoxia, Laura K. Smoyer, Douglas R. Dorer, Kenneth W. Nickerson, Alan C. Christensen Dec 2003

Phenotype Of The Triplo-Lethal Locus Of Drosophila Melanogaster And Its Suppression By Hyperoxia, Laura K. Smoyer, Douglas R. Dorer, Kenneth W. Nickerson, Alan C. Christensen

Papers in Microbiology

The Triplo-lethal locus (Tpl) of Drosophila is both triplo-lethal and haploinsufficient, but the function of the locus is unknown. We have examined Tpl-aneuploid embryos and find that, in both trisomics and monosomics, the midgut shows extensive cell death and the tracheae are abnormal. Shortly thereafter, all tissues die. PCR-based genotyping of individual embryos and larvae show that this phenotype occurs in the trisomics after hatching and in the monosomics before hatching. Weak alleles of the interacting gene Su(Tpl) delay the death of Tpl trisomics, but they still show the same tracheal and midgut phenotypes before dying. Hyperoxia (45 ...


Farnesol Biosynthesis In Candida Albicans: Cellular Response To Sterol Inhibition By Zaragozic Acid B, Jacob M. Hornby, Bessie W. Kebaara, Kenneth W. Nickerson Jul 2003

Farnesol Biosynthesis In Candida Albicans: Cellular Response To Sterol Inhibition By Zaragozic Acid B, Jacob M. Hornby, Bessie W. Kebaara, Kenneth W. Nickerson

Papers in Microbiology

The dimorphic fungus Candida albicans produces farnesol as a quorum-sensing molecule that regulates cellular morphology. The biosynthetic origin of farnesol has been resolved by treating these cells with zaragozic acid B, a potent inhibitor of squalene synthase in the sterol biosynthetic pathway. Treatment with zaragozic acid B leads to an eightfold increase in the amount of farnesol produced by C. albicans. Furthermore, C. albicans cell extracts contain enzymatic activity to convert [3H]farnesyl pyrophosphate to [3H]farnesol. Many common antifungal antibiotics (e.g., zaragozic acids, azoles, and allylamines) target steps in sterol biosynthesis. We suggest that the fungicidal ...


Sodium Dodecyl Sulfate Hypersensitivity Of Clpp And Clpb Mutants Of Escherichia Coli, Soumitra Rajagopal, Narasimhan Sudarsan, Kenneth W. Nickerson Aug 2002

Sodium Dodecyl Sulfate Hypersensitivity Of Clpp And Clpb Mutants Of Escherichia Coli, Soumitra Rajagopal, Narasimhan Sudarsan, Kenneth W. Nickerson

Papers in Microbiology

We studied the hypersensitivity of clpP and clpB mutants of Escherichia coli to sodium dodecyl sulfate (SDS). Both wild-type E. coli MC4100 and lon mutants grew in the presence of 10% SDS, whereas isogenic clpP and clpB single mutants could not grow above 0.5% SDS and clpA and clpX single mutants could not grow above 5.0% SDS. For wild-type E. coli, cellular ClpP levels as determined by Western immunoblot analysis increased ca. sixfold as the levels of added SDS increased from 0 to 2%. Capsular colanic acid, measured as uronic acid, increased ca. sixfold as the levels of ...


Membrane Glycoprotein Gp130 Of Dictyostelium Discoideum Is Lipid-Linked And Its Fate Altered In The Presence Of Tunicamycin, Betsy L. Barent, Catherine P. Chia Sep 2001

Membrane Glycoprotein Gp130 Of Dictyostelium Discoideum Is Lipid-Linked And Its Fate Altered In The Presence Of Tunicamycin, Betsy L. Barent, Catherine P. Chia

Papers in Microbiology

We are studying the structure and role in phagocytosis of gp130, a glycoprotein located on the extracellular surface of D. discoideum amoebae. Predictions from the protein sequence of gp130 deduced from the cDNA indicate its attachment to the membrane via a glycosylphosphatidylinositol (GP1)-anchor. This was confirmed when radiolabeled palmitic acid was incorporated into gp130 of axenically grown cells, and when nitrous acid deamination released gp130 from purified membranes. The GPI-anchor of gp130 resisted cleavage by bacterial and mammalian phosphoinositol-specific phospholipases. However, in the presence of protease inhibitors, we detected in vitro a time-dependent cleavage of gp 130 that was ...


Giant Vacuoles Observed In Dictyostelium Discoideum, Aidong Yuan, Catherine P. Chia Jan 2001

Giant Vacuoles Observed In Dictyostelium Discoideum, Aidong Yuan, Catherine P. Chia

Papers in Microbiology

Large intracellular vacuoles, >4 μm in diameter and either round or oval-shaped, were observed infrequently in Dictyostelium discoideum amoebae of axenically-grown strain AX2 (only 1 in 106–108 cells). These previously unreported single or multiple “giant” vacuoles were more common, however, in newly germinated KAX3 cells (0.55% of the population) and AT-Kneg, a strain that lacks an esterase (0.47% of the population). A vacuolar H+-ATPase was enriched in their membranes of intracellular giant vacuoles, indicating that the vacuoles were related possibly to both endosomes and the contractile vacuole compartment. When monitored over time, giant ...


Influence Of Infected Cell Growth State On Bacteriophage Reactivation Levels, Dana R. Kadavy, Julie J. Shaffer, Susan E. Lott, Thomas A. Wolf, Cathy E. Bolton, William H. Gallimore, Eugene L. Martin, Kenneth W. Nickerson, Tyler A. Kokjohn Dec 2000

Influence Of Infected Cell Growth State On Bacteriophage Reactivation Levels, Dana R. Kadavy, Julie J. Shaffer, Susan E. Lott, Thomas A. Wolf, Cathy E. Bolton, William H. Gallimore, Eugene L. Martin, Kenneth W. Nickerson, Tyler A. Kokjohn

Papers in Microbiology

Reactivation of UV-C-inactivated Pseudomonas aeruginosa bacteriophages D3C3, F116, G101, and UNL-1 was quantified in host cells infected during the exponential phase, during the stationary phase, and after starvation (1 day, 1 and 5 weeks) under conditions designed to detect dark repair and photoreactivation. Our experiments revealed that while the photoreactivation capacity of stationary-phase or starved cells remained about the same as that of exponential-phase cells, in some cases their capacity to support dark repair of UV-inactivated bacteriophages increased over 10-fold. This enhanced reactivation capacity was correlated with the ca. 30- fold-greater UV-C resistance of P. aeruginosa host cells that were ...


Natural Antibiotic Resistance Of Bacteria Isolated From Larvae Of The Oil Fly, Helaeomyia Petrolei, Dana R. Kadavy, Jacob M. Hornby, Terry Haverkost, Kenneth W. Nickerson Nov 2000

Natural Antibiotic Resistance Of Bacteria Isolated From Larvae Of The Oil Fly, Helaeomyia Petrolei, Dana R. Kadavy, Jacob M. Hornby, Terry Haverkost, Kenneth W. Nickerson

Papers in Microbiology

Helaeomyia petrolei (oil fly) larvae inhabit the asphalt seeps of Rancho La Brea in Los Angeles, Calif. The culturable microbial gut contents of larvae collected from the viscous oil were recently examined, and the majority (9 of 14) of the strains were identified as Providencia spp. Subsequently, 12 of the bacterial strains isolated were tested for their resistance or sensitivity to 23 commonly used antibiotics. All nine strains classified as Providencia rettgeri exhibited dramatic resistance to tetracycline, vancomycin, bacitracin, erythromycin, novobiocin, polymyxin, colistin, and nitrofurantoin. Eight of nine Providencia strains showed resistance to spectinomycin, six of nine showed resistance to ...


Integrity Of The Actin Cytoskeleton Required For Both Phagocytosis And Macropinocytosis In Dictyostelium Discoideum, Aidong Yuan, Catherine P. Chia Sep 2000

Integrity Of The Actin Cytoskeleton Required For Both Phagocytosis And Macropinocytosis In Dictyostelium Discoideum, Aidong Yuan, Catherine P. Chia

Papers in Microbiology

Filamentous (F-) actin is enriched in cellular extensions, such as phagocytic cups and macropioocytic crowns, of Dlctyostelium discoideum amebae. Previous studies of actin-disrupting agents that implicated the involvement of the actin cytoskeleton in Dictyostelium phagocytosis and pinocytosis, however, have yielded conflicting results. We show that the integrity of the actin cytoskeleton is required for both phagocytosis and macropinocytosis in D. discoideum with latrunculin A (IatA), which binds to monomeric actin, and cytochalasin A (cytA), which caps the plus end of actin filaments. Using rhodamine-phalloidin to visualize F-actin, cells treated for 30 min. with 1 to 4 pM of latA displayed ...


A Role For Calcineurin In Dictyostelium Discoideum Phagocytosis, Aidong Yuan, Catherine P. Chia Sep 2000

A Role For Calcineurin In Dictyostelium Discoideum Phagocytosis, Aidong Yuan, Catherine P. Chia

Papers in Microbiology

The Ca2+/calmodul1n-dependent protein phosphatase calcinewin is involved in the development of the cellular slime mold Dictyostelium discoideum. Because of its interactions with Ca2+, which appear to influence D. discoideum phagocytosis (Yuan and Chia, 1999, Mol. Biol. Cell 10, 220a), we undertook studies to test whether calcineurin also plays a role in Dictyostelium phagocytosis. The immunosuppressants cyclosporin A and FK506, through the formation of cyclosporin A-cyclophilin A and FK506- FK506-binding protein complexes, respectively, inhibited calcineurin activity. These two calcineurin inhibitors suppressed phagocytosis of fluorescently labeled yeast in a dose-dependent manner. Although it inhib~ted phagocytosis, cyclosporin A had an insignificant ...


A Receptor-Like Glycoprotein From Dictyostelium Discoideum: Functions In Phagocytosis And Cell Adhesion?, P. Christopher Larosa, Melissa B. Meirer, Catherine P. Chia Sep 2000

A Receptor-Like Glycoprotein From Dictyostelium Discoideum: Functions In Phagocytosis And Cell Adhesion?, P. Christopher Larosa, Melissa B. Meirer, Catherine P. Chia

Papers in Microbiology

The molecular mechanisms for the initial recognition and subsequent internalization of food and unicellular pathogens by phagocytes are incompletely understood We have hypothesized that a surface-exposed, glycosylated I30 kDa protein, gp130, that 1s concentrated on the plasma membrane and found In phagosomes, has a role In phagocytosis by D. discoideum amoebae. GpI30 appears to have a cytoskeletal association and has extracellular domains susceptible to proteolytic digestion. It is tightly bound to the plasma membrane probably via a carboxyterminal hydrophobic anchor predicted from the cDNA. Gp130 may be the same as a similarly sized protein, gp126, that was implicated as a ...


Tubulin And Neurofilament Proteins Are Transported Differently In Axons Of Chicken Motoneurons, Aidong Yuan, Roland G. Mills, Catherine P. Chia, John J. Bray Jun 2000

Tubulin And Neurofilament Proteins Are Transported Differently In Axons Of Chicken Motoneurons, Aidong Yuan, Roland G. Mills, Catherine P. Chia, John J. Bray

Papers in Microbiology

1. We previously showed that actin is transported in an unassembled form with its associated proteins actin depolymerizing factor, cofilin, and profilin. Here we examine the specific activities of radioactively labeled tubulin and neurofilament proteins in subcellular fractions of the chicken sciatic nerve following injection of L-[35S]methionine into the lumbar spinal cord. 2. At intervals of 12 and 20 days after injection, nerves were cut into 1-cm segments and separated into Triton X-100-soluble and particulate fractions. Analysis of the fractions by high-resolution two-dimensional gel electrophoresis, immunoblotting, fluorography, and computer densitometry showed that tubulin was transported as a unimodal ...


Role Of Esterase Gp70 And Its Influence On Growth And Development Of Dictyostelium Discoideum, Aidong Yuan, Catherine P. Chia Jan 2000

Role Of Esterase Gp70 And Its Influence On Growth And Development Of Dictyostelium Discoideum, Aidong Yuan, Catherine P. Chia

Papers in Microbiology

Gp70 is an esterase originally called crystal protein because of its presence in crystalline structures in aggregation-competent Dictyostelium discoideum cells. Although postulated to break down spore coats, the function of gp70 in vivo was incompletely investigated. Our immunolocalization and biochemical studies of vegetative D. discoideum amoebae show that gp70 was recruited to phagosomes and found in lysosomes. Purified gp70 was effective at hydrolyzing naphthyl substrates with acyl chains typical of lipids and lipopolysaccharides, indicating that the gp70 was involved in digesting endocytosed molecules. The activity of purified gp70 was inhibited by reductants that retarded its electrophoretic mobility and verified the ...


Microbiology Of The Oil Fly, Helaeomyia Petrolei, Dana R. Kadavy, Bradley A. Plantz, Christopher A. Shaw, Jill Myatt, Tyler A. Kokjohn, Kenneth Nickerson Apr 1999

Microbiology Of The Oil Fly, Helaeomyia Petrolei, Dana R. Kadavy, Bradley A. Plantz, Christopher A. Shaw, Jill Myatt, Tyler A. Kokjohn, Kenneth Nickerson

Papers in Microbiology

Helaeomyia petrolei larvae isolated from the asphalt seeps of Rancho La Brea in Los Angeles, Calif., were examined for microbial gut contents. Standard counts on Luria-Bertani, MacConkey, and blood agar plates indicated ca. 2 x 105 heterotrophic bacteria per larva. The culturable bacteria represented 15 to 20% of the total population as determined by acridine orange staining. The gut itself contained large amounts of the oil, had no observable ceca, and maintained a slightly acidic pH of 6.3 to 6.5. Despite the ingestion of large amounts of potentially toxic asphalt by the larvae, their guts sustained the ...


Nonspecific Interactions Alter Lipopolysaccharide Patterns And Protein Mobility On Sodium Dodecyl Sulfate Polyacrylamide Gels, Aidong Yuan, R L. Pardy, Catherine P. Chia Mar 1999

Nonspecific Interactions Alter Lipopolysaccharide Patterns And Protein Mobility On Sodium Dodecyl Sulfate Polyacrylamide Gels, Aidong Yuan, R L. Pardy, Catherine P. Chia

Papers in Microbiology

In testing whether bacterial lipopolysaccharide (LPS) was a natural substrate for an esterase from the soil amebae Dictyostelium discoideum, we observed altered banding patterns of the LPS and changed protein mobility on sodium dodecyl sulfate (SDS) polyacrylamide gels after incubation of LPS with the enzyme. The initial interpretation of these results was that the enzyme had removed ester-linked acyl chains from the LPS, leading to a change in its migration on gels. However, esterase inactivated by treatment with either dithiothreitol (DTT), heat, or SDS generated the same mobility shifts. Bovine serum albumin (BSA) also induced the same change in the ...


Cytoskeletal Association Of An Esterase In Dictyostelium Discoideum, Catherine P. Chia, Lothar Bomblies, Kristy K. Taylor Jan 1998

Cytoskeletal Association Of An Esterase In Dictyostelium Discoideum, Catherine P. Chia, Lothar Bomblies, Kristy K. Taylor

Papers in Microbiology

A 70-kDa glycoprotein, gp70, was found enriched in the detergentinsoluble cytoskeletal fraction of axenically grown Dictyostelium discoideum cells. Its N-terminal amino acid sequence identified it as “crystal protein” (L. Bomblies et al., 1990, J. Cell Biol. 110, 669– 679). This finding was corroborated when antibody to crystal protein cross-reacted with gp70 and its deglycosylated form. The postulated esterase activity of gp70/crystal protein was verified through comparative enzyme assays of extracts derived from cells that either overexpressed or lacked gp70. Gp70 cosedimented with cytoskeletons on sucrose gradients, suggesting an interaction with the cytoskeleton. Coisolation of gp70 with detergent-extracted cells, observed ...


Phagosomal Proteins Of Dictyostelium Discoideum, Betsy L. Rezabek, Juan M. Rodriguez-Paris, James A. Cardelli, Catherine P. Chia Jul 1997

Phagosomal Proteins Of Dictyostelium Discoideum, Betsy L. Rezabek, Juan M. Rodriguez-Paris, James A. Cardelli, Catherine P. Chia

Papers in Microbiology

In recognizing food particles, Dictyostelium cell-surface molecules initiate cytoskeletal rearrangements that result in phagosome formation. After feeding D. discoideum cells latex beads, early phagosomes were isolated on sucrose step gradients. Protein analyses of these vesicles showed that they contained glycoproteins and surface-labeled species corresponding to integral plasma membrane proteins. Cytoskeletal proteins also were associated with phagosomes, including myosin II, actin and a 30 kDa-actin bundling protein. As seen by the acridine orange fluorescence of vesicles containing bacteria, phagosomes were acidified rapidly by a vacuolar H+-ATPase that was detected by immunoblotting. Except for the loss of cytoskeletal proteins, few other ...


Bacillus Thuringiensis Hd-73 Spores Have Surface-Localized Cry1ac Toxin: Physiological And Pathogenic Consequences, Cheng Du, Kenneth Nickerson Oct 1996

Bacillus Thuringiensis Hd-73 Spores Have Surface-Localized Cry1ac Toxin: Physiological And Pathogenic Consequences, Cheng Du, Kenneth Nickerson

Papers in Microbiology

Spores from Cry+ strains of Bacillus thuringiensis bound fluorescein isothiocyanate-labeled antibodies specific for the 65-kDa activated Cry 1Ac toxin, whereas spores from Bacillus cereus and Cry- strains of B. thuringiensis did not. The Cry+ spores could be activated for germination by alkaline conditions (pH 10.3), whereas Cry- spores could not. Once the surrounding exosporia had been removed or permeabilized, Cry+ spores were able to bind the toxin receptor(s) from insect gut brush border membrane vesicle preparations, and their germination rates were increased ca. threefold in the presence of brush border membrane vesicles. A model is presented whereby in ...


Crystal Protein Expression During Vegetative Growth Of Dictyostelium, Kristy K. Taylor, Catherine P. Chia Sep 1996

Crystal Protein Expression During Vegetative Growth Of Dictyostelium, Kristy K. Taylor, Catherine P. Chia

Papers in Microbiology

We are studying interactions between membrane proteins and the cytoskeleton of Dictyostelium discoideum. Detergent (Triton X-100) -insoluble cytoskeletons from vegetative (AX2) amebae are enriched in a 70 kDa Concanavalin A (Con A)-binding protein (gp70). An enriched fraction of gp70 was prepared by Con A affinity chromatography of cytoskeletons from axenically-grown cells. N-terminal amino acid sequence of gel-purified gp70 established its identity to a 69 kDa 'crystal protein' previously characterized by Bomblies, et al. (1990, J. Cell Biol. 110:669 -679). Enzymatic deglycosylation of gp70 resulted in a 65 kDa product that did not bind Con A, verifying the predicted ...