scholarly journals Plasma Membrane Potential of Candida albicans Measured by Di-4-ANEPPS Fluorescence Depends on Growth Phase and Regulatory Factors

2019 ◽  
Vol 7 (4) ◽  
pp. 110 ◽  
Author(s):  
Suchodolski ◽  
Krasowska
Keyword(s):  
Candida Albicans ◽  
Plasma Membrane ◽  
Membrane Potential ◽  
Cell Membrane ◽  
Growth Phase ◽  
Fungal Pathogen ◽  
Physiological State ◽  
External Factors ◽  
Good Tool ◽  
Opportunistic Fungal Pathogen

The potential of the plasma membrane (Δѱ) regulates the electrochemical potential between the outer and inner sides of cell membranes. The opportunistic fungal pathogen, Candida albicans, regulates the membrane potential in response to environmental conditions, as well as the physiological state of the cell. Here we demonstrate a new method for detection of cell membrane depolarization/permeabilization in C. albicans using the potentiometric zwitterionic dye di-4-ANEPPS. Di-4-ANEPPS measures the changes in the cell Δѱ depending on the phases of growth and external factors regulating Δѱ, such as potassium or calcium chlorides, amiodarone or DM-11 (inhibitor of H+-ATPase). We also demonstrated that di-4-ANEPPS is a good tool for fast measurement of the influence of amphipathic compounds on Δѱ.

scholarly journals High-Throughput Nano-Biofilm Microarray for Antifungal Drug Discovery

mBio ◽  
2013 ◽  
Vol 4 (4) ◽  
Author(s):  
Anand Srinivasan ◽  
Kai P. Leung ◽  
Jose L. Lopez-Ribot ◽  
Anand K. Ramasubramanian
Keyword(s):  
Cell Culture ◽  
Candida Albicans ◽  
Drug Discovery ◽  
High Throughput ◽  
Fungal Pathogen ◽  
Microarray Platform ◽  
Microbial Cell ◽  
Content Type ◽  
Petri Dishes ◽  
Opportunistic Fungal Pathogen

ABSTRACT Micro- and nanoscale technologies have radically transformed biological research from genomics to tissue engineering, with the relative exception of microbial cell culture, which is still largely performed in microtiter plates and petri dishes. Here, we present nanoscale culture of the opportunistic fungal pathogen Candida albicans on a microarray platform. The microarray consists of 1,200 individual cultures of 30 nl of C. albicans biofilms (“nano-biofilms”) encapsulated in an inert alginate matrix. We demonstrate that these nano-biofilms are similar to conventional macroscopic biofilms in their morphological, architectural, growth, and phenotypic characteristics. We also demonstrate that the nano-biofilm microarray is a robust and efficient tool for accelerating the drug discovery process: (i) combinatorial screening against a collection of 28 antifungal compounds in the presence of immunosuppressant FK506 (tacrolimus) identified six drugs that showed synergistic antifungal activity, and (ii) screening against the NCI challenge set small-molecule library identified three heretofore-unknown hits. This cell-based microarray platform allows for miniaturization of microbial cell culture and is fully compatible with other high-throughput screening technologies. IMPORTANCE Microorganisms are typically still grown in petri dishes, test tubes, and Erlenmeyer flasks in spite of the latest advances in miniaturization that have benefitted other allied research fields, including genomics and proteomics. Culturing microorganisms in small scale can be particularly valuable in cutting down time, cost, and reagent usage. This paper describes the development, characterization, and application of nanoscale culture of an opportunistic fungal pathogen, Candida albicans. Despite a more than 2,000-fold reduction in volume, the growth characteristics and drug response profiles obtained from the nanoscale cultures were comparable to the industry standards. The platform also enabled rapid identification of new drug candidates that were effective against C. albicans biofilms, which are a major cause of mortality in hospital-acquired infections.

The hematoregulatory peptide, SK&F 107647, in combination with antifungal therapy in murine Candida albicans infections

1995 ◽  
Vol 73 (S1) ◽  
pp. 1199-1205 ◽  
Author(s):  
Peter L. DeMarsh ◽  
Scott K. Sucoloski ◽  
Carrie L. Frey ◽  
Louis M. Pelus ◽  
Pradip K. Bhatnagar ◽  
...  
Keyword(s):  
Candida Albicans ◽  
Amphotericin B ◽  
Antifungal Therapy ◽  
Fungal Pathogen ◽  
Prophylactic Treatment ◽  
Lethal Dose ◽  
Survival Rates ◽  
Low Molecular Weight ◽  
Opportunistic Fungal Pathogen ◽  
Immunosuppressed Mice

SK&F 107647, a novel synthetic low molecular weight peptide, has been shown to be a potent hematoregulatory agent. We have previously demonstrated that SK&F 107647 administration can prolong survival in both immunosuppressed and normal mice challenged with the opportunistic fungal pathogen Candida albicans. Additionally, we have determined the effect of prophylactic SK&F 107647 treatment combined with conventional antifungal therapy on the survival of mice challenged with a lethal dose of C. albicans. Prophylactic treatment with SK&F 107647 or therapeutic treatment with the antifungals fluconazole or amphotericin B significantly increased the survival rates of immunosuppressed mice challenged with a lethal dose of C. albicans. However, the combination of SK&F 107647 treatment followed by antifungal therapy resulted in statistically significant increases in survival over that observed with either therapy alone. These results indicated that the hematoregulatory factor(s) elicited by SK&F 107647 enhance the survival of mice treated with conventional therapies in a model of experimental systemic candidiasis. Key words: SK&F 107647, Candida albicans, hematoregulatory, fluconazole, amphotericin B.

scholarly journals Rac1 Dynamics in the Human Opportunistic Fungal Pathogen Candida albicans

PLoS ONE ◽  
2010 ◽  
Vol 5 (10) ◽  
pp. e15400 ◽  
Author(s):  
Romain Vauchelles ◽  
Danièle Stalder ◽  
Thomas Botton ◽  
Robert A. Arkowitz ◽  
Martine Bassilana
Keyword(s):  
Candida Albicans ◽  
Fungal Pathogen ◽  
Opportunistic Fungal Pathogen

scholarly journals Ascorbic Acid Inhibition of Candida albicans Hsp90-Mediated Morphogenesis Occurs via the Transcriptional Regulator Upc2

2014 ◽  
Vol 13 (10) ◽  
pp. 1278-1289 ◽  
Author(s):  
Frédérique Van Hauwenhuyse ◽  
Alessandro Fiori ◽  
Patrick Van Dijck
Keyword(s):  
Ascorbic Acid ◽  
Candida Albicans ◽  
Fungal Pathogen ◽  
Transcriptional Regulator ◽  
Hsp90 Inhibitor ◽  
Ergosterol Biosynthesis ◽  
Content Type ◽  
Temperature Changes ◽  
Opportunistic Fungal Pathogen ◽  
Hsp90 Function

ABSTRACTMorphogenetic transitions of the opportunistic fungal pathogenCandida albicansare influenced by temperature changes, with induction of filamentation upon a shift from 30 to 37°C. Hsp90 was identified as a major repressor of an elongated cell morphology at low temperatures, as treatment with specific inhibitors of Hsp90 results in elongated growth forms at 30°C. Elongated growth resulting from a compromised Hsp90 is considered neither hyphal nor pseudohyphal growth. It has been reported that ascorbic acid (vitamin C) interferes with the yeast-to-hypha transition inC. albicans. In the present study, we show that ascorbic acid also antagonizes the morphogenetic change caused by hampered Hsp90 function. Further analysis revealed that Upc2, a transcriptional regulator of genes involved in ergosterol biosynthesis, and Erg11, the target of azole antifungals, whose expression is in turn regulated by Upc2, are required for this antagonism. Ergosterol levels correlate with elongated growth and are reduced in cells treated with the Hsp90 inhibitor geldanamycin (GdA) and restored by cotreatment with ascorbic acid. In addition, we show that Upc2 appears to be required for ascorbic acid-mediated inhibition of the antifungal activity of fluconazole. These results identify Upc2 as a major regulator of ascorbic acid-induced effects inC. albicansand suggest an association between ergosterol content and elongated growth upon Hsp90 compromise.

Dual-color quantum dot detection of a heterotetrameric potassium channel (hKCa3.1)

2011 ◽  
Vol 300 (4) ◽  
pp. C843-C849 ◽  
Author(s):  
Daniel E. J. Waschk ◽  
Anke Fabian ◽  
Thomas Budde ◽  
Albrecht Schwab
Keyword(s):  
Quantum Dots ◽  
Plasma Membrane ◽  
Membrane Potential ◽  
Cell Membrane ◽  
Quantum Dot ◽  
Fluorescence Microscopy ◽  
Potassium Channel ◽  
Binomial Distribution ◽  
Hek293 Cells ◽  
F Cells

Potassium channels play a key role in establishing the cell membrane potential and are expressed ubiquitously. Today, more than 70 mammalian K+ channel genes are known. The diversity of K+ channels is further increased by the fact that different K+ channel family members may assemble to form heterotetramers. We present a method based on fluorescence microscopy to determine the subunit composition of a tetrameric K+ channel. We generated artificial “heteromers” of the K+ channel hKCa3.1 by coexpressing two differently tagged hKCa3.1 constructs containing either an extracellular hemagglutinin (HA) or an intracellular V5 epitope. hKCa3.1 channel subunits were detected in the plasma membrane of MDCK-F cells or HEK293 cells by labeling the extra- and intracellular epitopes with differently colored quantum dots (QDs). As previously shown for the extracellular part of hKCa3.1 channels, its intracellular domain can also bind only one QD label at a time. When both channel subunits were coexpressed, 27.5 ± 1.8% and 24.9 ± 2.1% were homotetramers consisting of HA- and V5-tagged subunits, respectively. 47.6 ± 3.2% of the channels were heteromeric and composed of both subunits. The frequency distribution of HA- and V5-tagged homo- and heteromeric hKCa3.1 channels is reminiscent of the binomial distribution ( a + b)2 = a2 + 2 ab + b2. Along these lines, our findings are consistent with the notion that hKCa3.1 channels are assembled from two homomeric dimers and not randomly from four independent subunits. We anticipate that our technique will be applicable to other heteromeric membrane proteins, too.

scholarly journals Drosophila melanogaster Thor and Response to Candida albicans Infection

2007 ◽  
Vol 6 (4) ◽  
pp. 658-663 ◽  
Author(s):  
A. Levitin ◽  
A. Marcil ◽  
G. Tettweiler ◽  
M. J. Laforest ◽  
U. Oberholzer ◽  
...  
Keyword(s):  
Innate Immunity ◽  
Drosophila Melanogaster ◽  
Candida Albicans ◽  
Host Defense ◽  
Translational Control ◽  
Fungal Pathogen ◽  
Transcriptional Profiling ◽  
S2 Cells ◽  
Candida Albicans Infection ◽  
Opportunistic Fungal Pathogen

ABSTRACT We used Drosophila melanogaster macrophage-like Schneider 2 (S2) cells as a model to study cell-mediated innate immunity against infection by the opportunistic fungal pathogen Candida albicans. Transcriptional profiling of S2 cells coincubated with C. albicans cells revealed up-regulation of several genes. One of the most highly up-regulated genes during this interaction is the D. melanogaster translational regulator 4E-BP encoded by the Thor gene. Analysis of Drosophila 4E-BP null mutant survival upon infection with C. albicans showed that 4E-BP plays an important role in host defense, suggesting a role for translational control in the D. melanogaster response to C. albicans infection.

Immunoelectron microscopic detection of mannan-antigen on yeast cells of Candida albicans

1992 ◽  
Vol 50 (1) ◽  
pp. 822-823
Author(s):  
Mercedes Edwards ◽  
Robert Trimble ◽  
William Samsonoff
Keyword(s):  
Cell Wall ◽  
Candida Albicans ◽  
Fungal Pathogen ◽  
Yeast Cells ◽  
Systemic Candidiasis ◽  
Worldwide Distribution ◽  
Circulating Antigen ◽  
Infection Mechanisms ◽  
Opportunistic Fungal Pathogen

Candida albicans (Ca) is an opportunistic fungal pathogen of worldwide distribution. In recent years, infections by this fungus have received increased attention due to their prevalence in immunocomproimsed hosts (e.g. AIDS patients). However, Ca infection mechanisms are still to be ascertained. Investigations on the antigenicity of this organism have found that mannan, a component of its cell wall, is a major circulating antigen in systemic candidiasis. A few previous reports using electron microscopy, have indicated that mannan is located at the cell wall, but the resolution of the published micrographs did not afford a conclusive determination of the antigen distribution.

scholarly journals Arginine-Induced Germ Tube Formation in Candida albicans Is Essential for Escape from Murine Macrophage Line RAW 264.7

2009 ◽  
Vol 77 (4) ◽  
pp. 1596-1605 ◽  
Author(s):  
Suman Ghosh ◽  
Dhammika H. M. L. P. Navarathna ◽  
David D. Roberts ◽  
Jake T. Cooper ◽  
Audrey L. Atkin ◽  
...  
Keyword(s):  
Candida Albicans ◽  
Fungal Pathogen ◽  
Germ Tube ◽  
Tube Formation ◽  
Raw 264.7 ◽  
Dependent Manner ◽  
Wild Type ◽  
Normal Flora ◽  
Opportunistic Fungal Pathogen ◽  
Germ Tubes

ABSTRACT The opportunistic fungal pathogen Candida albicans is a part of the normal flora but it also causes systemic candidiasis if it reaches the bloodstream. Upon being phagocytized by macrophages, an important component of innate immunity, C. albicans rapidly upregulates a set of arginine biosynthetic genes. Arginine, urea, and CO2 induced hyphae in a density-dependent manner in wild-type, cph1/cph1, and rim101/rim101 strains but not in efg1/efg1 or cph1/cph1 efg1/efg1 strains. Arginase (Car1p) converts arginine to urea, which in turn is degraded by urea amidolyase (Dur1,2p) to produce CO2, a signal for hyphal switching. We used a dur1,2/dur1,2 mutant (KWN6) and the complemented strain, KWN8 (dur1,2/dur1,2::DUR1,2/DUR1,2) to study germ tube formation. KWN6 could not make germ tubes in the presence of arginine or urea but did in the presence of 5% CO2, which bypasses Dur1,2p. We also tested the effect of arginine on the interaction between the macrophage line RAW 264.7 and several strains of C. albicans. Arginine activated an Efg1p-dependent yeast-to-hypha switch, enabling wild-type C. albicans and KWN8 to escape from macrophages within 6 h, whereas KWN6 was defective in this regard. Additionally, two mutants that cannot synthesize arginine, BWP17 and SN152, were defective in making hyphae inside the macrophages, whereas the corresponding arginine prototrophs, DAY286 and SN87, formed germ tubes and escaped from macrophages. Therefore, metabolism of arginine by C. albicans controls hyphal switching and provides an important mechanism for escaping host defense.

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