scholarly journals Candida albicans Zn Cluster Transcription Factors Tac1 and Znc1 Are Activated by Farnesol To Upregulate a Transcriptional Program Including the Multidrug Efflux Pump CDR1

2018 ◽  
Vol 62 (11) ◽  
Author(s):  
Zhongle Liu ◽  
John M. Rossi ◽  
Lawrence C. Myers
Keyword(s):  
Candida Albicans ◽  
Transcription Factors ◽  
Quorum Sensing ◽  
Efflux Pump ◽  
Close Relative ◽  
Multidrug Efflux ◽  
Multidrug Efflux Pump ◽  
Content Type ◽  
Dna Binding Specificity ◽  
Hyphal Formation

ABSTRACT Farnesol, a quorum-sensing molecule, inhibits Candida albicans hyphal formation, affects its biofilm formation and dispersal, and impacts its stress response. Several aspects of farnesol's mechanism of action remain incompletely uncharacterized. Among these are a thorough accounting of the cellular receptors and transporters for farnesol. This work suggests these processes are linked through the Zn cluster transcription factors Tac1 and Znc1 and their induction of the multidrug efflux pump Cdr1. Specifically, we have demonstrated that Tac1 and Znc1 are functionally activated by farnesol through a mechanism that mimics other means of hyperactivation of Zn cluster transcription factors. This is consistent with our observation that many genes acutely induced by farnesol are dependent on TAC1, ZNC1, or both. A related molecule, 1-dodecanol, invokes a similar TAC1-ZNC1 response, while several other proposed C. albicans quorum-sensing molecules do not. Tac1 and Znc1 both bind to and upregulate the CDR1 promoter in response to farnesol. Differences in inducer and DNA binding specificity lead to Tac1 and Znc1 having overlapping, but nonidentical, regulons. Induction of genes by farnesol via Tac1 and Znc1 was inversely related to the level of CDR1 present in the cell, suggesting a model in which induction of CDR1 by Tac1 and Znc1 leads to an increase in farnesol efflux. Consistent with this premise, our results show that CDR1 expression, and its regulation by TAC1 and ZNC1, facilitates growth in the presence of high farnesol concentrations in C. albicans and in certain strains of its close relative, C. dubliniensis.

scholarly journals Differential Requirement of the Transcription Factor Mcm1 for Activation of the Candida albicans Multidrug Efflux PumpMDR1by Its Regulators Mrr1 and Cap1

2011 ◽  
Vol 55 (5) ◽  
pp. 2061-2066 ◽  
Author(s):  
Selene Mogavero ◽  
Arianna Tavanti ◽  
Sonia Senesi ◽  
P. David Rogers ◽  
Joachim Morschhäuser
Keyword(s):  
Transcription Factor ◽  
Candida Albicans ◽  
Transcription Factors ◽  
Molecular Mechanisms ◽  
Efflux Pump ◽  
Multidrug Efflux ◽  
Multidrug Efflux Pump ◽  
Gain Of Function ◽  
Pathogenic Yeast ◽  
Content Type

ABSTRACTOverexpression of the multidrug efflux pump Mdr1 causes increased fluconazole resistance in the pathogenic yeastCandida albicans. The transcription factors Mrr1 and Cap1 mediateMDR1upregulation in response to inducing stimuli, and gain-of-function mutations in Mrr1 or Cap1, which render the transcription factors hyperactive, result in constitutiveMDR1overexpression. The essential MADS box transcription factor Mcm1 also binds to theMDR1promoter, but its role in inducible or constitutiveMDR1upregulation is unknown. Using a conditional mutant in which Mcm1 can be depleted from the cells, we investigated the importance of Mcm1 forMDR1expression. We found that Mcm1 was dispensable forMDR1upregulation by H2O2but was required for fullMDR1induction by benomyl. A C-terminally truncated, hyperactive Cap1 could upregulateMDR1expression both in the presence and in the absence of Mcm1. In contrast, a hyperactive Mrr1 containing a gain-of-function mutation depended on Mcm1 to causeMDR1overexpression. These results demonstrate a differential requirement for the coregulator Mcm1 for Cap1- and Mrr1-mediatedMDR1upregulation. When activated by oxidative stress or a gain-of-function mutation, Cap1 can induceMDR1expression independently of Mcm1, whereas Mrr1 requires either Mcm1 or an active Cap1 to cause overexpression of theMDR1efflux pump. Our findings provide more detailed insight into the molecular mechanisms of drug resistance in this important human fungal pathogen.

scholarly journals C. albicans Zn Cluster Transcription Factors Tac1 and Znc1 are Activated by Farnesol to Up Regulate a Transcriptional Program Including the Multi-Drug Efflux Pump CDR1

2018 ◽  
Author(s):  
Zhongle Liu ◽  
John M. Rossi ◽  
Lawrence C. Myers
Keyword(s):  
Transcription Factors ◽  
Quorum Sensing ◽  
Biofilm Formation ◽  
Target Genes ◽  
Efflux Pump ◽  
Close Relative ◽  
Drug Efflux ◽  
Dna Binding Specificity ◽  
Drug Efflux Pump ◽  
Hyphal Formation

AbstractFarnesol, a quorum-sensing molecule, inhibits C. albicans hyphal formation, affects its biofilm formation and dispersal, and impacts its stress response. Several aspects of farnesol’s mechanism of action remain incompletely uncharacterized. Among these are a thorough accounting of the cellular receptors and transporters for farnesol. This work suggests these themes are linked through the Zn cluster transcription factors Tac1 and Znc1, and their induction of the multi-drug efflux pump Cdr1. Specifically, we have demonstrated that Tac1 and Znc1 are functionally activated by farnesol through a mechanism that mimics other means of hyperactivation of Zn cluster transcription factors. This is consistent with our observation that many genes acutely induced by farnesol are dependent on TAC1, ZNC1, or both. A related molecule, 1-dodecanol, invokes a similar TAC1/ZNC1 response, while several other proposed C. albicans quorum sensing molecules do not. TAC1 and ZNC1 both bind to and up-regulate the CDR1 promoter in response to farnesol. Differences in inducer and DNA binding specificity lead to Tac1 and Znc1 having overlapping, but non-identical, regulons. TAC1 and ZNC1 dependent farnesol induction of their target genes was inversely related to the level of CDR1 present in the cell, suggesting a model in which induction of CDR1 by Tac1 and Znc1 leads to an increase in farnesol efflux. Consistent with this premise, our results show that CDR1 expression, and its regulation by TAC1 and ZNC1, facilitates growth in the presence of high farnesol concentrations in C. albicans, and certain strains of its close relative C. dubliniensis.

scholarly journals Inducible and Constitutive Activation of Two Polymorphic Promoter Alleles of the Candida albicans Multidrug Efflux PumpMDR1

2012 ◽  
Vol 56 (8) ◽  
pp. 4490-4494 ◽  
Author(s):  
Christoph Sasse ◽  
Rebecca Schillig ◽  
Alexandra Reimund ◽  
Julia Merk ◽  
Joachim Morschhäuser
Keyword(s):  
Candida Albicans ◽  
Transcription Factors ◽  
Loss Of Heterozygosity ◽  
Efflux Pump ◽  
Multidrug Efflux ◽  
Multidrug Efflux Pump ◽  
Fluconazole Resistance ◽  
Content Type ◽  
Constitutive Activation ◽  
Strain Sc5314

ABSTRACTOverexpression of the multidrug efflux pumpMDR1confers resistance to the antifungal drug fluconazole onCandida albicans. It has been reported that two types ofMDR1promoters exist inC. albicansand that homozygosity for the allele with higher activity may promote fluconazole resistance. We found that the twoMDR1promoter alleles in strain SC5314 were equally well activated by inducing chemicals or hyperactive forms of the transcription factors Mrr1 and Cap1, which controlMDR1expression. In addition, no loss of heterozygosity at theMDR1locus was observed inMDR1-overexpressing clinicalC. albicansstrains that developed fluconazole resistance during therapy.

scholarly journals SAGA/ADA Complex Subunit Ada2 Is Required for Cap1- but Not Mrr1-Mediated Upregulation of the Candida albicans Multidrug Efflux PumpMDR1

2014 ◽  
Vol 58 (9) ◽  
pp. 5102-5110 ◽  
Author(s):  
Bernardo Ramírez-Zavala ◽  
Selene Mogavero ◽  
Eva Schöller ◽  
Christoph Sasse ◽  
P. David Rogers ◽  
...  
Keyword(s):  
Transcription Factor ◽  
Candida Albicans ◽  
Efflux Pump ◽  
Wild Type ◽  
Multidrug Efflux ◽  
Multidrug Efflux Pump ◽  
Gain Of Function ◽  
Content Type ◽  
Zinc Cluster ◽  
Fluconazole Resistant

ABSTRACTOverexpression of the multidrug efflux pumpMDR1is one mechanism by which the pathogenic yeastCandida albicansdevelops resistance to the antifungal drug fluconazole. The constitutive upregulation ofMDR1in fluconazole-resistant, clinicalC. albicansisolates is caused by gain-of-function mutations in the zinc cluster transcription factor Mrr1. It has been suggested that Mrr1 activatesMDR1transcription by recruiting Ada2, a subunit of the SAGA/ADA coactivator complex. However,MDR1expression is also regulated by the bZIP transcription factor Cap1, which mediates the oxidative stress response inC. albicans. Here, we show that a hyperactive Mrr1 containing a gain-of-function mutation promotesMDR1overexpression independently of Ada2. In contrast, a C-terminally truncated, hyperactive Cap1 causedMDR1overexpression in a wild-type strain but only weakly in mutants lackingADA2. In the presence of benomyl or H2O2, compounds that induceMDR1expression in an Mrr1- and Cap1-dependent fashion,MDR1was upregulated with the same efficiency in wild-type andada2Δ cells. These results indicate that Cap1, but not Mrr1, recruits Ada2 to theMDR1promoter to induce the expression of this multidrug efflux pump and that Ada2 is not required forMDR1overexpression in fluconazole-resistantC. albicansstrains containing gain-of-function mutations in Mrr1.

scholarly journals The Quorum-Sensing Molecule Farnesol Is a Modulator of Drug Efflux Mediated by ABC Multidrug Transporters and Synergizes with Drugs in Candida albicans

2011 ◽  
Vol 55 (10) ◽  
pp. 4834-4843 ◽  
Author(s):  
Monika Sharma ◽  
Rajendra Prasad
Keyword(s):  
Candida Albicans ◽  
Quorum Sensing ◽  
Efflux Pump ◽  
Major Facilitator Superfamily ◽  
Simultaneous Increase ◽  
Drug Efflux ◽  
Multidrug Efflux Pump ◽  
Content Type ◽  
Multidrug Transporters ◽  
Quorum Sensing Molecule

ABSTRACTOverexpression of theCaCDR1-encoded multidrug efflux pump protein CaCdr1p (Candidadrug resistance protein 1), belonging to the ATP binding cassette (ABC) superfamily of transporters, is one of the most prominent contributors of multidrug resistance (MDR) inCandida albicans. Thus, blocking or modulating the function of the drug efflux pumps represents an attractive approach in combating MDR. In the present study, we provide first evidence that the quorum-sensing molecule farnesol (FAR) is a specific modulator of efflux mediated by ABC multidrug transporters, such as CaCdr1p and CaCdr2p ofC. albicansand ScPdr5p ofSaccharomyces cerevisiae. Interestingly, FAR did not modulate the efflux mediated by the multidrug extrusion pump protein CaMdr1p, belonging to the major facilitator superfamily (MFS). Kinetic data revealed that FAR competitively inhibited rhodamine 6G efflux in CaCdr1p-overexpressing cells, with a simultaneous increase in an apparentKmwithout affecting theVmaxvalues and the ATPase activity. We also observed that when used in combination, FAR at a nontoxic concentration synergized with the drugs at their respective nonlethal concentrations, as was evident from their <0.5 fractional inhibitory concentration index (FICI) values and from the drop of 14- to 64-fold in the MIC80values in the wild-type strain and in azole-resistant clinical isolates ofC. albicans. Our biochemical experiments revealed that the synergistic interaction of FAR with the drugs led to reactive oxygen species accumulation, which triggered early apoptosis, and that both could be partly reversed by the addition of an antioxidant. Collectively, FAR modulates drug extrusion mediated exclusively by ABC proteins and is synergistic to fluconazole (FLC), ketoconazole (KTC), miconazole (MCZ), and amphotericin (AMB).

scholarly journals Cryo-EM Structures of a Gonococcal Multidrug Efflux Pump Illuminate a Mechanism of Drug Recognition and Resistance

mBio ◽  
2020 ◽  
Vol 11 (3) ◽  
Author(s):  
Meinan Lyu ◽  
Mitchell A. Moseng ◽  
Jennifer L. Reimche ◽  
Concerta L. Holley ◽  
Vijaya Dhulipala ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Drug Resistance ◽  
Neisseria Gonorrhoeae ◽  
Antimicrobial Agents ◽  
Efflux Pump ◽  
Multidrug Efflux ◽  
Multidrug Efflux Pump ◽  
Transmitted Infection ◽  
Content Type ◽  
Cryo Electron Microscopy

ABSTRACT Neisseria gonorrhoeae is an obligate human pathogen and causative agent of the sexually transmitted infection (STI) gonorrhea. The most predominant and clinically important multidrug efflux system in N. gonorrhoeae is the multiple transferrable resistance (Mtr) pump, which mediates resistance to a number of different classes of structurally diverse antimicrobial agents, including clinically used antibiotics (e.g., β-lactams and macrolides), dyes, detergents and host-derived antimicrobials (e.g., cationic antimicrobial peptides and bile salts). Recently, it has been found that gonococci bearing mosaic-like sequences within the mtrD gene can result in amino acid changes that increase the MtrD multidrug efflux pump activity, probably by influencing antimicrobial recognition and/or extrusion to elevate the level of antibiotic resistance. Here, we report drug-bound solution structures of the MtrD multidrug efflux pump carrying a mosaic-like sequence using single-particle cryo-electron microscopy, with the antibiotics bound deeply inside the periplasmic domain of the pump. Through this structural approach coupled with genetic studies, we identify critical amino acids that are important for drug resistance and propose a mechanism for proton translocation. IMPORTANCE Neisseria gonorrhoeae has become a highly antimicrobial-resistant Gram-negative pathogen. Multidrug efflux is a major mechanism that N. gonorrhoeae uses to counteract the action of multiple classes of antibiotics. It appears that gonococci bearing mosaic-like sequences within the gene mtrD, encoding the most predominant and clinically important transporter of any gonococcal multidrug efflux pump, significantly elevate drug resistance and enhance transport function. Here, we report cryo-electron microscopy (EM) structures of N. gonorrhoeae MtrD carrying a mosaic-like sequence that allow us to understand the mechanism of drug recognition. Our work will ultimately inform structure-guided drug design for inhibiting these critical multidrug efflux pumps.

scholarly journals Control of Biofilm Formation in Healthcare: Recent Advances Exploiting Quorum-Sensing Interference Strategies and Multidrug Efflux Pump Inhibitors

Materials ◽  
2018 ◽  
Vol 11 (9) ◽  
pp. 1676 ◽  
Author(s):  
Bindu Subhadra ◽  
Dong Kim ◽  
Kyungho Woo ◽  
Surya Surendran ◽  
Chul Choi
Keyword(s):  
Quorum Sensing ◽  
Biofilm Formation ◽  
Efflux Pump ◽  
Financial Burden ◽  
Multidrug Efflux ◽  
Multidrug Efflux Pump ◽  
Persistent Infections ◽  
Healthcare Settings ◽  
Recent Advances ◽  
Multidrug Efflux Pumps

Biofilm formation in healthcare is an issue of considerable concern, as it results in increased morbidity and mortality, imposing a significant financial burden on the healthcare system. Biofilms are highly resistant to conventional antimicrobial therapies and lead to persistent infections. Hence, there is a high demand for novel strategies other than conventional antibiotic therapies to control biofilm-based infections. There are two approaches which have been employed so far to control biofilm formation in healthcare settings: one is the development of biofilm inhibitors based on the understanding of the molecular mechanism of biofilm formation, and the other is to modify the biomaterials which are used in medical devices to prevent biofilm formation. This review will focus on the recent advances in anti-biofilm approaches by interrupting the quorum-sensing cellular communication system and the multidrug efflux pumps which play an important role in biofilm formation. Research efforts directed towards these promising strategies could eventually lead to the development of better anti-biofilm therapies than the conventional treatments.

scholarly journals Multidrug Adaptive Resistance of Pseudomonas aeruginosa Swarming Cells

2019 ◽  
Vol 64 (3) ◽  
Author(s):  
Shannon R. Coleman ◽  
Travis Blimkie ◽  
Reza Falsafi ◽  
Robert E. W. Hancock
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Antibiotic Resistance ◽  
Efflux Pump ◽  
Iron Acquisition ◽  
Polymyxin B ◽  
Rna Seq ◽  
Multidrug Efflux ◽  
Multidrug Efflux Pump ◽  
Content Type ◽  
Adaptive Resistance

ABSTRACT Swarming surface motility is a complex adaptation leading to multidrug antibiotic resistance and virulence factor production in Pseudomonas aeruginosa. Here, we expanded previous studies to demonstrate that under swarming conditions, P. aeruginosa PA14 is more resistant to multiple antibiotics, including aminoglycosides, β-lactams, chloramphenicol, ciprofloxacin, tetracycline, trimethoprim, and macrolides, than swimming cells, but is not more resistant to polymyxin B. We investigated the mechanism(s) of swarming-mediated antibiotic resistance by examining the transcriptomes of swarming cells and swarming cells treated with tobramycin by transcriptomics (RNA-Seq) and reverse transcriptase quantitative PCR (qRT-PCR). RNA-Seq of swarming cells (versus swimming) revealed 1,581 dysregulated genes, including 104 transcriptional regulators, two-component systems, and sigma factors, numerous upregulated virulence and iron acquisition factors, and downregulated ribosomal genes. Strain PA14 mutants in resistome genes that were dysregulated under swarming conditions were tested for their ability to swarm in the presence of tobramycin. In total, 41 mutants in genes dysregulated under swarming conditions were shown to be more resistant to tobramycin under swarming conditions, indicating that swarming-mediated tobramycin resistance was multideterminant. Focusing on two genes downregulated under swarming conditions, both prtN and wbpW mutants were more resistant to tobramycin, while the prtN mutant was additionally resistant to trimethoprim under swarming conditions; complementation of these mutants restored susceptibility. RNA-Seq of swarming cells treated with subinhibitory concentrations of tobramycin revealed the upregulation of the multidrug efflux pump MexXY and downregulation of virulence factors.

scholarly journals A New Natural Product Analog of Blasticidin S Reveals Cellular Uptake Facilitated by the NorA Multidrug Transporter

2017 ◽  
Vol 61 (6) ◽  
Author(s):  
Jack R. Davison ◽  
Katheryn M. Lohith ◽  
Xiaoning Wang ◽  
Kostyantyn Bobyk ◽  
Sivakoteswara R. Mandadapu ◽  
...  
Keyword(s):  
Efflux Pump ◽  
Bacterial Strains ◽  
Multidrug Efflux ◽  
Multidrug Efflux Pump ◽  
Gene Encoding ◽  
Content Type ◽  
Bacterial Membranes ◽  
Development Of Resistance ◽  
Antibiotic Compounds ◽  
Blasticidin S

ABSTRACT The permeation of antibiotics through bacterial membranes to their target site is a crucial determinant of drug activity but in many cases remains poorly understood. During screening efforts to discover new broad-spectrum antibiotic compounds from marine sponge samples, we identified a new analog of the peptidyl nucleoside antibiotic blasticidin S that exhibited up to 16-fold-improved potency against a range of laboratory and clinical bacterial strains which we named P10. Whole-genome sequencing of laboratory-evolved strains of Staphylococcus aureus resistant to blasticidin S and P10, combined with genome-wide assessment of the fitness of barcoded Escherichia coli knockout strains in the presence of the antibiotics, revealed that restriction of cellular access was a key feature in the development of resistance to this class of drug. In particular, the gene encoding the well-characterized multidrug efflux pump NorA was found to be mutated in 69% of all S. aureus isolates resistant to blasticidin S or P10. Unexpectedly, resistance was associated with inactivation of norA, suggesting that the NorA transporter facilitates cellular entry of peptidyl nucleosides in addition to its known role in the efflux of diverse compounds, including fluoroquinolone antibiotics.

scholarly journals Molecular Mechanism of MBX2319 Inhibition of Escherichia coli AcrB Multidrug Efflux Pump and Comparison with Other Inhibitors

2014 ◽  
Vol 58 (10) ◽  
pp. 6224-6234 ◽  
Author(s):  
Attilio V. Vargiu ◽  
Paolo Ruggerone ◽  
Timothy J. Opperman ◽  
Son T. Nguyen ◽  
Hiroshi Nikaido
Keyword(s):  
Efflux Pump ◽  
Efflux Pumps ◽  
Multidrug Efflux ◽  
Multidrug Efflux Pump ◽  
Efflux Pump Inhibitor ◽  
Content Type ◽  
X Ray Crystallography ◽  
Resistance Nodulation Division ◽  
Hydrophobic Portion ◽  
Dynamics Simulations

ABSTRACTEfflux pumps of the resistance nodulation division (RND) superfamily, such as AcrB, make a major contribution to multidrug resistance in Gram-negative bacteria. The development of inhibitors of the RND pumps would improve the efficacy of current and next-generation antibiotics. To date, however, only one inhibitor has been cocrystallized with AcrB. Thus,in silicostructure-based analysis is essential for elucidating the interaction between other inhibitors and the efflux pumps. In this work, we used computer docking and molecular dynamics simulations to study the interaction between AcrB and the compound MBX2319, a novel pyranopyridine efflux pump inhibitor with potent activity against RND efflux pumps ofEnterobacteriaceaespecies, as well as other known inhibitors (D13-9001, 1-[1-naphthylmethyl]-piperazine, and phenylalanylarginine-β-naphthylamide) and the binding of doxorubicin to the efflux-defective F610A variant of AcrB. We also analyzed the binding of a substrate, minocycline, for comparison. Our results show that MBX2319 binds very tightly to the lower part of the distal pocket in the B protomer of AcrB, strongly interacting with the phenylalanines lining the hydrophobic trap, where the hydrophobic portion of D13-9001 was found to bind by X-ray crystallography. Additionally, MBX2319 binds to AcrB in a manner that is similar to the way in which doxorubicin binds to the F610A variant of AcrB. In contrast, 1-(1-naphthylmethyl)-piperazine and phenylalanylarginine-β-naphthylamide appear to bind to somewhat different areas of the distal pocket in the B protomer of AcrB than does MBX2319. However, all inhibitors (except D13-9001) appear to distort the structure of the distal pocket, impairing the proper binding of substrates.

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