Assessing risk factors for systemic fungal infections

Trehalose, a nonreducing disaccharide which accumulates dramatically during stationary phase or under oxidative stress, is well known as a stress protectant in several organisms. Here we investigated the putative correlation of trehalose with Cap1p, which is a basic region-leucine zipper (bZip) transcription factor participating in oxidative stress tolerance in Candida albicans. HPLC-MS analysis showed that trehalose did not accumulate in the cap1/cap1 mutant during stationary phase. When the mutant was exposed to high concentration of H 2 O 2 , trehalose accumulation was still not induced. Under both of the conditions above, the cap1/cap1 mutant showed high sensitivity to H 2 O 2 , and the cell viability was rather low. Furthermore, when exogenous trehalose was added to the culture of the cap1/cap1 mutant, the tolerance of this strain to oxidative stress was increased. Real time reverse transcription polymerase chain reaction (RT-PCR) analysis revealed that the transcript levels of TPS2 and TPS3 were increased in the wild type strain compared to that in cap1/cap1 mutant when exposed to H 2 O 2 . These results indicated that trehalose accumulation is important to the oxidative stress tolerance mediated by Cap1p in C. albicans.

Section: Level Of the Trehalose Content And Cell Viability In Responsmentioning

confidence: 99%

“…1,2) A major mechanism of the host defense system against fungal infection is via production of reactive oxygen species (ROS) by phagocytes to kill phagocytosed fungal cells. 3) Moreover, ROS generation is involved in the mechanisms of many antifungal agents.…”

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confidence: 99%

Trehalose Is an Important Mediator of Cap1p Oxidative Stress Response in Candida albicans

Cao

Wang

Dai

et al. 2008

“…[1][2][3] Triazole antifungal agents, such as fluconazole (FLC) and itraconazole, are considered to be the first-choice agents for treatment and prophylaxis because of their relatively low side effects and high effectiveness on mucosal infections. However, with prolonged exposure to azoles, drug resistance becomes a serious problem in clinical practice.…”

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confidence: 99%

Berberine Inhibits Fluphenazine-Induced Up-Regulation of <i>CDR1</i> in <i>Candida albicans</i>

Zhu

Yan

Zhang

et al. 2014

Over-expression of the Candida drug resistance gene CDR1 is a common mechanism generating azoleresistant Candida albicans in clinical isolates. CDR1 is transcriptionally activated through the binding of the transcription factor Tac1p to the cis-acting drug-responsive element (DRE) in its promoter. We previously demonstrated that the combination of fluconazole (FLC) and berberine (BBR) produced significant synergy when used against FLC-resistant C. albicans in vitro. In this study, we found that BBR inhibited both the up-regulation of CDR1 mRNA and the transport function of Cdr1p induced by fluphenazine (FNZ). Further, electrophoretic mobility shift assays suggested that the transcription activation complex of protein-DRE was disrupted by BBR, and electrospray ionization mass spectrometry analysis showed that BBR bound to the DRE of CDR1. Thus we propose that BBR inhibits the FNZ-induced transcriptional activation of CDR1 in C. albicans by blocking transcription factor binding to the DRE of CDR1. These results contribute to our understanding of the mechanism of synergistic effect of BBR and FLC. Key words Candida albicans; berberine; CDR1; drug-responsive element; fluphenazineCandida albicans is one of the major opportunistic fungal pathogens in humans, and causes candidiasis ranging from superficial mucosal infections to life-threatening systemic disorders.1-3) Triazole antifungal agents, such as fluconazole (FLC) and itraconazole, are considered to be the first-choice agents for treatment and prophylaxis because of their relatively low side effects and high effectiveness on mucosal infections. However, with prolonged exposure to azoles, drug resistance becomes a serious problem in clinical practice. 4,5) Several genes have been confirmed to contribute to the development of multidrug resistance including ERG11, CDR1, CDR2, MDR1, RTA2, Among them, upregulation of CDR1, a member of the ATP-binding cassette (ABC) transporter super-family in C. albicans, is a common occurrence in clinical C. albicans isolates resistant to antifungal azoles.13) Cdr1p pumps azoles out of C. albicans cells to reduce intracellular accumulation, which can be indirectly identified by the efflux of rhodamine 6G (R6G), which is a fluorescent substrate transported by Cdr1p. 14) In the promoter of CDR1, the basal expression element (BEE) (−960-−710) is responsible for basal transcription of CDR1, and the drugresponsive element (DRE) (−264-−244) is responsible for drug-induced transcription.15) The DRE (5′-CGG ATA TCG GAT ATT TTT TTT-3′) contains CGG triplets that are often recognized by Zinc finger proteins. 16,17) It has been demonstrated that the DRE of CDR1 can be recognized and bound by the transcription controller Tac1p that is characterized by a highly conserved Zn(II) 2 Cys 6 motif within the DNA binding domain (DBD). 15,16) Small molecules which can regulate the expression of CDR1 become a promising strategy to combat azole resistance in C. albicans. Berberine (BBR) is a plant-derived isoquinoline alkaloid from the roots and bark o...

“…[1][2][3] Coincident with the increased use of antifungal drugs, the incidences of drug resistance have also increased. [4][5][6] In recent years, the development of multidrug resistance in clinical isolates has challenged effective treatment of the infections.…”

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confidence: 99%

Fcr1p Inhibits Development of Fluconazole Resistance in Candida albicans by Abolishing CDR1 Induction

Shen

Wang

et al. 2007

Candida albicans (C. albicans), the major opportunistic fungal pathogen in humans, causes diseases varying from superficial mucosal complaints to life-threatening systemic disorders. [1][2][3] Coincident with the increased use of antifungal drugs, the incidences of drug resistance have also increased. [4][5][6] In recent years, the development of multidrug resistance in clinical isolates has challenged effective treatment of the infections. Specifically, the extensive and repetitive use of antifungal azole derivatives such as fluconazole (FLC) has allowed C. albicans to utilize many mechanisms of resistance in order to ensure its survival. 7,8) This situation highlights the need for elucidating the mechanism of drug resistance in C. albicans to develop new antifungal agents.Overexpression of efflux pumps, either ATP-binding cassette (ABC) or major facilitator superfamily (MFS) transporters, has been shown to be one of the major mechanisms of drug resistance in clinical isolates because if causes active extrusion of the drug out of the cell. 9-11) The Candida drug resistance 1 (CDR1) gene, which encodes an ABC efflux pump, is identified by complementation of the pleiotropic drug resistance 5 (pdr5) mutant, which is hypersensitive to cycloheximide, chloramphenicol, and azole drugs, in Saccharomyces cerevisiae (S. cerevisiae). 12) C. albicans cdr1 mutant resulted in increasing susceptibilities to azole drugs, 13) which is consistent with the observation that overexpression of CDR1 contributes to the drug resistance of clinical isolates of C. albicans. 6,14) Furthermore, the existence of trans-regulatory factors of CDR1 has also been suggested. 15) However, the molecular mechanism and the gene network regulating the expression of CDR1 and drug resistance are poorly understood.C. albicans Fcr1p (for fluconazole resistance 1 protein), a member of the family of zinc cluster proteins, is characterized by a highly conserved Zn(II) 2 Cys 6 zinc finger motif within the N-terminal DNA binding domain (DBD). 16) Well known Pdr1p and Pdr3p (pleiotropic drug resistance protein) zinc cluster proteins regulate the expression of several multidrug ABC transporter genes including PDR5, SNQ2, and YOR1, causing azole resistance in S. cerevisiae. 17-28) C. albicans Fcr1p displays significant sequence homology with S. cerevisiae Pdr1p and Pdr3p, and it could confer azole resistance in S. cerevisiae pdr1 pdr3 mutant by regulating PDR5. 16) We hypothesized that Fcr1p maybe involved in the development of C. albicans azole resistance as well as Pdr1p and Pdr3p in S. cerevisiae.In the present study we investigated the role of Fcr1p in the development of C. albicans azole resistance and possible mechanisms in vitro and in vivo. The results showed that Fcr1p inhibited development of fluconazole resistance in C. albicans through abolishing the induction of CDR1 expression by FLC, and in contrast FLC resistance development was accelerated resulting from the deletion of FCR1. (Table 1). MATERIALS AND METHODS Strains and MediaThe strains were cul...