The rising number of invasive fungal infections caused by drug-resistant Candida strains is one of the greatest challenges for the development of novel antifungal strategies. The scarcity of available antifungals has drawn attention to the potential of natural products as antifungals and in combinational therapies. One of these is catechins-polyphenolic compounds-flavanols, found in a variety of plants. In this work, we evaluated the changes in the susceptibility of Candida glabrata strain characterized at the laboratory level and clinical isolates using the combination of catechin and antifungal azoles. Catechin alone had no antifungal activity within the concentration range tested. Its use in combination with miconazole resulted in complete inhibition of growth in the sensitive C. glabrata isolate and a significant growth reduction in the azole resistant C. glabrata clinical isolate. Simultaneous use of catechin and miconazole leads to increased intracellular ROS generation. The enhanced susceptibility of C. glabrata clinical isolates to miconazole by catechin was accompanied with the intracellular accumulation of ROS and changes in the plasma membrane permeability, as measured using fluorescence anisotropy, affecting the function of plasma membrane proteins.
Závažný medicínsky problém v súčasnosti predstavujú mykotické infekcie. Život ohrozujúce stavy, ktoré sú asociované najmä s imunokompromitovanými pacientami, spôsobujú vo veľkej miere zástupcovia z rodu Candida. Najčastejším pôvodcom je kvasinka Candida albicans, v posledných rokoch však dochádza k výraznému posunu k druhu Candida glabrata a ďalším tzv. non-albicans Candida kvasinkám (napr. Candida tropicalis, Candida parapsilosis). S vysokou mortalitou sú asociované invazívne infekcie spôsobované multirezistentnou kvasinkou Candida auris. Medzi kvasinkou C. glabrata a ostatnými pôvodcami kandidóz existuje niekoľko rozdielov v biologických vlastnostiach, ako aj vo faktoroch prispievajúcich k virulencii. Vrodene zvýšená rezistencia voči azolom je, spolu so schopnosťou rýchlo nadobúdať rezistenciu aj voči ostatným skupinám antifungálnych látok, nebezpečnou kombináciou pri zvládaní úspešnej liečby kandidových infekcií. Poznanie faktorov podieľajúcich sa na virulencii a mechanizmov vedúcich k rezistencii patogéna voči antifungálnym látkam môže viesť k lepšiemu zvládaniu kandidových infekcií. Nemenej dôležitým je hľadanie nových cieľových miest pre antifungálnu terapiu. Predložená práca stručne sumarizuje existujúce poznatky, ktoré sa venujú uvedenej problematike.
Fungal infections are currently a serious health concern. Life-threatening conditions that occur mainly in immunocompromised patients are largely caused by representatives of the genus Candida. The most common causative agent is the yeast Candida albicans, but in recent years there has been a significant shift towards Candida glabrata and other so-called non-albicans Candida yeasts (e.g. Candida tropicalis or Candida parapsilosis). Invasive infections caused by the multidrug-resistant yeast Candida auris are associated with high mortality. There are several differences between C. glabrata and other causative agents of candidiasis in biological characteristics and virulence factors. The innate increased resistance to azoles along with the ability to rapidly acquire resistance to other groups of antifungal agents is a dangerous combination which makes it difficult to manage Candida infections. A better understanding of the virulence factors and mechanisms of resistance to antifungal agents can benefit the management of Candida infections. Equally important is the search for new target sites for antifungal therapy. The present work briefly summarizes the existing knowledge in this area.
- MeSH
- antifungální látky farmakologie terapeutické užití MeSH
- biofilmy MeSH
- Candida glabrata * MeSH
- Candida MeSH
- faktory virulence MeSH
- fungální léková rezistence * MeSH
- kandidóza * farmakoterapie mikrobiologie MeSH
- lidé MeSH
- mikrobiální testy citlivosti MeSH
- virulence MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- přehledy MeSH
KlUpc2p, a transcription factor belonging to the fungal binuclear cluster family, is an important regulator of ergosterol biosynthesis and azole drug resistance in Kluyveromyces lactis. In this work, we show that the absence of KlUpc2p generates Rag- phenotype and modulates the K. lactis susceptibility to oxidants and calcofuor white. The KlUPC2 deletion leads to increased expression of KlMGA2 gene, encoding an important regulator of hypoxic and lipid biosynthetic genes in K. lactis and also KlHOG1 gene. The absence of KlUpc2p does not lead to statistically significant changes in glycerol, corroborating the expression of KlGPD1 gene, encoding NAD+-dependent glycerol-3-phosphate dehydrogenase, that is similar in both the deletion mutant and the parental wild-type strain. Increased sensitivity of Klupc2 mutant cells to brefeldin A accompanied with significant increase in KlARF2 gene expression point to the involvement of KlUpc2p in intracellular signaling. Our observations highlight the connections between ergosterol and fatty acid metabolism to modulate membrane properties and point to the possible involvement of KlUpc2p in K. lactis oxidative stress response.
In yeast, the STB5 gene encodes a transcriptional factor belonging to binuclear cluster class (Zn2Cys6) of transcriptional regulators specific to ascomycetes. In this study, we prepared the Kluyveromyces lactis stb5Δ strain and assessed its responses to different stresses. We showed that KlSTB5 gene is able to complement the deficiencies of Saccharomyces cerevisiae stb5Δ mutant. The results of phenotypic analysis suggested that KlSTB5 gene deletion did not sensitize K. lactis cells to oxidative stress inducing compounds but led to Klstb5Δ resistance to 4-nitroquinoline-N-oxide and hygromycin B. Expression analysis indicated that the loss of KlSTB5 gene function induced the transcription of drug efflux pump encoding genes that might contribute to increased 4-nitroquinoline-N-oxide and hygromycin B tolerance. Our results show that KlStb5p functions as negative regulator of some ABC transporter genes in K. lactis.
- MeSH
- 4-nitrochinolin-1-oxid farmakologie MeSH
- delece genu MeSH
- fungální proteiny genetika metabolismus MeSH
- Kluyveromyces účinky léků genetika metabolismus MeSH
- oxidační stres účinky léků MeSH
- regulace genové exprese u hub účinky léků MeSH
- Saccharomyces cerevisiae - proteiny genetika metabolismus MeSH
- Saccharomyces cerevisiae účinky léků genetika metabolismus MeSH
- transkripční faktory genetika metabolismus MeSH
- Publikační typ
- časopisecké články MeSH
We investigated the effect of Kluyveromyces lactis ERG6 gene deletion on plasma membrane function and showed increased susceptibility of mutant cells to salt stress, cationic drugs and weak organic acids. Contrary to Saccharomyces cerevisiae, Klerg6 mutant cells exhibited increased tolerance to tunicamycin. The content of cell wall polysacharides did not significantly vary between wild-type and mutant cells. Although the expression of the NAD+-dependent glycerol 3-phosphate dehydrogenase (KlGPD1) in the Klerg6 mutant cells was only half of that in the parental strain, it was induced in the presence of calcofluor white. Also, cells exposed to this drug accumulated glycerol. The absence of KlErg6p led to plasma membrane hyperpolarization but had no statistically significant influence on the plasma membrane fluidity. We propose that the phenotype of Klerg6 mutant cells to a large extent was a result of the reduced activity of specific plasma membrane proteins that require proper lipid composition for full activity.
- MeSH
- delece genu MeSH
- fungální proteiny genetika metabolismus MeSH
- fyziologická adaptace * MeSH
- kationické antimikrobiální peptidy metabolismus MeSH
- Kluyveromyces účinky léků enzymologie genetika fyziologie MeSH
- kyseliny karboxylové toxicita MeSH
- methyltransferasy genetika metabolismus MeSH
- osmotický tlak MeSH
- regulace genové exprese u hub * MeSH
- tolerance léku MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Multidrug transporters are often responsible for failure of medical treatment, since they expel a variety of structurally and functionally unrelated drugs out of the cell. We found that the fluorescent probe diS-C3(3) is a substrate of not only Pdr5p of Saccharomyces cerevisiae (ScPdr5p) but also of its less-explored Kluyveromyces lactis homologue (KlPdr5p). This enabled us to compare the ability of azoles to competitively inhibit the Pdr5p-mediated probe efflux in the two species. In K. lactis, these azoles completely inhibit probe transport by KlPdr5p and also compete with each other for transport. This indicates that the probe and the azoles are bound by the same site(s) of the KlPdr5p binding pocket. On the other hand, the azoles' capacity to inhibit the probe transport by ScPdr5p is limited, as a result of their partial cotransport with the probe. While the azoles bind to only one or two separate binding sites, the probe is able to bind to all three of them. Moreover, the bulky ScPdr5p substrate enniatin B, which effectively inhibits both probe and azole transport by the pump, has negligible effect on KlPdr5p. Our data point to a tighter arrangement of the KlPdr5p binding pocket compared to that of ScPdr5p.
- MeSH
- ABC transportéry chemie genetika metabolismus MeSH
- azoly chemie farmakologie MeSH
- biologický transport MeSH
- fluorescenční barviva MeSH
- fluorescenční protilátková technika MeSH
- Kluyveromyces účinky léků metabolismus MeSH
- kompetitivní vazba MeSH
- Saccharomyces cerevisiae - proteiny chemie metabolismus MeSH
- Saccharomyces cerevisiae účinky léků metabolismus MeSH
- substrátová specifita MeSH
- vazba proteinů MeSH
- vazebná místa * MeSH
- Publikační typ
- časopisecké články MeSH
Boron is an essential micronutrient for living cells, yet its excess causes toxicity. To date, the mechanisms of boron toxicity are poorly understood. Recently, the ScATR1 gene has been identified encoding the main boron efflux pump in Saccharomyces cerevisiae. In this study, we analyzed the ScATR1 ortholog in Kluyveromyces lactis--the KNQ1 gene, to understand whether it participates in boron stress tolerance. We found that the KNQ1 gene, encoding a permease belonging to the major facilitator superfamily, is required for K. lactis boron tolerance. Deletion of the KNQ1 gene led to boron sensitivity and its overexpression increased K. lactis boron tolerance. The KNQ1 expression was induced by boron and the intracellular boron concentration was controlled by Knq1p. The KNQ1 promoter contains two putative binding motifs for the AP-1-like transcription factor KlYap1p playing a central role in oxidative stress defense. Our results indicate that the induction of the KNQ1 expression requires the presence of KlYap1p and that Knq1p like its ortholog ScAtr1p in S. cerevisiae functions as a boron efflux pump providing boron resistance in K. lactis.
- MeSH
- bor metabolismus MeSH
- fungální proteiny genetika metabolismus MeSH
- homeostáza MeSH
- Kluyveromyces enzymologie genetika metabolismus MeSH
- membránové transportní proteiny genetika metabolismus MeSH
- promotorové oblasti (genetika) MeSH
- regulace genové exprese u hub MeSH
- Saccharomyces cerevisiae genetika metabolismus MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- MeSH
- antifungální látky * MeSH
- buněčná membrána * metabolismus MeSH
- ergosterol MeSH
- fosfatidylethanolaminy MeSH
- fosfolipidy metabolismus MeSH
- imunitní systém * MeSH
- kardiolipiny metabolismus MeSH
- kvasinky * klasifikace MeSH
- lidé MeSH
- metabolismus lipidů * MeSH
- Saccharomyces cerevisiae MeSH
- sfingolipidy metabolismus MeSH
- steroly metabolismus MeSH
- xenobiotika * MeSH
- Check Tag
- lidé MeSH
