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DNA Methylation regulates phenotype-dependent transcriptional activity in

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Abstract

DNA methylation is a common epigenetic signaling mechanism associated with silencing of repeated DNA and transcriptional regulation in eukaryotes. Here we report that DNA methylation in the human fungal pathogen Candida albicans is primarily localized within structural genes and modulates transcriptional activity. Major repeat sequences and multigene families are largely free of DNA methylation. Among the genes subject to DNA methylation are those associated with dimorphic transition between yeast and hyphal forms, switching between white and opaque cells, and iron metabolism. Transcriptionally repressed methylated loci showed increased frequency of C-to-T transitions during asexual growth, an evolutionarily stable pattern of repression associated mutation that could bring about genetic alterations under changing environmental or host conditions. Dynamic differential DNA methylation of structural genes may be one factor contributing to morphological plasticity that is cued by nutrition and host interaction.

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... Differently, most of TEs in each subclass were methylated ( Figure 6D). For example, 83.8-87.9% of Gypsy members A c c e p t e d M a n u s c r i p t 13 have been found methylated in CpG context with average methylation level of 50.1-56.7%. The unexpressed elements of "Others" LTR retrotransposons subclass had the average methylation level of 87.4-97.1%. ...
... With the transcriptomic profiles determined, we then switched to profiling DNA cytosine methylation, for which a growing body of evidence have demonstrated its dynamic patterns and diverse functionalities in fungi [7,8,9,10,11,12,13]. A range of global methylation levels of 3.3% to 5.2% was observed in the four conditions of H. parviporum with a pronounced preference for CpG dinucleotides (6.7-9.3%) over non-CpG nucleotide contexts (2.0-3.7%), which is a phenomenon also shared by the majority of other species having methylation pattern studied [6,7,12]. ...
... Recently, the presence of cytosine methylation in gene bodies has been increasingly reported in many eukaryotic organisms including fungi [13,48]. In H. parviporum, the occurrences of A c c e p t e d M a n u s c r i p t 22 methylation in genes, markedly increased methylation level in gene flanking regions, together with varied methylation level in different conditions, particularly in NECT and SAP conditions ( Figure 2C,D) made us speculate that DNA methylation may, to a certain extent, play a role in condition- specific gene expression. ...
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Heterobasidion parviporum is the most devastating fungal pathogen of conifer forests in Northern Europe. The fungus has dual life strategies, necrotrophy on living trees and saprotrophy on dead woods. DNA cytosine methylation is an important epigenetic modification in eukaryotic organisms. Our presumption is that the lifestyle transition and asexual development in H. parviporum could be driven by epigenetic effects. Involvements of DNA methylation in the regulation of aforementioned processes have never been studied thus far. RNA-seq identified lists of highly induced genes enriched in carbohydrate-active enzymes during necrotrophic interaction with host trees and saprotrophic sawdust growth. It also highlighted signaling- and transcription factor-related genes potentially associated with the transition of saprotrophic to necrotrophic lifestyle and groups of primary cellular activities throughout asexual development. Whole-genome bisulfite sequencing revealed that DNA methylation displayed pronounced preference in CpG dinucleotide context across the genome and mostly targeted transposable element (TE)-rich regions. TE methylation level demonstrated a strong negative correlation with TE expression, reinforcing the protective function of DNA methylation in fungal genome stability. Small groups of genes putatively subject to methylation transcriptional regulation in response to saprotrophic and necrotrophic growth in comparison with free-living mycelia were also explored. Our study reported on the first methylome map of a forest pathogen. Analysis of transcriptome and methylome variations associated with asexual development and different lifestyle strategies provided further understanding of basic biological processes in H. parviporum. More importantly, our work raised additional potential roles of DNA methylation in fungi apart from controlling the proliferation of TEs. © 2019, © 2019 The Author(s). Published by Informa UK Limited, trading as Taylor & Francis Group.
... DNA methylation is a common, but not universal, epigenetic modification that plays an important role in multiple cellular processes, including genome structure rearrangements [1][2][3], development [2,4,5], or the response to environmental factors [6,7]. DNA methylation relies on covalent addition of methyl groups to cytosine (5 mC), and less frequently-to adenosine (6 mA) or to uracil (hydroxyl-mU; the latter is specific for dinoflagellates) [8]. ...
... However, genome methylation in different fungal species is still a matter of debate. The occurrence of this epigenetic modification has been confirmed by different technical approaches in several yeast species, including Komagataella phaffii [13], 2 of 16 Candida albicans [1], and Saccharomyces cerevisiae [14,15]. On the other hand, the vast majority of reports deny the presence of genomic DNA methylation in S. cerevisiae [8,16] and K. phaffii [13], and many other yeast species. ...
... Prior to the 5 mC (%) calculation, the amount of 5 mC (ng) was quantified using the slope (OD450 nm/ng) of the standard curve. The global DNA methylation was calculated using the following formula: 5 mC% = 5 mC amount (ng) amount of input sample DNA (ng) (1) All results were normalized to the readout for negative control (DNA devoid of any methylation) provided by the kit's manufacturer, by subtraction of the control sample value from the readout for the unknown samples. The results were expressed as the mean values ± SD of the three independent biological replicates, all analyzed in technical duplicate. ...
Article
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DNA methylation is a common, but not universal, epigenetic modification that plays an important role in multiple cellular processes. While definitely settled for numerous plant, mammalian, and bacterial species, the genome methylation in different fungal species, including widely studied and industrially-relevant yeast species, Yarrowia lipolytica, is still a matter of debate. In this paper, we report a differential DNA methylation level in the genome of Y. lipolytica subjected to sequential subculturing and to heat stress conditions. To this end, we adopted repeated batch bioreactor cultivations of Y. lipolytica subjected to thermal stress in specific time intervals. To analyze the variation in DNA methylation between stressed and control cultures, we (a) quantified the global DNA methylation status using an immuno-assay, and (b) studied DNA methylation patterns through whole-genome sequencing. Primarily, we demonstrated that 5 mC modification can be detected using a commercial immuno-assay, and that the modifications are present in Y. lipolytica’s genome at ~0.5% 5 mC frequency. On the other hand, we did not observe any changes in the epigenetic response of Y. lipolytica to heat shock (HS) treatment. Interestingly, we identified a general phenomenon of decreased 5 mC level in Y. lipolytica’s genome in the stationary phase of growth, when compared to a late-exponential epigenome. While this study provides an insight into the subculturing stress response and adaptation to the stress at epigenetic level by Y. lipolytica, it also leaves an open question of inability to detect any genomic DNA methylation level (either in CpG context or context-less) through whole-genome sequencing. The results of ONT sequencing, suggesting that 5 mC modification is either rare or non-existent in Y. lipolytica genome, are contradicted with the results of the immunoassay.
... Epigenetic factors, such as DNA methylation, can regulate the cellular response in absence of sequence changes in the DNA, and these factors can be modified and reversed by internal and external cellular stimuli, such as pollutants, oxidative stress, temperature, and nutrients (Byun and Baccarelli, 2014). Currently the only data available on C. albicans epigenetic regulation focus on its nuclear genome, mainly on modulation of morphology and other virulence factors, such as white-opaque switching (Zordan et al., 2006;Mishra et al., 2011;Zhang et al., 2013;Kim et al., 2015;Tscherner et al., 2015;Freire-Benéitez et al., 2016). Mapping of C. albicans nuclear genome hypermethylated sites identified genes involved in morphogenesis and hyphal growth (16.7%), ...
... white-opaque switching (3.3%), iron use (6.7%), drugs resistance and signaling (12%), stress response (7.3%), and genes involved in regulatory activities such as chromatin organization (3.3%), cycle or cell division (7.3%), biogenesis and protein transport (12.7%), DNA/RNA processing (5.3%), pathogenesis or virulence (2%), and carbohydrate metabolism (1.3%). It was also observed that, in this species, methylation occurs at both CpG and CH sites (H = Adenine, Cytosine, or Thymine), mainly in the gene bodies, instead of the promoters (Mishra et al., 2011). ...
... Also, for some genes, within a predominantly methylated population, the occurrence of unmethylated copies between them is common. This indicates that methylases may have a limiting rate, which results in incomplete methylation or that the transition from active to inactive transcription occurs through a passive dilution of methylated copies during replication (Mishra et al., 2011). ...
Article
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The commensal yeast Candida albicans is an opportunistic pathogen. In order to successfully colonize or infect the human body, the fungus must adapt to the host’s environmental conditions, such as low oxygen tension (hypoxia), temperature (37°C) and the different carbon sources available. Previous studies demonstrated the adaptive importance of C. albicans genetic variability for its pathogenicity, although the contributions of epigenetic and the influence of environmental factors are not fully understood. Mitochondria play important roles in fungal energetic metabolism, regulation of nuclear epigenetic mechanisms and pathogenicity. However, the specific impact of inter-strain mitochondrial genome variability and mitochondrial epigenetics in pathogenicity is unclear. Here we draw attention to this relevant organelle and its potential role in C. albicans pathogenicity and provide preliminary evidence, for the first time, for methylation of the yeast mitochondrial genome. Our results indicate that environmental conditions, such as continuous exposure for 12 weeks to hypoxia and 37°C, decrease the mitochondrial genome methylation in strains SC5314 and L757. However, the methylation decrease is quantitatively different in specific genome positions when strains SC5314 and L757 are compared. We hypothesize that this phenomenon can be promising for future research to understand how physical factors of the host affect the C. albicans mitochondrial genome and its possible impact on adaptation and pathogenicity.
... albicans epigenetic plasticity as an adaptive mechanism (Freire-Benéitez et al., 2016;Lopes da Rosa and Kaufman, 2012;Tscherner et al., 2015;Zordan et al., 2006). To date, the few studies on C. albicans epigenetic regulation focus on the epigenetics of its nuclear genome, mainly on the modulation of its morphology and other virulence factors, such as white-opaque switching (Freire-Benéitez et al., 2016;Kim et al., 2015;Mishra et al., 2011;Tscherner et al., 2015;Zhang et al., 2013;Zordan et al., 2006). There is increasing evidence of mtDNA methylation role in several diseases and its potential use as a biomarker for harmful environmental and nutritional factors (Iacobazzi et al., 2013). ...
... It was also observed that, in this species, methylation occurs at both CpG and CH sites (H = Adenine, Cytosine or Thymine), and mainly in the gene bodies, instead of the promoters. Mapping of hypermethylated sites in C. albicans genome revealed that the majority were within genes (82%), followed by non-repetitive intergenic regions (13%) and repetitive regions (5%), suggesting that methylation in this yeast genome plays an important role in gene regulation (Mishra et al., 2011). However, to date, there are no studies on C. albicans mitochondrial genome methylation. ...
... Also, for some genes, within a predominantly methylated population, the occurrence of unmethylated copies between them is common. This indicates that methylases may have a limiting rate, which results in incomplete methylation or that the transition from active to inactive transcription occurs through a passive dilution of methylated copies during replication (Mishra et al., 2011). ...
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Infection by Candida albicans requires its adaption to physical constraints in the human body, such as low oxygen tension (hypoxia), increased temperature (37°C) and different carbon sources. Previous studies demonstrated that the genetic variability of C. albicans isolates is an important adaptive mechanism, although little is known about the dynamics of this genetic diversity, and the influence of these environmental conditions on its mitochondrial genome (mtDNA). To test the synergistic effect of these stress conditions on C. albicans genome, reference strain SC5314 was subjected to an in vitro evolution scheme under hypoxia and 37°C, with two different carbon sources (glycerol and dextrose) for up to 48 weeks (approximately 4,000 generations). Experimental evolution results showed no sequence or copy number changes in the mtDNA, although sequence variants were detected on its nuclear genome by Multilocus sequence typing (MLST) and whole genome sequencing (WGS). After 12 weeks of experimental evolution, sample GTH12, grown under hypoxia at 37°C in glycerol, showed inferior growth and respiratory rates as compared to other conditions tested. Although WGS of GTH12 revealed no variants in its mtDNA, WGS with sodium bisulfite showed a significant reduction in mtDNA methylation in GTH12 in both non-coding and coding regions. Our results provide the first whole mitochondrial genome methylation map of C. albicans and show that environmental conditions promote the selective growth of specific variants and affect the methylation patterns of the mtDNA in a strain-specific manner.
... For instance, DNA methylation in the model fungus Neurospora crassa is restricted to transposable elements that have been targeted by a genome defence system (Repeat Induced Point mutations; RIP) (Selker et al. 2003;Lewis et al. 2009). Nevertheless, gene methylation has also been observed in few fungi Mishra, Baum and Carbon 2011;Jeon et al. 2015). In the human pathogen, Candida albicans, gene methylation modulates the transcriptional activity of genes associated with morphological plasticity and iron metabolism, which is important for signalling and pathogenicity (Mishra, Baum and Carbon 2011). ...
... Nevertheless, gene methylation has also been observed in few fungi Mishra, Baum and Carbon 2011;Jeon et al. 2015). In the human pathogen, Candida albicans, gene methylation modulates the transcriptional activity of genes associated with morphological plasticity and iron metabolism, which is important for signalling and pathogenicity (Mishra, Baum and Carbon 2011). Similarly, exons in the saprophytic fungus Uncinocarpus reesii can be methylated in certain genomic contexts, and this methylation is correlated with transcription . ...
Article
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Fungal secondary metabolites are small molecules that exhibit diverse biological activities exploited in medicine, industry and agriculture. Their biosynthesis is governed by co-expressed genes that often co-localize in gene clusters. Most of these secondary metabolite gene clusters are inactive under laboratory conditions, which is due to a tight transcriptional regulation. Modifications of chromatin, the complex of DNA and histone proteins influencing DNA accessibility, play an important role in this regulation. However, tinkering with well-characterised chemical and genetic modifications that affect chromatin alters the expression of only few biosynthetic gene clusters, and thus the regulation of the vast majority of biosynthetic pathways remains enigmatic. In the past, attempts to activate silent gene clusters in fungi mainly focused on histone acetylation and methylation, while in other eukaryotes many other post-translational modifications are involved in transcription regulation. Thus, how chromatin regulates the expression of gene clusters remains a largely unexplored research field. In this review, we argue that focusing on only few well-characterised chromatin modifications is significantly hampering our understanding of the chromatin-based regulation of biosynthetic gene clusters. Research on underexplored chromatin modifications and on the interplay between different modifications is timely to fully explore the largely untapped reservoir of fungal secondary metabolites.
... Incomplete capabilities for histone methylation are widespread among hemiascomycetes. For example, the best-understood eukaryotic model organism, S. cerevisiae, lacks H3K9, H3K27, and cytosine DNA methylation; another widely used model, S. pombe, lacks H3K27 and cytosine DNA methylation; and one model for human opportunistic pathogens, Candida albicans, lacks H3K9 and H3K27 methylation but may have cytosine DNA methylation (95). Within the filamentous fungi several lineages have lost the genes required for H3K27 methylation (e.g., Mucor, Rhizopus, Aspergillus), and at least some strains of Zymoseptoria tritici lack the ability for cytosine DNA methylation (28). ...
... Zymoseptoria tritici has no cytosine methylation (28), but sister species have intact genes for DNMTs. No obvious DNMTs are found in the Candida genome, yet there have been reports on cytosine DNA methylation (95). Cytosine DNA methylation has now been found in Cryptococcus neoformans, catalyzed by the novel DNMT5, for which there are genes in genera that had been thought to be devoid of cytosine methylation (Aspergillus, Penicillium, Histoplasma, Coccidioides). ...
Article
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Histone-modifying enzymes are responsible for regulating transcription, recombination, DNA repair, DNA replication, chromatid cohesion, and chromosome segregation. Fungi are ideally suited for comparative chromatin biology because sequencing of numerous genomes from many clades is coupled to existing rich methodology that allows truly holistic approaches, integrating evolutionary biology with mechanistic molecular biology and ecology, promising applications in medicine or plant pathology. While genome information is rich, mechanistic studies on histone modifications are largely restricted to two yeasts, Saccharomyces cerevisiae and Schizosaccharomyces pombe, and one filamentous fungus, Neurospora crassa-three species that arguably are not representative of this diverse kingdom. Here, histone methylation serves as a paradigm to illustrate the roles chromatin modifications may play in more complex fungal life cycles. This review summarizes recent advances in our understanding of histone H3 methylation at two sites associated with active transcription, lysine 4 and lysine 36 (H3K4, H3K36); a site associated with the formation of constitutive heterochromatin, lysine 9 (H3K9); and a site associated with the formation of facultative heterochromatin, lysine 27 (H3K27). Special attention is paid to differences in how methylation marks interact in different taxa. Expected final online publication date for the Annual Review of Microbiology Volume 71 is September 8, 2017. Please see http://www.annualreviews.org/page/journal/pubdates for revised estimates.
... Currently, DNA methylation has been studied in a few yeasts species. Some studies postulated the absence of this epigenetic modification while some others demonstrated the existence of a variable degree of DNA methylation [30,[39][40][41][42]. Despite these investigations, so far, no studies have been ...
... albida), as a possible response to cold stress. DNA methylation has recently been studied in Candida albicans; Metschnikowia reukaufii; Cryptococcus laurentii (now Papiliotrema laurentii); and some species within the Kluyveromyces, Candida, Schizosaccharomyces, and Saccharomyces genera [39,40,42,52]. Despite the involvement of DNA methylation in the rapid response to biotic and abiotic stresses has been widely investigated in plants [46,[53][54][55][56][57], none of the above-mentioned studies have taken into consideration the possible involvement of DNA methylation as a possible response induced in yeast species by cold stress. ...
Article
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The involvement of DNA methylation in the response to cold stress of two different yeast species (Naganishia antarctica, psychrophilic, and Naganishia albida, psychrotolerant), exhibiting different temperature aptitudes, has been studied. Consecutive incubations at respective optimum temperatures, at 4 °C (cold stress) and at optimum temperatures again, were performed. After Methylation Sensitive Amplified Polymorphism (MSAP) fingerprints a total of 550 and 423 clear and reproducible fragments were amplified from N. antarctica and N. albida strains, respectively. The two Naganishia strains showed a different response in terms of level of DNA methylation during cold stress and recovery from cold stress. The percentage of total methylated fragments in psychrophilic N. antarctica did not show any significant change. On the contrary, the methylation of psychrotolerant N. albida exhibited a nonsignificant increase during the incubation at 4 °C and continued during the recovery step, showing a significant difference if compared with control condition, resembling an uncontrolled response to cold stress. A total of 12 polymorphic fragments were selected, cloned, and sequenced. Four fragments were associated to genes encoding for elongation factor G and for chitin synthase export chaperon. To the best of our knowledge, this is the first study on DNA methylation in the response to cold stress carried out by comparing a psychrophilic and a psychrotolerant yeast species.
... Elevated gene body (intron/exon) methylation in PN conditions may be important to maintain homeostasis, while decreased gene body methylation may allow for more rapid changes to gene expression under NPN conditions (Figure 3). Such gene body methylation is still poorly understood in fungi, however, a study in Candida albicans suggested that DNA methylation is predominantly located in the gene body and modulates transcriptional activity by acting as a repressive mark (Mishra et al., 2011). A majority of methylated genes in C. albicans encoded proteins involved in environmentally cued pathways, suggesting gene body methylation plays an important role in the regulation of transcription under changing environmental conditions (Mishra et al., 2011) Similarly, in Ganoderma sinense, transcriptional repression of BGC was linked to DNA methylation in their gene bodies (Zhu et al., 2015). ...
... Such gene body methylation is still poorly understood in fungi, however, a study in Candida albicans suggested that DNA methylation is predominantly located in the gene body and modulates transcriptional activity by acting as a repressive mark (Mishra et al., 2011). A majority of methylated genes in C. albicans encoded proteins involved in environmentally cued pathways, suggesting gene body methylation plays an important role in the regulation of transcription under changing environmental conditions (Mishra et al., 2011) Similarly, in Ganoderma sinense, transcriptional repression of BGC was linked to DNA methylation in their gene bodies (Zhu et al., 2015). The methylation density in G. sinense was similar to F. graminearum at 1.8%. ...
Article
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Histone modifications play a significant role in the regulation of biosynthetic gene clusters (BGCs) in the phytopathogen Fusarium graminearum, by contrast, epigenetic regulation by DNA methyltransferases (DNMTs) is less documented. In this study, we characterized two DNMTs (FgDIM-2 and FgRID) in F. graminearum, with homologies to “Deficient in methylation” (DIM-2) and “Repeat-induced point (RIP) deficient” (RID) from Neurospora. The loss of DNMTs resulted in not only a decrease in average methylation density in the nutrient-poor, compared to nutrient-rich conditions, but also differences in the genes expressed between the WT and the DNMT mutant strains, implicating the external environment as an important trigger in altering DNA methylation patterns. Consequently, we observed significant changes in the regulation of multiple BGCs and alterations in the pathogenicity of the fungus.
... Comparison of DNA methylomes across more than 20 eukaryote genomes (including those of animals, plants and fungi) has revealed that most of the genomes undergo DNA methylation, while only a few eukaryotes show zero or little evidence of DNA methylation, i.e., yeast (Saccharomyces cerevisiae), fruit fly (Drosophila melanogaster) and roundworm (Caenorhabditis elegans) (Jeltsch, 2010). DNA methylation was also detected in some model fungal species (Filippovich et al., 2004;Mishra et al., 2011;Montanini et al., 2014). For example, DNA methylation in Neurospora crassa (red bread mold; Ascomycota) is crucial for balancing the formation of sexual and asexual reproductive structures (Filippovich et al., 2004). ...
... For example, DNA methylation in Neurospora crassa (red bread mold; Ascomycota) is crucial for balancing the formation of sexual and asexual reproductive structures (Filippovich et al., 2004). In the human fungal pathogen Candida albicans, DNA methylation has been recently reported to be primarily located within structural genes and associated with the dimorphic transition between yeast and hyphal forms, switching between white and opaque cells, and iron metabolism (Mishra et al., 2011). In black truffle, Tuber melanosporum, the DNA methylation pattern revealed selective targeting of transposable elements (TEs) rather than gene bodies, and demethylation treatment changed its phenotype (Montanini et al., 2014). ...
Article
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DNA methyltransferases (DNMTs) are a group of proteins that catalyze DNA methylation by transferring a methyl group to DNA. The genetic variation in DNMTs results in differential DNA methylation patterns associated with various biological processes. In fungal species, DNMTs and their DNA methylation profiles were found to be very diverse and have gained many research interests. We reviewed fungal DNMTs in terms of their biological functions, protein domain structures, and their associated epigenetic regulations compared to those known in plant and animal systems. In addition, we summarized recent reports on potential RNA-directed DNA methylation (RdDM) related to DNMT5 in fungi. We surveyed up to 40 fungal species with published genome-wide DNA methylation profiles (methylomes) and presented the associations between the specific patterns of fungal DNA methylation and their DNMTs based on a phylogenetic tree of protein domain structures. For example, the main DNMTs in Basidiomycota, DNMT1 with RFD domain + DNMT5, contributing to CG methylation preference, were distinct from RID + Dim-2 in Ascomycota, resulting in a non-CG methylation preference. Lastly, we revealed that the dynamic methylation involved in fungal life stage changes was particularly low in mycelium and DNA methylation was preferentially located in transposable elements (TEs). This review comprehensively discussed fungal DNMTs and methylomes and their connection with fungal development and taxonomy to present the diverse usages of DNA methylation in fungal genomes.
... methylation of genome in Tuber melanosporum [53]. More recent research showed that in fungi, the DNA methylation is around the genes [49][50][51][52], which have more complex function than we have observed in the past; they are also present in more complicated and divergent patterns, for example, they show their role in defense of genome like in bacteria [49,51,53], and also, research showed that they have dynamic epigenetic entity, play their role in the development of M.oryzea [49,50], and play role in the secondary metabolism and regulation of G. sinense [51] and cell morphology delay in Candida Albicans [54] by changing the transcriptional activities of related genes. DNA methylation is related to different lifestyles for example Decomposition, parasitism and life cycles of fungi, in H, parvipoerum 96,026 it is related to asexual (mycelia and conidia) lifestyle which was confirmed by Zhen Zeng et al in H, parvipoerum that DNA methylation have a role in the transcriptional regulation of asexual patterns, and N.carsaa before going to sexual cycle the spore fuses and before nuclear fusion take place a defense system is activated which works to prevent the repetitive sequencing such as TEs [55]. ...
... Methylation of histone is widespread among the hemiasco mycetes, best model organism for the understanding is S. cerevisiae don't have H3K9, H3K27 and cytosine methylation, another model organism S. pombe lack H3K27 and methylation of histone but the specie Candida albicans don't have H3K9 and H3K27 but they might have cytosine DNA methylation [54]. ...
Article
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DNA methylation is a process in which methyl (CH3) groups are added to the DNA molecule. The DNA segment does not change in the sequence, but DNA methylation could alter the action of DNA. Different enzymes like DNA methyltransferases (DNMTs) take part in methylation of cytosine/adenine nucleosides in DNA. In prokaryotes, DNA methylation is performed to prevent the attack of phage and also plays a role in the chromosome replication and repair. In fungi, DNA methylation is studied to see the transcriptional changes, as in insects, the DNA methylation is not that well-known, it plays a different role like other organisms. In mammals, the DNA methylation is related to different types of cancers and plays the most important role in the placental development and abnormal DNA methylation connected with diseases like cancer, autoimmune diseases, and rheumatoid arthritis.
... It is estimated that the level of DNA methylation of Candida albicans is only 0.5%. It is believed that DNA methylation plays in fungi a major role in silencing of repeated sequences (Mishra et al., 2011). ...
... The authors believe that dynamic changes in the methylation profile may explain the morphological plasticity of Candida albicans cells. Although the applied method of assessing the level of Candida albicans genome methylation has not revealed its role in the development of resistance to azoles, it showed similar level of methylation, suggested by other authors (Mishra et al., 2011). ...
Article
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A phenomenon of increasing resistance of Candida spp. to azoles has been observed for several years now. One of the mechanisms of lack of sensitivity to azoles is associated with CDR1, CDR2, MRD1 genes (their products are active transport pumps conditioning drug efflux from pathogen's cell), and ERG11 gene (encoding lanosterol 14α-demethylase). Test material was 120 strains of Candida albicans (60 resistant and 60 susceptible to azole drugs) obtained from clinical samples. The first stage of experiment assessed the expression of CDR1, CDR2, MDR1 and ERG11 genes by Q-PCR. The impact of ERG11 gene's mutations on the expression of this gene was analysed. The final stage of the experiment assessed the level of genome methylation of Candida albicans strains. An increase in the expression of CDR2, MDR1 and ERG11 was observed in azole-resistant strains of Candida albicans in comparison to strains sensitive to this class of drugs. Furthermore, 19 changes in the sequence of ERG11 were detected in tested strains. Four of the discovered mutations: T495A, A530C, G622A and A945C led to the following amino acid substitutions: D116E, K128T, V159I and E266D, respectively. It has also been found that statistically five mutations: T462C, G1309A, C216T, C1257T and A945C affected the expression of ERG11. The applied method of assessing the level of methylation of Candida albicans genome did not confirm its role in the development of resistance to azoles. The results indicate however, that resistance of Candida albicans strains to azole drugs is multifactorial.
... The histone acetyl transferase Hat1 was found to mediate oxidative stress tolerance and resistance to azoles in C. albicans [61]. Moreover, DNA methylation in C. albicans was linked to morphological switching and iron metabolism, both processes important for virulence [62]. Therefore, epigenetic modulations add another level of plasticity, which may result in strain-specific differences. ...
Article
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The incidence of human infections caused by the opportunistic fungal pathogen Candida albicans is on the rise due to increasing numbers of immunosuppressed patients. The importance of the immune system in preventing overgrowth of the colonizing fungus and thereby limiting infection is well recognized and host protective mechanisms widely investigated. Only recently, it was recognized that the natural diversity in the fungal species could also influence the outcome of the interaction between the fungus and the host. C. albicans strain-specific differences are complex and their regulation at the genomic, genetic, and epigenetic level and by environmental factors is only partially understood. In this review, we provide an overview of the natural diversity of C. albicans and discuss how it impacts host-fungal interactions and thereby affects the balance between commensalism versus disease.
... In Neurospora, DNA methylation is mainly regarded as a genome defense mechanism to silence transposable elements and DNA repeats (Martienssen and Colot, 2001;Selker et al., 2003). In Candida albicans, DNA methylation takes place predominantly in structural genes and regulates transcriptional activity, with repeat regions largely devoid of methylation (Mishra et al., 2011). In Magnaporthe oryzae, DNA methylation serves as a dynamic epigenetic modification functioning in genome defense and fungal development (Jeon et al., 2015) and DNA methylation was shown to be a dynamic process during sexual development in Cordyceps militaris ( Wang et al., 2015). ...
Article
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In eukaryotic genomes, DNA methylation is an important type of epigenetic modification that plays crucial roles in many biological processes. To investigate the impact of a hypovirus infection on the methylome of Cryphonectria parasitica, the chestnut blight fungus, whole-genome bisulfite sequencing (WGBS) was employed to generate single-base resolution methylomes of the fungus with/without hypovirus infection. The results showed that hypovirus infection alters methylation in all three contexts (CG, CHG, and CHH), especially in gene promoters. A total of 600 differentially methylated regions (DMRs) were identified, of which 144 could be annotated to functional genes. RNA-seq analysis revealed that DNA methylation in promoter is negatively correlated with gene expression. Among DMRs, four genes were shown to be involved in conidiation, orange pigment production, and virulence. Taken together, our DNA methylomes analysis provide valuable insights into the understanding of the relationship between DNA methylation and hypovirus infection, as well as phenotypic traits in C. parasitica.
... Moreover, DNA methylation is not present in a variety of unicellular yeasts such as Saccharomyces cerevisiae and Schizosaccharomyces pombe, and also in some filamentous ascomycetes evolutionarily related to Neurospora such as Aspergillus nidulans and Aspergillus flavus [16,17]. Another unusual situation occurs in the dimorphic yeast Candida albicans, where methylation has been shown to target and modulate the transcription of genes [18]. ...
Article
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Background We investigated how an extremely transposon element (TE)-rich organism such as the plant-symbiotic ascomycete truffle Tuber melanosporum exploits DNA methylation to cope with the more than 45,000 repeated elements that populate its genome.ResultsWhole-genome bisulfite sequencing performed on different developmental stages reveals a high fraction of methylated cytosines with a strong preference for CpG sites. The methylation pattern is highly similar among samples and selectively targets TEs rather than genes. A marked trend toward hypomethylation is observed for TEs located within a 1 kb distance from expressed genes, rather than segregated in TE-rich regions of the genome. Approximately 300 hypomethylated or unmethylated TEs are transcriptionally active, with higher expression levels in free-living mycelium compared to fruitbody. Indeed, multiple TE-enriched, copy number variant regions bearing a significant fraction of hypomethylated and expressed TEs are found almost exclusively in free-living mycelium. A reduction of DNA methylation, restricted to non-CpG sites and accompanied by an increase in TE expression, is observed upon treatment of free-living mycelia with 5-azacytidine.Conclusions Evidence derived from analysis of the T. melanosporum methylome indicates that a non-exhaustive, partly reversible, methylation process operates in truffles. This allows for the existence of hypomethylated, transcriptionally active TEs that are associated with copy number variant regions of the genome. Non-exhaustive TE methylation may reflect a role of active TEs in promoting genome plasticity and the ability to adapt to sudden environmental changes.
... However, this report [8] could not completely rule out the existence of DNA methylation in C. elegans, because the techniques of HPLC-UV used have a limit of detection (LOD) of 0.01 % 5-mdC, which leaves room for about 3500 molecules of 5-mdC being undetected in C. elegans [9]. A low level of methylation may still be important in gene regulation, because several studies have shown that a few methylated residues in DNA are sufficient to modulate genetic switches [10,11]. ...
Article
Since 1986 till now, the popular research model organism Caenorhabditis elegans has been thought to completely lack DNA methylation and seems to lost DNA methylation enzymes from its genomes. Here we report the development of a sensitive and selective assay based on LC-MS/MS to simultaneously measure 5-methyl-2'-deoxycytidine (5-mdC) and 5-hydroxymethyl-2'-deoxycytidine (5-hmdC) in DNA hydrolysates. With the use of isotope internal standards (d3-5-mdC and d3-5-hmdC) and online solid-phase extraction, the detection limits of 5-mdC and 5-hmdC were estimated to be 0.01 and 0.02 pg, respectively, which correspond to 0.000006% and 0.00001% methylation and hydroxymethylation level. This method was applied to investigate whether the DNA methylation/hydroxymethylation exists in C. elegans. Our work for the first time demonstrates that 5-mdC is present in C. elegans genomic DNA (0.0019-0.0033% of cytosine methylated) using LC-MS/MS, while another epigenetic modification 5-hmdC is not detectable. Furthermore, we found that C. elegans DNA was hypomethylated or hypermethylated in a dose-dependent manner by the DNA methyltransferase (DNMT)-inhibiting drug decitabine (5-aza-2'-deoxycytidine) or cadmium, respectively. Our data support the possible existence of active DNA methylation mechanism in C. elegans, in which unidentified DNMTs could be involved. Our work highlights the importance that the evolutionary conservation of DNA methylation machinery in the nematodes that were traditionally considered to lack functional DNA methylation, needs to be re-evaluated.
... Epigenetic modifications serve as a connection between genetic components and environmental changes (Flanagan et al., 2006;Bock and Lengauer, 2008). Although not all fungi have a significant level of DNA methylation in their genomes (Liu et al., 2012), genome-wide DNA methylation has been profiled in several fungi using bisulfite sequencing (Mishra et al., 2011;Jeon et al., 2015). The present study contributed to our understanding of the DNA methylation status of C. parasitica. ...
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Mutation in CpBck1, an ortholog of the cell wall integrity mitogen-activated protein kinase kinase kinase (MAPKKK) of Saccharomyces cerevisiaee, in the chestnut blight fungus Cryphonectria parasitica resulted in a sporadic sectorization as culture proceeded. The progeny from the sectored area maintained the characteristics of the sector, showing a massive morphogenetic change, including robust mycelial growth without differentiation. Epigenetic changes were investigated as the genetic mechanism underlying this sectorization. Quantification of DNA methylation and whole-genome bisulfite sequencing revealed genome-wide DNA methylation of the wild-type at each nucleotide level and changes in DNA methylation of the sectored progeny. Compared to the wild-type, the sectored progeny exhibited marked genome-wide DNA hypomethylation but increased methylation sites. Expression analysis of two DNA methyltransferases, including two representative types of DNA methyltransferase (DNMTase), demonstrated that both were significantly down-regulated in the sectored progeny. However, functional analysis using mutant phenotypes of corresponding DNMTases demonstrated that a mutant of CpDmt1, an ortholog of RID of Neurospora crassa, resulted in the sectored phenotype but the CpDmt2 mutant did not, suggesting that the genetic basis of fungal sectorization is more complex. The present study revealed that a mutation in a signaling pathway component resulted in sectorization accompanied with changes in genome-wide DNA methylation, which suggests that this signal transduction pathway is important for epigenetic control of sectorization via regulation of genes involved in DNA methylation.
... The diploid C. albicans WO-1 genome is composed of 2.1% retroelements and contains only two copies each of a Tc1/mariner-like DNA transposon and a (defective) Mutator-like element (Butler et al. 2009). However, endogenous transposition activity has not been reported for any of these elements, despite a report that C. albicans transposons are not modified by DNA methylation ( Mishra et al. 2011). In this study, we explored the suitability of the maize Ac/Ds transposons for in vivo trans- poson insertion mutagenesis in C. albicans. ...
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In non-model systems genetic research is often limited by the lack of techniques for the generation and identification of gene mutations. One approach to overcome this bottleneck is the application of transposons for gene tagging. We have established a two-element transposon tagging system, based on the transposable elements Activator (Ac)/Dissociation (Ds) from maize, for in vivo insertion mutagenesis in the fungal human pathogen Candidaalbicans A non-autonomous Ds transposon carrying a selectable marker was constructed into the ADE2 promoter on chromosome 3 and a codon usage-adapted Ac transposase gene was inserted into the neutral NEUT5L locus on chromosome 5. In C. albicans cells expressing the transposase the Ds element efficiently excised and reintegrated elsewhere in the genome, which makes the Ac/Ds transposons promising tools for saturating insertion mutagenesis in clinical strains of C. albicans.
... Histone and other chromatin modifications as well as small RNAs are known to control the switch between the replicative and non-replicative stages of pathogens such as Toxoplasma gondii (Bougdour et al., 2009). Recent evidence indicates that DNA methylation causes Candida albicans to switch from the yeast to the hyphal form (Mishra et al., 2011). Bacterial pathogens lack chromatin but they have evolved a range of effector molecules and virulence factors that allow them to overcome host defenses, e.g. by reprograming the host epigenome to suppress anti-inflammatory responses (Mujtaba et al., 2013). ...
Article
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Epigenetic inheritance refers to changes in gene expression that are heritable across generations but are not caused by changes in the DNA sequence. Many environmental factors are now known to cause epigenetic changes, including the presence of pathogens, parasites, harmful chemicals and other stress factors. There is increasing evidence that transcriptional reprograming caused by epigenetic modifications can be passed from parents to offspring. Indeed, diseases such as cancer can occur in the offspring due to epigenetically-inherited gene expression profiles induced by stress experienced by the parent. Empirical studies to investigate the role of epigenetics in trans-generational gene regulation and disease require appropriate model organisms. In this review, we argue that selected insects can be used as models for human diseases with an epigenetic component because the underlying molecular mechanisms (DNA methylation, histone acetylation and the expression of microRNAs) are evolutionarily conserved. Insects offer a number of advantages over mammalian models including ethical acceptability, short generation times and the potential to investigate complex interacting parameters such as fecundity, longevity, gender ratio, and resistance to pathogens, parasites and environmental stress. Copyright © 2015. Published by Elsevier Ltd.
... Variation in genome methylation patterns suggests that the role of DNA methylation is not strictly conserved among different fungal species. DNA methylation inhibits elongation, but not initiation, of transcription in Neurospora crassa and regulates phenotype-dependent transcriptional activity in Candida albicans (Rountree & Selker 1997;Mishra et al. 2011). In Magnaporthe oryzae, DNA methylation plays a role in fungal development and genome defence throughout development of the asexual life cycle (Jeon et al. 2015). ...
Article
DNA methylation is a basic epigenetic mechanism found in eukaryotes, but its patterns and roles vary significantly among diverse taxa. In fungi, DNA methylation has various effects on diverse biological processes. However, its function in the sexual development of fungi remains unclear. Cordyceps militaris, readily performs sexual reproduction and thus provides a remarkably rich model for understanding epigenetic processes in sexual development. Here, we surveyed the methylome of C. militaris at single-base resolution to assess DNA methylation patterns during sexual development using genomic bisulfite sequencing (BS-Seq). The results showed that approximately 0.4 % of cytosines are methylated, similar to the DNA methylation level (0.39 %) during asexual development. Importantly, we found that DNA methylation in the fungi undergoes global reprogramming during fungal development. Moreover, RNA-Seq analysis indicated that the differentially methylated regions (DMRs) have no correlation with the genes that have roles during fungal sexual development in C. militaris. These results provide a comprehensive characterization of DNA methylation in the sexual development of C. militaris, which will contribute to future investigations of epigenetics in fungi.
... Coverage of the A. fumigatus genome was improved and fairly even across all 8 chromosomes supporting the idea that little of its genome is methylated at CCGG sites (S2 Fig). In contrast, the dimorphic yeast C. albicans uses cytosine methylation to modulate the transition between yeast and hyphal forms among other transcription events [28]. The presence of CpG methylation in C. albicans correlates with the lower genome enrichment of 5.5 fold relative to that of A. fumigatus (Fig 3A). ...
Article
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We describe continuing work to develop restriction endonucleases as tools to enrich targeted genomes of interest from diverse populations. Two approaches were developed in parallel to segregate genomic DNA based on cytosine methylation. First, the methyl-sensitive endonuclease HpaII was used to bind non-CG methylated DNA. Second, a truncated fragment of McrB was used to bind CpG methylated DNA. Enrichment levels of microbial genomes can exceed 100-fold with HpaII allowing improved genomic detection and coverage of otherwise trace microbial genomes from sputum. Additionally, we observe interesting enrichment results that correlate with the methylation states not only of bacteria, but of fungi, viruses, a protist and plants. The methods presented here offer promise for testing biological samples for pathogens and global analysis of population methylomes.
... It should be noted, however, that in the genomes of some RIP-competent species, including P. anserina and L. maculans , a clear association between RIP and methylation has not been reported (Graia et al. 2001 ;Idnurm and Howlett 2003 ;Arnaise et al. 2008 ). It is also worth noting that 5mC methylation in fungi can result in increased levels of C → T transitions (Mishra et al. 2011 ), via the deamination of 5mC (Nabel et al. 2012 ), which are not attributable to RIP. The determining factor for the targeted de novo methylation of "RIP-relics" appears to be the depletion of G:C content that results from RIP. ...
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Repeat-induced point mutation (RIP) is a form of genome mutation that is targeted towards repeated DNA sequences and which is observed only in certain fungal taxa; the Pezizomycotina (filamentous Ascomycota) and some species of the Basidiomycota. RIP is widely believed to have evolved to protect fungal genomes against transposon replication. RIP was first observed during the sexual reproductive cycle of the ascomycete Neurospora crassa. During the haploid dikaryotic stage that occurs following fertilization and prior to meiosis, the RIP process selectively mutated duplicated sequences in both DNA strands by inducing single-nucleotide point (SNP) mutations that converted C:G base pairs to T:A. This often led to the introduction of nonsense or missense mutations which affected the expression of these sequences. The precise mechanism by which detection and subsequent mutation of duplicated regions occurs is still unknown, but is dependent on a cytosine methyltransferase. RIP has also been observed to influence the evolution of fungal genes. With the exception of ribosomal DNA repeats, RIP acts upon repeated genomic regions including tandem repeats, unlinked repeats, and large segmental duplications including endogenous genes. RIP has the potential to either enhance or impede the generation of genetic diversity. Species exhibiting high levels of RIP are observed to be deficient in gene family diversity, whereas diversity among gene families in species with low levels of RIP is increased. Furthermore, RIP has also been reported to affect non-duplicated genes adjacent to RIP-affected repetitive DNA sequences, which may drive the evolution of these genes and can promote rapid adaptation to selection pressures in some species.
... DNA methylation is speculated to occur transiently during the sexual stage , DNA methylation may be not exist in Aspergillus flavus. DNA methylation prevents extension, transcription in Neurospora crassa and Candida albicans (Rountree and Selker, 1997;Mishra et al., 2011). In Magnaporthe oryzae, methylcytosines content (0.4%) and level (0.39%) of DNA methylation are similar during asexual development (Jeon et al., 2015). ...
Article
The methylation sensitive amplified polymorphism (MSAP) was used to induce DNA methylation transformation in mushroom mycelia. DNA obtained from the mycelial stages of P. eryngii subsp. tuoliensis was digested with isoschizomers Msp I or Hpa II (mixture of EcoR I), the ability to digest the sequence CpCpGpG as influenced by their methylation state. The data analysis demonstrated that full-methylated and unmethylation modifications were primary and the hemi-methylated ratio was significantly lower. These results indicated that the pattern of CG hypermethylation is abundant in P. eryngii subsp. tuoliensis. All fragments that were differentially amplified upon low temperature induction illustrated the feasibility of MSAP in edible mushrooms. Moreover, this study confirmed that genetic and epigenetic changes in P. eryngii subsp. tuoliensis were induced under low temperature.
... The variation in genome methylation levels suggests that DNA methylation is not strictly conserved among different fungal species. In N. crassa and Candida albicans, DNA methylation silences transposable elements and repetitive DNA sequences as a genome defense mechanism (Mishra et al. 2011;Selker et al. 2003). However, DNA methylation plays a role in fungal development throughout the asexual life cycle of M. oryzae (Jeon et al. 2015). ...
Article
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DNA methylation is an important epigenetic mark in mammals, plants, and fungi and depends on multiple genetic pathways involving de novo and maintenance DNA methyltransferases (DNMTases). Metarhizium robertsii, a model system for investigating insect-fungus interactions, has been used as an environmentally friendly alternative to chemical insecticides. However, little is known concerning the molecular basis for DNA methylation. Here, we report on the roles of two DNMTases (MrRID and MrDIM-2) by characterizing ΔMrRID, ΔMrDIM-2, and ΔRID/ΔDIM-2 mutants. The results showed that approximately 71, 10, and 8% of (m)C sites remained in the ΔMrRID, ΔMrDIM-2, and ΔRID/ΔDIM-2 strains, respectively, compared with the wild-type (WT) strain. Further analysis showed that MrRID regulates the specificity of DNA methylation and MrDIM-2 is responsible for most DNA methylation, implying an interaction or cooperation between MrRID and MrDIM-2 for DNA methylation. Moreover, the ΔMrDIM-2 and ΔRID/ΔDIM-2 strains showed more defects in radial growth and conidial production compared to the WT. Under ultraviolet (UV) irradiation or heat stress, an obvious reduction in spore viability was observed for all the mutant strains compared to the WT. The spore median lethal times (LT50s) for the ΔMrDIM-2 and ΔRID/ΔDIM-2 strains in the greater wax moth, Galleria mellonella, were decreased by 47.7 and 65.9%, respectively, which showed that MrDIM-2 is required for full fungal virulence. Our data advances the understanding of the function of DNMTase in entomopathogenic fungi, which should contribute to future epigenetic investigations in fungi.
... Table 1 includes documented regulatory associations, where, in the case of iron in C. albicans, it is based on the work of 2011), as well as potential regulation based on the presence of putative DNA-binding sites in the promoter region of genes. In addition to the aforementioned transcription factors, transcription of iron acquisition genes such as RBT5 is also inhibited by methylation during iron-replete conditions (Mishra et al., 2011). Interestingly, these authors also observed an increased frequency of mutation in genes that are repressed under iron-replete conditions. ...
Article
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Iron is an absolute requirement for both the host and most pathogens alike and is needed for normal cellular growth. The acquisition of iron by biological systems is regulated to circumvent toxicity of iron overload, as well as the growth deficits imposed by iron deficiency. In addition, hosts, such as humans, need to limit the availability of iron to pathogens. However, opportunistic pathogens such as Candida albicans are able to adapt to extremes of iron availability, such as the iron replete environment of the gastrointestinal tract and iron deficiency during systemic infection. C. albicans has developed a complex and effective regulatory circuit for iron acquisition and storage to circumvent iron limitation within the human host. As C. albicans can form complex interactions with both commensal and pathogenic co-inhabitants, it can be speculated that iron may play an important role in these interactions. In this review, we highlight host iron regulation as well as regulation of iron homeostasis in C. albicans. In addition, the review argues for the need for further research into the role of iron in polymicrobial interactions. Lastly, the role of iron in treatment of C. albicans infection is discussed.
... In Toxoplasma gondii infection, histone acetylation was considered to be responsible for the switch between replicative and non-replicative stages of the pathogen (Dixon et al. 2010). DNA methylation in the human fungal pathogen Candida albicans was shown to be related to the transition between its yeast and hyphal forms (Mishra, Baum and Carbon 2011). On the other hand, host epigenomic state can control microbial virulence by some intracellular pathogens (Gomez-Diaz et al. 2012). ...
Article
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Epigenetic mechanisms have rapidly and controversially emerged as silent modulators of host defenses that can lead to a more prominent immune response and shape the course of inflammation in host. Thus, the epigenetics can both drive the production of specific inflammatory mediators and control the magnitude of host response. The epigenetic actions that are predominantly shown to modulate the host defense against microbial pathogens are DNA methylation, histone modification, and the activity of non-coding RNAs. There is also growing evidence that opportunistic chronic pathogens, such as Porphyromonas gingivalis, as a microbial host subversion strategy, can epigenetically interfere with the host DNA machinery for successful colonization. Similarly, the novel involvement of small molecule “danger signals”, which are released by stressed or infected cells, in the center of host-pathogen interplay and epigenetics is developing. In this review, we systematically examined the latest knowledge within the field of epigenetics in the context of host-derived danger molecule and purinergic signaling, with a particular focus on host microbial defenses and infection-driven chronic inflammation.
... Other forms of epigenetic modification represent interesting fields of research; some, such as DNA methylation, have been investigated in pathogenic fungi (Liu et al., 2012;Mishra et al., 2011). Of note is the recently reported ability of some pathogenic bacteria to modify a host's gene expression through histone modifications and DNA methylation. ...
Article
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Opportunistic fungal infections are an increasing threat for global health, and for immunocompromised patients in particular. These infections are characterized by interaction between fungal pathogen and host cells. The exact mechanisms and the attendant variability in host and fungal pathogen interaction remain to be fully elucidated. The field of systems biology aims to characterize a biological system, and utilize this knowledge to predict the system's response to stimuli such as fungal exposures. A multi-omics approach, for example, combining data from genomics, proteomics, metabolomics, would allow a more comprehensive and pan-optic "two systems" biology of both the host and the fungal pathogen. In this review and literature analysis, we present highly specialized and nascent methods for analysis of multiple -omes of biological systems, in addition to emerging single-molecule visualization techniques that may assist in determining biological relevance of multi-omics data. We provide an overview of computational methods for modeling of gene regulatory networks, including some that have been applied towards the study of an interacting host and pathogen. In sum, comprehensive characterizations of host-fungal pathogen systems are now possible, and utilization of these cutting-edge multi-omics strategies may yield advances in better understanding of both host biology and fungal pathogens at a systems scale.
... Furthermore, the phytopathogenic fungus Cryphonectria parasitica shows epigenetic diversity among different haplotypes with distinctive expansion and invasion traits (68). Interestingly enough, genes mediating morphogenetic changes are enriched with heterogeneous DNA methylation patterns in C. albicans (69). Hence, hereditary epigenetic modifications, known as "epigenetic memory," provide a swift adaptive-fitness advantage in response to environmental cues or host immune defenses and enable the phenotypic plasticity of clonal (sub)populations. ...
Article
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The surge in antimicrobial drug resistance in some bacterial and fungal pathogens constitutes a significant challenge to health care facilities. The emerging human fungal pathogen Candida auris has been particularly concerning, as isolates can display pan-antifungal resistance traits against all drugs, including echinocandins.
... In addition, the sequence of C. albicans SC5314 is transcribed by previous studies so that C. albicans SC5314 is frequently used as a wild-type control derived from common laboratory [4]. Although previous studies did not indicate why C. albicans would be separated into different strains, strains SC5314 and WO1 are estimated to be separated from each other by a divergence time of one million years [5]. Both strains of C. albicans might exist in human body experiencing constant evolution to adapt for host microenvironment. ...
Article
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Candida albicans (C. albicans) is the most prevalent fungal species. Although it is a healthy microbiota, genetic and epigenetic alterations in host and pathogen, and microenvironment changes would lead to thrush, vaginal yeast infection, and even hematogenously disseminated infection. Despite the fact that cytotoxicity is well-characterized, few studies discuss the genome-wide genetic and epigenetic molecular mechanisms between host and C. albicans. The aim of this study is to identify drug targets and design a multiple-molecule drug to prevent the infection from C. albicans. To investigate the common and specific pathogenic mechanisms in human oral epithelial OKF6/TERT-2 cells during the C. albicans infection in different strains, systems modeling and big databases mining were used to construct candidate host–pathogen genetic and epigenetic interspecies network (GEIN). System identification and system order detection are applied on two-sided next generation sequencing (NGS) data to build real host–pathogen cross-talk GEINs. Core host–pathogen cross-talk networks (HPCNs) are extracted by principal network projection (PNP) method. By comparing with core HPCNs in different strains of C. albicans, common pathogenic mechanisms were investigated and several drug targets were suggested as follows: orf19.5034 (YBP1) with the ability of anti-ROS; orf19.939 (NAM7), orf19.2087 (SAS2), orf19.1093 (FLO8) and orf19.1854 (HHF22) with high correlation to the hyphae growth and pathogen protein interaction; orf19.5585 (SAP5), orf19.5542 (SAP6) and orf19.4519 (SUV3) with the cause of biofilm formation. Eventually, five corresponding compounds—Tunicamycin, Terbinafine, Cerulenin, Tetracycline and Tetrandrine—with three known drugs could be considered as a potential multiple-molecule drug for therapeutic treatment of C. albicans.
... Genetic studies have been conducted on C. albicans, revealing potential new targets for antifungal therapy (Becker et al., 2010;da et al., 2010;Mishra, Baum, & Carbon, 2011;Wurtele et al., 2010). In particular, Wurtele et al. (2010) detected that preserving the expression of the enzyme HST3 (histone deacetylase that removes H3K56ac genomewide) reduced H3K56ac compared with cells with lower expression of this enzyme, resulting in cells with the hyphae phenotype. ...
Article
Novel treatments are needed to prevent candidiasis/candidemia infection due to the emergence of Candida species resistant to current antifungals. Considering the yeast‐to‐hyphae switch is a critical factor to Candida albicans virulence, phenols common in plant sources have been reported to demonstrating their ability to prevent dimorphism. Therefore, phenols present in many agricultural waste stress (ferulic (FA) and gallic (GA) acid) were initially screened in isolation for their yeast‐to‐hyphae inhibitory properties at times 3, 6, and 24 hr. Both FA and GA inhibited 50% of hyphae formation inhibitory concentration (IC50) but at a concentration of 8.0 ± 0.09 and 90.6 ± 1.05 mM, respectively, at 24 hr. However, the inhibitory effect of FA increased by 1.9–2.6 fold when combined with different GA concentrations. GA and FA values decreased even lower when sinapic acid (SA) was added as a third component. As evidenced by concave isobolograms and combination indexes less than 1, both GA:F A and GA:FA:SA combinations acted synergistically to inhibit 50% hyphae formation at 24 hr. Lastly, acetylation of histone H3 lysine 56 acetylation (H3K56) was higher in response to the triple phenolic cocktail (using the IC50 24 hr inhibitory concentration level) comparable with the nontreated samples, indicating that the phenols inhibited hyphal growth in part by targeting H3K56 acetylation.
... The digestion pattern in control and 5-azacytidine treated samples were indicative of methylation in CCG and CG motifs, generally found in the genomes of plant and fungal systems (Fulnecek and Kovarík, 2014). Earlier studies have also identified CG methylation of the fungal genome of U. reesii and extensive gene body methylation in both CpG and CpN sites in C. albicans (Mishra et al., 2011). Jeon et al. (2015) observed around 0.22 % of total cytosines to be methylated with average methylation at 5-methylcytosine sites between 20-30 % in Magnoporthe oryzae genome. ...
Article
Fungal endophytes, a major component of the plant host microbiome, are known to synthesize plant-derived metabolites in vitro. However, attenuation of metabolite production upon repeated sub-culturing is a major drawback towards utilizing them as an alternative for plant-derived metabolites. In this study, we isolated Diaporthe perseae, a fungal endophyte from Gloriosa superba tubers, which showed the production of colchicine in axenic cultures. Mass spectrometry, Nuclear Magnetic Resonance spectroscopy, and tubulin polymerization assays confirmed the compound to be colchicine. Repeated sub-culturing of the endophyte for 10 generations led to a reduction in the yield of the metabolite from 55.25 μg/g to 2.32 μg/g of mycelial dry weight. Treatment of attenuated cultures with DNA methylation inhibitor 5-azacytidine resulted in increased metabolite concentration (39.68 μg/g mycelial dry weight) in treated samples compared to control (2.61 μg/g mycelial dry weight) suggesting that 5-azacytidine can induce demethylation of the fungal genome to overcome the phenomenon of attenuation of metabolite synthesis. Reduced levels of global methylation were observed upon 5-azacytidine treatment in attenuated cultures (0.41 % of total cytosines methylated) as compared to untreated control (0.78 % of total cytosines methylated). The results provide a significant breakthrough in utilizing fungal endophytes as a veritable source of plant-derived metabolites from critically endangered plants.
... Indeed, others made a similar observation in single-cell fungi. While the environmental yeasts studied herein and previously lacked any modifications (Capuano et al., 2014), the most frequent commensal yeast pathogen Candida albicans contained as sole yeast species 5mdC (Mishra et al., 2011). This result is interesting, because it could mean that host-pathogen interactions could select for similar DNA modifications in the pathogen as in the host. ...
Article
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Interpreting the function and metabolism of enzymatic DNA modifications requires both position-specific and global quantities. Sequencing-based techniques that deliver the former have become broadly accessible, but analytical methods for the global quantification of DNA modifications have thus far been applied mostly to individual problems. We established a mass spectrometric method for the sensitive and accurate quantification of multiple enzymatic DNA modifications. Then, we isolated DNA from 124 archean, bacterial, fungal, plant, and mammalian species, and several tissues and created a resource of global DNA modification quantities. Our dataset provides insights into the general nature of enzymatic DNA modifications, reveals unique biological cases, and provides complementary quantitative information to normalize and assess the accuracy of sequencing-based detection of DNA modifications. We report that only three of the studied DNA modifications, methylcytosine (5mdC), methyladenine (N6mdA) and hydroxymethylcytosine (5hmdC), were detected above a picomolar detection limit across species, and dominated in higher eukaryotes (5mdC), in bacteria (N6mdA), or the vertebrate central nervous systems (5hmdC). All three modifications were detected simultaneously in only one of the tested species, Raphanus sativus. In contrast, these modifications were either absent or detected only at trace quantities, across all yeasts and insect genomes studied. Further, we reveal interesting biological cases. For instance, in Allium cepa, Helianthus annuus, or Andropogon gerardi, more than 35% of cytosines were methylated. Additionally, next to the mammlian CNS, 5hmdC was also detected in plants like Lepidium sativum and was found on 8% of cytosines in the Garra barreimiae brain samples. Thus, identifying unexpected levels of DNA modifications in several wild species, our resource underscores the need to address biological diversity for studying DNA modifications.
... Collectively, these findings are concord with the assertion of previous works that although multiple systems of gene regulation operate, epigenetic control plays an important role in regulating host/pathogen interactions [37,41,42]. Our study also adds to the growing body of evidence suggesting the importance of dynamic DNA methylation as a regulator of phenotypic plasticity of plant pathogens and interaction with the host [43]. ...
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Pathogenic fungi can lose virulence after protracted periods of culture, but little is known of the underlying mechanisms. Here, we present the first analysis of DNA methylation flux at a single-base resolution for the plant pathogen B. cinerea and identify differentially methylated genes/genomic regions associated with virulence erosion during in vitro culture. Cultures were maintained for eight months, with subcultures and virulence testing every month. Methylation-sensitive amplified polymorphisms were performed at monthly intervals to characterise global changes to the pathogen’s genome during culture and also on DNA from mycelium inoculated onto Arabidopsis thaliana after eight months in culture. Characterisation of culture-induced epialleles was assessed by whole-genome re-sequencing and whole-genome bisulfite sequencing. Virulence declined with time in culture and recovered after inoculation on A. thaliana. Variation detected by methylation-sensitive amplified polymorphisms followed virulence changes during culture. Whole-genome (bisulfite) sequencing showed marked changes in global and local methylation during culture but no significant genetic changes. We imply that virulence is a non-essential plastic character that is at least partly modified by the changing levels of DNA methylation during culture. We hypothesise that changing DNA methylation during culture may be responsible for the high virulence/low virulence transition in B. cinerea and speculate that this may offer fresh opportunities to control pathogen virulence.
... Similarly to S. cerevisiae, C. albicans is devoid of H3K9me and H3K27me [33]. Although 5mC mark has been detected in C. albicans, it is unclear whether DNA methylation is associated with heterochromatic regions in this organism [34]. Instead, chromatin profiling studies have demonstrated that C. albicans heterochromatic regions are characterised by low levels of both histone acetylation and methylation [31] The post-translation modification of histone proteins is a dynamic and reversible process catalysed by "writer" and "eraser" enzymes that add and remove epigenetic marks ( Figure 2). ...
Article
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The human fungal pathogen Candida albicans is a dimorphic opportunistic pathogen that colonises most of the human population without creating any harm. However, this fungus can also cause life-threatening infections in immunocompromised individuals. The ability to successfully colonise different host niches is critical for establishing infections and pathogenesis. C. albicans can live and divide in various morphological forms critical for its survival in the host. Indeed, C. albicans can grow as both yeast and hyphae and can form biofilms containing hyphae. The transcriptional regulatory network governing the switching between these different forms is complex but well understood. In contrast, non-DNA based epigenetic modulation is emerging as a crucial but still poorly studied regulatory mechanism of morphological transition. This review explores our current understanding of chromatin-mediated epigenetic regulation of the yeast to hyphae switch and biofilm formation. We highlight how modification of chromatin structure and non-coding RNAs contribute to these morphological transitions.
... DNA methylation in C. albicans is restricted to structural genes that modulate transcriptional activities, whereas repeat sequences and multigene families are comparatively free of DNA methylation; for instance, studies report methylation of INP51, MUC1, and LIP8 genes, which are related to pathogenicity and virulence (Mishra et al. 2011). The cell wall protein b-glucans induce functional reprogramming of monocytes by elevating H3K4me3 levels at the promoters of TNF-a, IL -6, and IL-18 through dectin -1/ Raf -1 pathway. ...
Article
The interaction of microbiota with its host has the ability to alter the cellular functions of both, through several mechanisms. Recent work, from many laboratories including our own, has shown that epigenetic mechanisms play an important role in the alteration of these cellular functions. Epigenetics broadly refers to change in the phenotype without a corresponding change in the DNA sequence. This change is usually brought by epigenetic modifications of the DNA itself, the histone proteins associated with the DNA in the chromatin, non-coding RNA or the modifications of the transcribed RNA. These modifications, also known as epigenetic code, do not change the DNA sequence but alter the expression level of specific genes. Microorganisms seem to have learned how to modify the host epigenetic code and modulate the host transcriptome in their favour. In this review, we explore the literature that describes the epigenetic interaction of bacteria, fungi and viruses, with their mammalian hosts.
... Silencing transposon appears to be the main purpose of DNA methylation in fungi. Unusual deposition patterns occur in the dimorphic yeast Candida albicans, where DNA methylation has been shown to target and modulate the transcription of genes [37]. ...
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Botrytis cinerea is one of the most destructive fungal pathogens that cause gray mold rot in horticultural products, including fresh fruits, vegetables, and flowers, leading to serious economic losses. B. cinerea is difficult to control because it has strong stress resistance and complex infection modes. The pathogenic mechanisms of B. cinerea have been revealed at multiple levels, but little is known at the epigenetic level. In this study, we first revealed the important role of DNA methyltransferases in regulating the development and pathogenicity of B. cinerea. We showed that two DNA methyltransferases, BcDIM2 and BcRID2, showed a strong synergistic effect in regulating the pathogenicity of B. cinerea. The double knockout mutant △Bcdim2rid2 showed slower mycelial growth, lower spore germination, attenuated oxidative tolerance, and complete pathogenicity loss on various hosts, which is related to the reduced expression of virulence-related genes in △Bcdim2rid2 and the induced resistance of the host. Although B. cinerea has multiple DNA methyltransferases, the global methylation level is very low, and few 5mC sites can be detected by BS-seq. These results first revealed the important role and the action mode of DNA methyltransferases in B. cinerea.
... Previous studies of CGIs of yeast by Sharif et al. [41] noticed that low CpG promoters are targeted by DNA methylation. Furthermore, Mishra et al. [42] in their study attempted that the methylated gene region in fungi are likely involved in pathogenesis or virulence. Consistent with the present analysis, previously, low G + C content was reported from fungi family Clavicipitaceae [43] and S. auriculariicola [40]. ...
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Background Pest control strategies almost entirely rely on chemical insecticides, which cause environmental problems such as biosphere deterioration and emergence of resistant pests. Bio-pesticide is an alternative approach, which uses organisms such as entomopathogenic fungi, Metarhizium anisopliae, to control pests. Screening such potential organism at a molecular level and understanding their gene regulation mechanism is an important approach to reduce emergence of pesticide resistance and worsening of the biosphere. Understanding promoter regions which play a pivotal role in gene regulation is crucial. In particular, identification of the promoter regions in M. anisopliae Strain ME1 remains poorly understood. To our knowledge, the mitogenome trn gene clusters of M. anisopliae Strain ME1 were not characterized. Here, we used machine learning approach to identify and characterize the promoter regions, regulatory elements, and CpG island densities of 15 protein coding genes of entomopathogenic fungi, M. anisolpliae Strain ME1. Results The current analysis revealed multiple transcription start sites (TSS) for all utilized sequences, except for promoter region genes of Pro-cob and Pro-nad5. With reference to the start codon (ATG), 85.3% of TSS was located above – 500 bp. Based on the standard predictive score at cut off value of 0.8a, the current study revealed 54.7% of predictive score greater than or equal from 0.9 promoter prediction score. Expectation maximization algorithm output identified five candidate motifs. Nonetheless, of all candidate motifs, MtrnI was revealed as the common promoter region motif with a value of 76.9% both in terms of size of binding sites and with an E value of 9.1E−054. Accordingly, we perceived that MtrnI serve as the binding site for tryptophan cluster with P value 0.0044 and C4 type zinc fingers functions as the binding site to regulate gene expression of M. anisopliae Strain ME1. The analysis revealed that mitogenome trn gene clusters of M. anisopliae Strain ME1 showed homologues evolutionary ancestor supported with a bootstrap value of 100%. Conclusion Identified common candidate motifs and binding transcription factors through in silico approach are likely expected to contribute for better understanding of gene expression and strain improvement of M. anisopliae Strain ME1 for its bio-pesticides role.
... Previous studies of CGIs of yeast by Sharif et al. [41] noticed that low CpG promoters are targeted by DNA methylation. Furthermore, Mishra et al. [42] in their study attempted that the methylated gene region in fungi are likely involved in pathogenesis or virulence. Consistent with the present analysis, previously, low G + C content was reported from fungi family Clavicipitaceae [43] and S. auriculariicola [40]. ...
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Background Pest control strategies almost entirely rely on chemical insecticides, which cause environmental problems such as biosphere deterioration and emergence of resistant pests. Bio-pesticide is an alternative approach, which uses organisms such as entomopathogenic fungi, Metarhizium anisopliae , to control pests. Screening such potential organism at a molecular level and understanding their gene regulation mechanism is an important approach to reduce emergence of pesticide resistance and worsening of the biosphere. Understanding promoter regions which play a pivotal role in gene regulation is crucial. In particular, identification of the promoter regions in M. anisopliae Strain ME1 remains poorly understood. To our knowledge, the mitogenome trn gene clusters of M. anisopliae Strain ME1 were not characterized. Here, we used machine learning approach to identify and characterize the promoter regions, regulatory elements, and CpG island densities of 15 protein coding genes of entomopathogenic fungi, M. anisolpliae Strain ME1. Results The current analysis revealed multiple transcription start sites (TSS) for all utilized sequences, except for promoter region genes of Pro-cob and Pro-nad5. With reference to the start codon (ATG), 85.3% of TSS was located above – 500 bp. Based on the standard predictive score at cut off value of 0.8 a , the current study revealed 54.7% of predictive score greater than or equal from 0.9 promoter prediction score. Expectation maximization algorithm output identified five candidate motifs. Nonetheless, of all candidate motifs, MtrnI was revealed as the common promoter region motif with a value of 76.9% both in terms of size of binding sites and with an E value of 9.1E−054. Accordingly, we perceived that MtrnI serve as the binding site for tryptophan cluster with P value 0.0044 and C4 type zinc fingers functions as the binding site to regulate gene expression of M. anisopliae Strain ME1. The analysis revealed that mitogenome trn gene clusters of M. anisopliae Strain ME1 showed homologues evolutionary ancestor supported with a bootstrap value of 100%. Conclusion Identified common candidate motifs and binding transcription factors through in silico approach are likely expected to contribute for better understanding of gene expression and strain improvement of M. anisopliae Strain ME1 for its bio-pesticides role.
... Methylation of gene bodies is found in plants [36,37], mammals [36,37], possibly fungi [42], and insects [7,36,37]. This broad phylogenetic distribution suggests that gene body methylation is one of the oldest conserved features of eukaryotic DNA methylation systems. ...
... 73 Sequence analysis for differentially regulated genes 74 Fastq files, obtained from the sequencing, were analyzed for qual-75 ity using FastQC v0.11.5; (Andrews 2010) and the reads with 76 low quality (<Q30) were discarded, using PRINSEQ-lite v0.20.4 77 (Schmieder and Edwards 2011). The sequences were aligned to 78 the C. albicans SC5314 genome assembly 21 (Skrzypek et al. 2017) 79 using TopHat2 (Trapnell et al. 2012;Kim et al. 2013) with the fr-80 secondstrand, -r 250 mate-std-I 10000 -G option (Dutton et al. (Trapnell et al. 2012) was also used to gen-87 erate differential expression data using aligned reads obtained 88 from TopHat2 (Trapnell et al. 2012;Kim et al. 2013) 105 were analyzed with the NanoString nCounter analysis system 106 (Geiss et al. 2008) at the University of the Witwatersrand, Depart-107 ment of Internal Medicine, using a gene expression TagSet that 108 targets 36 genes including three housekeeping genes (orf19.1191, 109 SLF1, orf19.2184). ...
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Candida albicans is an opportunistic yeast pathogen within the human microbiota with significant medical importance because of its pathogenic potential. The yeast produces highly resistant biofilms, which are crucial for maintaining infections. Though antifungals are available, their effectiveness is dwindling due to resistance. Alternate options that comprise the combination of existing azoles and polyunsaturated fatty acids, such as arachidonic acid (AA), have been shown to increase azoles susceptibility of C. albicans biofilms; however, the mechanisms are still unknown. Therefore, transcriptome analysis was conducted on biofilms exposed to sub-inhibitory concentrations of AA alone, fluconazole alone, and AA combined with fluconazole to understand the possible mechanism involved with the phenomenon. P rotein AN alysis TH rough E volutionary R elationships (PANTHER) analysis from the differentially expressed genes revealed that the combination of AA and fluconazole influence biological processes associated with essential processes including methionine synthesis processes and those involved in ATP generation, such as AMP biosynthesis, fumarate metabolism and fatty acid oxidation. These observation suggests that the interference of AA with these processes may be possible mechanisms to induce increased antifungal susceptibility
... It is unknown whether and how these γH2A-enriched regions are continuously repaired and whether DNA damage repair at these sites contributes to C. albicans genome instability. DNA that is methylated on cytosine (5mC) has been detected at the rDNA locus and subtelomeric regions, suggesting that DNA methylation is an important contributor to C. albicans heterochromatin structure [85]. It will be important to identify the DNA methyltransferase(s) that establish and maintain this modification as this will allow dissection of the role of DNA methylation in the regulation of gene expression and genome stability in C. albicans. ...
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Human fungal pathogens, such as Candida albicans, Aspergillus fumigatus and Cryptococcus neoformans, are a public health problem, causing millions of infections and killing almost half a million people annually. The ability of these pathogens to colonise almost every organ in the human body and cause life-threating infections relies on their capacity to adapt and thrive in diverse hostile host-niche environments. Stress-induced genome instability is a key adaptive strategy used by human fungal pathogens as it increases genetic diversity, thereby allowing selection of genotype(s) better adapted to a new environment. Heterochromatin represses gene expression and deleterious recombination and could play a key role in modulating genome stability in response to environmental changes. However, very little is known about heterochromatin structure and function in human fungal pathogens. In this review, I use our knowledge of heterochromatin structure and function in fungal model systems as a road map to review the role of heterochromatin in regulating genome plasticity in the most common human fungal pathogens: Candida albicans, Aspergillus fumigatus and Cryptococcus neoformans.
... A notable exception includes the fission yeast Schizosaccharomyces pombe genome that contains a Dnmt2-like homolog. To date, the presence of 5-mC in the ascomycetous yeast genomes remains controversial [20,21]. ...
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DNA methyltransferases are ubiquitous enzymes conserved in bacteria, plants and opisthokonta. These enzymes, which methylate cytosines, are involved in numerous biological processes, notably development. In mammals and higher plants, methylation patterns established and maintained by the cytosine DNA methyltransferases (DMTs) are essential to zygotic development. In fungi, some members of an extensively conserved fungal-specific DNA methyltransferase class are both mediators of the Repeat Induced Point mutation (RIP) genome defense system and key players of sexual reproduction. Yet, no DNA methyltransferase activity of these purified RID (RIP deficient) proteins could be detected in vitro. These observations led us to explore how RID-like DNA methyltransferase encoding genes would play a role during sexual development of fungi showing very little genomic DNA methylation, if any. To do so, we used the model ascomycete fungus Podospora anserina. We identified the PaRid gene, encoding a RID-like DNA methyltransferase and constructed knocked-out ΔPaRid defective mutants. Crosses involving P. anserina ΔPaRid mutants are sterile. Our results show that, although gametes are readily formed and fertilization occurs in a ΔPaRid background, sexual development is blocked just before the individualization of the dikaryotic cells leading to meiocytes. Complementation of ΔPaRid mutants with ectopic alleles of PaRid, including GFP-tagged, point-mutated and chimeric alleles, demonstrated that the catalytic motif of the putative PaRid methyltransferase is essential to ensure proper sexual development and that the expression of PaRid is spatially and temporally restricted. A transcriptomic analysis performed on mutant crosses revealed an overlap of the PaRid-controlled genetic network with the well-known mating-types gene developmental pathway common to an important group of fungi, the Pezizomycotina.
... This is estimated because iron with a concentration of 5% is probably too large to be induced to Candida albicans. According to Mishra et al. (9), the condition of excess iron in Candida can increase the methylation of DNA, which in turn can cause the activity of the iron receiving gene to be depressed. In addition, according to Lan et al. (5), the condition of iron overload can suppress the expression of the Sfu1 gene and inhibit Sef1 expression so that it also suppresses the iron receiving gene. ...
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Background: Oral candidiasis is one of the most common fungal infections, which attack the mucosa of the oral cavity. These lesions are mostly caused by the fungal species Candida albicans. Candida albicans is included in the normal oral microorganisms that are opportunistic pathogens, and its presence is quite large, which can reach 75% of the total oral fungal population. Research on specific proteins of Candida biofilm can be an alternative to early prevention of oral infections such as Oral Candidiasis. This biofilm protein can be used as a reference in making kits to detect the presence of microbes that cause infectious diseases. The purpose of this study was to determine molecular weight of Candida albicans biofilm protein induced by 5% glucose, 5% lactose, soy protein, and 5% iron. Material and methods: This experimental laboratory study used SDS-PAGE electrophoresis to determine the molecular weight of Candida albicans biofilm proteins induced by glucose 5%, lactose 5%, soy protein, and iron 5%. Results: Biofilm induced by 5% glucose shows four protein bands: 71,6 kDa; 56,1 kDa; 49,7 kDa; and 41 kDa. Biofilm induced by 5% lactose shows seven protein bands: 71 kDa; 61,2 kDa; 57,7 kDa; 55,3 kDa; 48,9 kDa; 39,5 kDa; and 29,8 kDa. Biofilm induced by soy protein shows one protein band: 49,4 kDa. Biofilm induced by 5% iron shows one protein band: 51,1 kDa. Conclusions: Candida albicans biofilm induced by 5% glucose has four protein band candidates, 5% lactose has seven candidates of protein band, and soy protein and 5% iron each has a candidate of protein band, which can be used as a target for the detection of oral Candidiasis. Key words:Biofilm protein, Candida albicans, molecular weight, oral candidiasis.
... Unlike S. pombe, the genome of C. albicans does not encode an HP1/Swi6-like protein, an H3K9 methyltransferase like Clr4 and components of a fully functional RNAi machinery (FREIRE-BENEITEZ et al. 2016). There is no evidence of DNA methylation at the CEN DNA in C. albicans (BAUM et al. 2006;MISHRA et al. 2011). The reversible silencing of the expression of a marker gene, URA3, captured by 5-Fluoroorotic acid (5-FOA) counter-selection, has been observed upon its integration into the CENPA binding region of the centromere in C. albicans endowing it a transcriptionally flexible status (THAKUR and SANYAL 2013). ...
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The diploid budding yeast Candida albicans harbors unique CENPA-rich 3- to 5-kb regions that form the centromere (CEN) core on each of its eight chromosomes. The epigenetic nature of these CENs does not permit the stabilization of a functional kinetochore on an exogenously introduced CEN plasmid. The flexible nature of such centromeric chromatin is exemplified by the reversible silencing of a transgene upon its integration into the CENPA-bound region. The lack of a conventional heterochromatin machinery and the absence of defined boundaries of CENPA chromatin makes the process of CEN specification in this organism elusive. Additionally, upon native CEN deletion, C. albicans can efficiently activate neocentromeres proximal to the native CEN locus, hinting at the importance of CEN-proximal regions. In this study, we examine this CEN-proximity effect and identify factors for CEN specification in C. albicans. We exploit a counterselection assay to isolate cells that can silence a transgene when integrated into the CEN-flanking regions. We show that the frequency of reversible silencing of the transgene decreases from the central core of CEN7 to its peripheral regions. Using publicly available C. albicans high-throughput chromosome conformation capture data, we identify a 25-kb region centering on the CENPA-bound core that acts as CEN-flanking compact chromatin (CFCC). Cis- and trans-chromosomal interactions associated with the CFCC spatially segregates it from bulk chromatin. We further show that neocentromere activation on chromosome 7 occurs within this specified region. Hence, this study identifies a specialized CEN-proximal domain that specifies and restricts the centromeric activity to a unique region.
... Epigenetics is an important regulatory mechanism, in which functionally relevant, heritable, phenotypic modifications occur without changes in the nucleotide sequence (Jablonka and Raz 2009;Weinhold 2006). Epigenetic marks are broadly classified into two groups; nucleotide modifications such as DNA methylation (Bird 2002;Mishra et al. 2011), and the post-translational modifications (PTMs) of proteins (Muller 2018). Epigenetic regulation has been implicated in fundamental processes such as DNA replication, gene expression, cell cycle progression and chromosome segregation (Cullati and Gould 2019;Henikoff and Greally 2016;Jaenisch and Bird 2003;Mishra et al. 2007). ...
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Centromere identity is specified epigenetically by specialized nucleosomes containing the evolutionarily conserved centromeric histone H3 variant (Cse4 in budding yeast, CENP-A in humans) which is essential for faithful chromosome segregation. However, the mechanisms of epigenetic regulation of Cse4 have not been clearly defined. We have identified two kinases, Cdc5 (Plk1 in humans) and Ipl1 (Aurora B kinase in humans) that phosphorylate Cse4 to prevent chromosomal instability (CIN). Cdc5 associates with Cse4 in mitosis and Cdc5-mediated phosphorylation of Cse4 is coincident with the centromeric enrichment of Cdc5 during metaphase. Defects in Cdc5-mediated Cse4 phosphorylation causes CIN, whereas constitutive association of Cdc5 with Cse4 results in lethality. Cse4 is also a substrate for Ipl1 and phospho-mimetic cse4 mutants suppress growth defects of ipl1 and Ipl1 kinetochore substrate mutants, namely dam1 spc34 and ndc80. Ipl1-mediated phosphorylation of Cse4 regulates kinetochore–microtubule interactions and chromosome biorientation. We propose that collaboration of Cdc5- and Ipl1-mediated phosphorylation of Cse4 modulates kinetochore structure and function, and chromosome biorientation. These findings demonstrate how phosphorylation of Cse4 regulates the integrity of the kinetochore, and acts as an epigenetic marker for mitotic control.
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Cytosine methylation, a common form of DNA modification that antagonizes transcription, is found at transposons and repeats in vertebrates, plants and fungi. Here we have mapped DNA methylation in the entire Arabidopsis thaliana genome at high resolution. DNA methylation covers transposons and is present within a large fraction of A. thaliana genes. Methylation within genes is conspicuously biased away from gene ends, suggesting a dependence on RNA polymerase transit. Genic methylation is strongly influenced by transcription: moderately transcribed genes are most likely to be methylated, whereas genes at either extreme are least likely. In turn, transcription is influenced by methylation: short methylated genes are poorly expressed, and loss of methylation in the body of a gene leads to enhanced transcription. Our results indicate that genic transcription and DNA methylation are closely interwoven processes.
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The 10.9x genomic sequence of Candida albicans, the most important human fungal pathogen, was published in 2004. Assembly 19 consisted of 412 supercontigs, of which 266 were a haploid set, since this fungus is diploid and contains an extensive degree of heterozygosity but lacks a complete sexual cycle. However, sequences of specific chromosomes were not determined. Supercontigs from Assembly 19 (183, representing 98.4% of the sequence) were assigned to individual chromosomes purified by pulse-field gel electrophoresis and hybridized to DNA microarrays. Nine Assembly 19 supercontigs were found to contain markers from two different chromosomes. Assembly 21 contains the sequence of each of the eight chromosomes and was determined using a synteny analysis with preliminary versions of the Candida dubliniensis genome assembly, bioinformatics, a sequence tagged site (STS) map of overlapping fosmid clones, and an optical map. The orientation and order of the contigs on each chromosome, repeat regions too large to be covered by a sequence run, such as the ribosomal DNA cluster and the major repeat sequence, and telomere placement were determined using the STS map. Sequence gaps were closed by PCR and sequencing of the products. The overall assembly was compared to an optical map; this identified some misassembled contigs and gave a size estimate for each chromosome. Assembly 21 reveals an ancient chromosome fusion, a number of small internal duplications followed by inversions, and a subtelomeric arrangement, including a new gene family, the TLO genes. Correlations of position with relatedness of gene families imply a novel method of dispersion. The sequence of the individual chromosomes of C. albicans raises interesting biological questions about gene family creation and dispersion, subtelomere organization, and chromosome evolution.