Chemical modification of nucleotide bases in DNA provides one mechanism for conveying information in addition to the genetic code. 5-methylcytosine (5mC) represents the most common chemically ...modified base in eukaryotic genomes. Sometimes referred to simply as DNA methylation, in eukaryotes 5mC is most prevalent at CpG dinucleotides and is frequently associated with transcriptional repression of transposable elements. However, 5mC levels and distributions are variable across phylogenies, and emerging evidence suggests that the functions of DNA methylation may be more diverse and complex than was previously appreciated. We summarize the current understanding of DNA methylation profiles and functions in different eukaryotic lineages.
Although DNA methyltransferase content is generally conserved across eukaryotes, there is extensive variation in how this modified base is used for a variety of cellular processes.The manner in which DNA methyltransferases are recruited to target sequences leads to a diversity of genome-wide DNA methylation patterns across eukaryotes.Continued exploration of DNA methylation patterns and DNA methyltransferase content in diverse eukaryotic lineages will lead to an expanded understanding of the mechanism by which the modified base functions in genomes.
Single nucleotide polymorphism (SNP) discovery and genotyping are essential to genetic mapping. There remains a need for a simple, inexpensive platform that allows high-density SNP discovery and ...genotyping in large populations. Here we describe the sequencing of restriction-site associated DNA (RAD) tags, which identified more than 13,000 SNPs, and mapped three traits in two model organisms, using less than half the capacity of one Illumina sequencing run. We demonstrated that different marker densities can be attained by choice of restriction enzyme. Furthermore, we developed a barcoding system for sample multiplexing and fine mapped the genetic basis of lateral plate armor loss in threespine stickleback by identifying recombinant breakpoints in F(2) individuals. Barcoding also facilitated mapping of a second trait, a reduction of pelvic structure, by in silico re-sorting of individuals. To further demonstrate the ease of the RAD sequencing approach we identified polymorphic markers and mapped an induced mutation in Neurospora crassa. Sequencing of RAD markers is an integrated platform for SNP discovery and genotyping. This approach should be widely applicable to genetic mapping in a variety of organisms.
Polycomb group (PcG) proteins form multiple complexes that direct assembly of transcriptionally repressed chromatin, an essential process for multicellular development. Recent studies in plants and ...animals have uncovered surprising complexity in the form of multiple, variant PcG complexes governed by an extensive regulatory network. PcG proteins are absent from major yeast models, but recent research revealed minimal PcG systems in several well-studied fungi. In both animals and fungi, PcG complexes are responsive to local chromatin structure, but important aspects of PcG regulation remain unknown. Moreover, how PcG complexes repress transcription is poorly understood. The emergence of new fungal models to study PcG proteins is an important development that will accelerate understanding of this important chromatin regulation pathway.
Histone posttranslational modifications (HPTMs) are involved in chromatin-based regulation of fungal secondary metabolite biosynthesis (SMB) in which the corresponding genes-usually physically linked ...in co-regulated clusters-are silenced under optimal physiological conditions (nutrient-rich) but are activated when nutrients are limiting. The exact molecular mechanisms by which HPTMs influence silencing and activation, however, are still to be better understood. Here we show by a combined approach of quantitative mass spectrometry (LC-MS/MS), genome-wide chromatin immunoprecipitation (ChIP-seq) and transcriptional network analysis (RNA-seq) that the core regions of silent A. nidulans SM clusters generally carry low levels of all tested chromatin modifications and that heterochromatic marks flank most of these SM clusters. During secondary metabolism, histone marks typically associated with transcriptional activity such as H3 trimethylated at lysine-4 (H3K4me3) are established in some, but not all gene clusters even upon full activation. KdmB, a Jarid1-family histone H3 lysine demethylase predicted to comprise a BRIGHT domain, a zinc-finger and two PHD domains in addition to the catalytic Jumonji domain, targets and demethylates H3K4me3 in vivo and mediates transcriptional downregulation. Deletion of kdmB leads to increased transcription of about ~1750 genes across nutrient-rich (primary metabolism) and nutrient-limiting (secondary metabolism) conditions. Unexpectedly, an equally high number of genes exhibited reduced expression in the kdmB deletion strain and notably, this group was significantly enriched for genes with known or predicted functions in secondary metabolite biosynthesis. Taken together, this study extends our general knowledge about multi-domain KDM5 histone demethylases and provides new details on the chromatin-level regulation of fungal secondary metabolite production.
Globally, there are several million individuals with life-threatening invasive fungal diseases such as candidiasis, aspergillosis, cryptococcosis, Pneumocystis pneumonia (PCP), and mucormycosis. The ...mortality rate for these diseases generally exceeds 40%. Annual medical costs to treat these invasive fungal diseases in the United States exceed several billion dollars. In addition to AIDS patients, the risks of invasive mycoses are increasingly found in immune-impaired individuals or in immunosuppressed patients following stem cell or organ transplant or implantation of medical devices. Current antifungal drug therapies are not meeting the challenge, because (1) at safe doses, they do not provide sufficient fungal clearance to prevent reemergence of infection; (2) most become toxic with extended use; (3) drug-resistant fungal isolates are emerging; and (4) only one new class of antifungal drugs has been approved for clinical use in the last 2 decades. DectiSomes represent a novel design of drug delivery to drastically increase drug efficacy. Antifungals packaged in liposomes are targeted specifically to where the pathogen is, through binding to the fungal cell walls or exopolysaccharide matrices using the carbohydrate recognition domains of pathogen receptors. Relative to untargeted liposomal drug, DectiSomes show order of magnitude increases in the binding to and killing of Candida albicans, Cryptococcus neoformans, and Aspergillus fumigatus in vitro and similarly improved efficacy in mouse models of pulmonary aspergillosis. DectiSomes have the potential to usher in a new antifungal drug treatment paradigm.
The segregation of eukaryotic genomes into euchromatin and heterochromatin represents a fundamental and poorly understood process. Here, we demonstrate that genome-wide establishment of ...heterochromatin is triggered by the maternal to zygotic transition (MZT) during zebrafish embryogenesis. We find that prior to MZT, zebrafish lack hallmarks of heterochromatin including histone H3 lysine 9 trimethylation (H3K9me3) and condensed chromatin ultrastructure. Global establishment of heterochromatic features occurs following MZT and requires both activation of the zygotic genome and degradation of maternally deposited RNA. Mechanistically, we demonstrate that zygotic transcription of the micro RNA miR-430 promotes degradation of maternal RNA encoding the chromatin remodeling protein Smarca2, and that clearance of Smarca2 is required for global heterochromatin establishment in the early embryo. Our results identify MZT as a key developmental regulator of heterochromatin establishment during vertebrate embryogenesis and uncover functions for Smarca2 in protecting the embryonic genome against heterochromatinization.
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Combatting the ever-increasing threat from invasive fungal pathogens faces numerous fundamental challenges, including constant human exposure to large reservoirs of species in the ...environment, the increasing population of immunocompromised or immunosuppressed individuals, the unsatisfactory efficacy of current antifungal drugs and their associated toxicity, and the scientific and economic barriers limiting a new antifungal pipeline. DectiSomes represent a new drug delivery platform that enhances antifungal efficacy for diverse fungal pathogens and reduces host toxicity for current and future antifungals. DectiSomes employ pathogen receptor proteins − C-type lectins − to target drug-loaded liposomes to conserved fungal cognate ligands and away from host cells. DectiSomes represent one leap forward for urgently needed effective pan-antifungal therapy. Herein, we discuss the problems of battling fungal diseases and the state of DectiSome development.
Around one-third of Americans reported they were unwilling to get a COVID-19 vaccine in April 2021. This focus group study aimed to provide insights on the factors contributing to unvaccinated ...adults' hesitancy or refusal to get vaccinated with COVID-19 vaccines.
Ipsos recruited 59 unvaccinated US adults who were vaccine hesitant (i.e., conflicted about or opposed to receiving a COVID-19 vaccination) using the Ipsos KnowledgePanel. Trained facilitators led a total of 10 focus groups via video-conference in March and April 2021. Two coders manually coded the data from each group using a coding frame based on the focus group discussion guide. The coding team collaborated in analyzing the data for key themes.
Data analysis of transcripts from the focus groups illuminated four main themes associated with COVID-19 vaccine hesitancy: lack of trust in experts and institutions; concern about the safety of COVID-19 vaccines; resistance towards prescriptive guidance and restrictions; and, despite personal reluctance or unwillingness to get vaccinated, acceptance of others getting vaccinated.
Vaccine confidence communication strategies should address individual concerns, describe the benefits of COVID-19 vaccination, and highlight evolving science using factural and neutral presentations of information to foster trust.
Trimethylated lysine 27 on histone H3 (H3K27me3) is present in Drosophila, Arabidopsis. worms, and mammals, but is absent from yeasts that have been examined. We identified and analyzed H3K27me3 in ...the filamentous fungus Neurospora crassa and in other Neurospora species. H3K27me3 covers 6.8% of the N. crassa genome, encompassing 223 domains, including 774 genes, all of which are transcriptionally silent N. crassa H3K27me3-marked genes are less conserved than unmarked genes and only ~35% of genes marked by H3K27me3 in N. crassa are also H3K27me3-marked in Neurospora discreta and Neurospora tetrasperma. We found that three components of the Neurospora Polycomb repressive complex 2 (PRC2)—Su-(var) 3-9; E(z); Trrthorax (SET)-7, embryonic ectoderm development (EED), and SU(Z)12 (suppressor of zeste12)—are required for H3K27me3, whereas the fourth component Neurospora protein 55 (an N. crassa homolog of p55/RbAp48), is critical for H3K27me3 only at subtelomeric domains. Loss of H3K27me3, caused by deletion of the gene encoding the catalytic PRC2 subunit, set-7, resulted in up-regulation of 130 genes, including genes in both H3K27me3-marked and unmarked regions.
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