Deletion of the Candida albicans TLO gene family using CRISPR-Cas9 mutagenesis allows characterisation of functional differences in α-, β- and γ- TLO gene function.

The Candida albicans genome contains between ten and fifteen distinct TLO genes that all encode a Med2 subunit of Mediator. In order to investigate the biological role of Med2/Tlo in C. albicans we deleted all fourteen TLO genes using CRISPR-Cas9 mutagenesis.

Stress combined with loss of the Candida albicans SUMO protease Ulp2 triggers selection of aneuploidy via a two-step process.

A delicate balance between genome stability and instability ensures genome integrity while generating genetic diversity, a critical step for evolution. Indeed, while excessive genome instability is harmful, moderated genome instability can drive adaptation to novel environments by maximising genetic variation. Candida albicans, a human fungal pathogen that colonises different parts of the human body, adapts rapidly and frequently to different hostile host microenvironments.

On and Off: Epigenetic Regulation of Morphological Switches.

The human fungal pathogen 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.

The Genome of the CTG(Ser1) Yeast Is Plastic.

Microorganisms need to adapt to environmental changes, and genome plasticity can lead to rapid adaptation to hostile environments by increasing genetic diversity. Here, we investigate genome plasticity in the CTG(Ser1) yeast , an organism with an enormous potential for second-generation biofuel production. We demonstrate that has an intrinsically plastic genome and that different isolates have genomes with distinct chromosome organizations.

Chromatin-Mediated Regulation of Genome Plasticity in Human Fungal Pathogens.

Human fungal pathogens, such as , and , 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.

Chromatin Profiling of the Repetitive and Nonrepetitive Genomes of the Human Fungal Pathogen Candida albicans.

Eukaryotic genomes are packaged into chromatin structures that play pivotal roles in regulating all DNA-associated processes. Histone posttranslational modifications modulate chromatin structure and function, leading to rapid regulation of gene expression and genome stability, key steps in environmental adaptation. , a prevalent fungal pathogen in humans, can rapidly adapt and thrive in diverse host niches.

Sir2 regulates stability of repetitive domains differentially in the human fungal pathogen Candida albicans.

DNA repeats, found at the ribosomal DNA locus, telomeres and subtelomeric regions, are unstable sites of eukaryotic genomes. A fine balance between genetic variability and genomic stability tunes plasticity of these chromosomal regions. This tuning mechanism is particularly important for organisms such as microbial pathogens that utilise genome plasticity as a strategy for adaptation.

The Chromatin of Candida albicans Pericentromeres Bears Features of Both Euchromatin and Heterochromatin.

Centromeres, sites of kinetochore assembly, are important for chromosome stability and integrity. Most eukaryotes have regional centromeres epigenetically specified by the presence of the histone H3 variant CENP-A. CENP-A chromatin is often surrounded by pericentromeric regions packaged into transcriptionally silent heterochromatin.

Candida albicans repetitive elements display epigenetic diversity and plasticity.

Transcriptionally silent heterochromatin is associated with repetitive DNA. It is poorly understood whether and how heterochromatin differs between different organisms and whether its structure can be remodelled in response to environmental signals. Here, we address this question by analysing the chromatin state associated with DNA repeats in the human fungal pathogen Candida albicans.

The RFTS domain of Raf2 is required for Cul4 interaction and heterochromatin integrity in fission yeast.

Centromeric heterochromatin assembly in fission yeast is critical for faithful chromosome segregation at mitosis. Its assembly requires a concerted pathway of events whereby the RNA interference (RNAi) pathway guides H3K9 methylation to target sequences. H3K9 methylation, a hallmark of heterochromatin structure, is mediated by the single histone methyltransferase Clr4 (equivalent to metazoan Suv3-9), a component of the CLRC complex.

Distinct roles for Sir2 and RNAi in centromeric heterochromatin nucleation, spreading and maintenance.

Epigenetically regulated heterochromatin domains govern essential cellular activities. A key feature of heterochromatin domains is the presence of hypoacetylated nucleosomes, which are methylated on lysine 9 of histone H3 (H3K9me). Here, we investigate the requirements for establishment, spreading and maintenance of heterochromatin using fission yeast centromeres as a paradigm.

Raf1 Is a DCAF for the Rik1 DDB1-like protein and has separable roles in siRNA generation and chromatin modification.

Non-coding transcription can trigger histone post-translational modifications forming specialized chromatin. In fission yeast, heterochromatin formation requires RNAi and the histone H3K9 methyltransferase complex CLRC, composed of Clr4, Raf1, Raf2, Cul4, and Rik1. CLRC mediates H3K9 methylation and siRNA production; it also displays E3-ubiquitin ligase activity in vitro.

Building centromeres: home sweet home or a nomadic existence?

Centromere assembly and propagation is governed by genetic and epigenetic mechanisms. A centromere-specific histone H3 variant, CENP-A is strongly favored as the epigenetic mark that specifies centromere identity. Despite the critical importance of centromere function, centromeric sequences are not conserved.

Hairpin RNA induces secondary small interfering RNA synthesis and silencing in trans in fission yeast.

RNA interference (RNAi) is widespread in eukaryotes and regulates gene expression transcriptionally or post-transcriptionally. In fission yeast, RNAi is tightly coupled to template transcription and chromatin modifications that establish heterochromatin in cis. Exogenous double-stranded RNA (dsRNA) triggers seem to induce heterochromatin formation in trans only when certain silencing proteins are overexpressed.

Molecular biology: silencing unlimited.

Heterochromatin domains are essential for normal chromosome functions. The Eri1 ribonuclease is a negative regulator of the RNA interference machinery; recent studies have shown that, in fission yeast lacking Eri1, heterochromatin formation is more promiscuous.

X-chromosome targeting and dosage compensation are mediated by distinct domains in MSL-3.

In Drosophila, dosage compensation of X-linked genes is achieved by transcriptional upregulation of the male X chromosome. Genetic and biochemical studies have demonstrated that male-specific lethal (MSL) proteins together with roX RNAs regulate this process. Here, we show that MSL-3 is essential for cell viability and that three domains in the protein have distinct roles in dosage compensation.

Nuclear pore components are involved in the transcriptional regulation of dosage compensation in Drosophila.

Dosage compensation in Drosophila is dependent on MSL proteins and involves hypertranscription of the male X chromosome, which ensures equal X-linked gene expression in both sexes. Here, we report the purification of enzymatically active MSL complexes from Drosophila embryos, Schneider cells, and human HeLa cells. We find a stable association of the histone H4 lysine 16-specific acetyltransferase MOF with the RNA/protein containing MSL complex as well as with an evolutionary conserved complex.

A general precursor ion-like scanning mode on quadrupole-TOF instruments compatible with chromatographic separation.

MS protein identification and quantitation are key proteomic techniques in biological research. Besides identification of proteins, MS is used increasingly to characterize secondary protein modifications. This often requires trimming the analytical strategy to a specific type of modification.

Structure of the chromo barrel domain from the MOF acetyltransferase.

We report here the structure of the putative chromo domain from MOF, a member of the MYST family of histone acetyltransferases that acetylates histone H4 at Lys-16 and is part of the dosage compensation complex in Drosophila. We found that the structure of this domain is a beta-barrel that is distinct from the alpha + beta fold of the canonical chromo domain. Despite the differences, there are similarities that support an evolutionary relationship between the two domains, and we propose the name "chromo barrel.

MOF-regulated acetylation of MSL-3 in the Drosophila dosage compensation complex.

Dosage compensation ensures equal expression of X-linked genes in males and females. In Drosophila, equalization is achieved by hypertranscription of the male X chromosome. This process requires an RNA/protein containing dosage compensation complex (DCC).

Identification of the enhancer binding protein MBF-1 of the sea urchin modulator alpha-H2A histone gene.

The modulator of the sea urchin alpha-H2A histone gene promoter is the only enhancer identified in the alpha-histone gene cluster. Binding of a single factor, denoted MBF-1, has previously detected in nuclear extracts from morula and gastrula embryos. Here, we describe the cloning of MBF-1 by screening a cDNA expression library with a tandem array of modulator binding sites.