The next-generation sequencing-chess problem.

The development of next-generation sequencing (NGS) technologies paved the way for studying the spatiotemporal coordination of cellular processes along the genome. However, data sets are commonly limited to a few time points, and missing information needs to be interpolated. Most models assume that the studied dynamics are similar between individual cells, so that a homogeneous cell culture can be represented by a population-wide average.

Mediator complex in transcription regulation and DNA repair: Relevance for human diseases.

The Mediator complex is an essential coregulator of RNA polymerase II transcription. More recent developments suggest Mediator functions as a link between transcription regulation, genome organisation and DNA repair mechanisms including nucleotide excision repair, base excision repair, and homologous recombination. Dysfunctions of these processes are frequently associated with human pathologies, and growing evidence shows Mediator involvement in cancers, neurological, metabolic and infectious diseases.

A genome-wide comprehensive analysis of nucleosome positioning in yeast.

In eukaryotic cells, the one-dimensional DNA molecules need to be tightly packaged into the spatially constraining nucleus. Folding is achieved on its lowest level by wrapping the DNA around nucleosomes. Their arrangement regulates other nuclear processes, such as transcription and DNA repair.

Functional interplay between Mediator and RSC chromatin remodeling complex controls nucleosome-depleted region maintenance at promoters.

Chromatin organization is crucial for transcriptional regulation in eukaryotes. Mediator is an essential and conserved co-activator thought to act in concert with chromatin regulators. However, it remains largely unknown how their functions are coordinated.

A quantitative modelling approach for DNA repair on a population scale.

The great advances of sequencing technologies allow the in vivo measurement of nuclear processes-such as DNA repair after UV exposure-over entire cell populations. However, data sets usually contain only a few samples over several hours, missing possibly important information in between time points. We developed a data-driven approach to analyse CPD repair kinetics over time in Saccharomyces cerevisiae.

Genomic analysis of Rad26 and Rad1-Rad10 reveals differences in their dependence on Mediator and RNA polymerase II.

Mediator is a conserved coregulator playing a key role in RNA polymerase (Pol) II transcription. Mediator also links transcription and nucleotide excision repair (NER) via a direct contact with Rad2/ERCC5(XPG) endonuclease. In this work, we analyzed the genome-wide distribution of Rad26/ERCC6(CSB) and Rad1-Rad10/ERCC4(XPF)-ERCC1, addressing the question of a potential link of these proteins with Mediator and Pol II in yeast Our genomic analyses reveal that Rad1-Rad10 and Rad26 are present on the yeast genome in the absence of genotoxic stress, especially at highly transcribed regions, with Rad26 binding strongly correlating with that of Pol II.

Microfluidic platform for monitoring mutation accumulation.

Mutations in DNA have large-ranging consequences, from evolution to disease. Many mechanisms contribute to mutational processes such as dysfunctions in DNA repair pathways and exogenous or endogenous mutagen exposures. Model organisms and mutation accumulation (MA) experiments are indispensable to study mutagenesis.

A Role for the Mre11-Rad50-Xrs2 Complex in Gene Expression and Chromosome Organization.

Mre11-Rad50-Xrs2 (MRX) is a highly conserved complex with key roles in various aspects of DNA repair. Here, we report a new function for MRX in limiting transcription in budding yeast. We show that MRX interacts physically and colocalizes on chromatin with the transcriptional co-regulator Mediator.

Mediator Roles Going Beyond Transcription.

Dysfunctions of nuclear processes including transcription and DNA repair lead to severe human diseases. Gaining an understanding of how these processes operate in the crowded context of chromatin can be particularly challenging. Mediator is a large multiprotein complex conserved in eukaryotes with a key coactivator role in the regulation of RNA polymerase (Pol) II transcription.

Mammalian Mediator as a Functional Link between Enhancers and Promoters.

In this issue of Cell, Casellas and colleagues provide insights into the structural and functional aspects of the mammalian multi-subunit Mediator complex, a conserved and essential transcriptional coregulator. Combining cryo-EM, genetic, and genomic analyses, the work sheds light on Mediator's mode of action as a functional bridge between enhancers and promoters.

Functional interplay between Mediator and RNA polymerase II in Rad2/XPG loading to the chromatin.

Transcription and maintenance of genome integrity are fundamental cellular functions. Deregulation of transcription and defects in DNA repair lead to serious pathologies. The Mediator complex links RNA polymerase (Pol) II transcription and nucleotide excision repair via Rad2/XPG endonuclease.

Transcription regulation by the Mediator complex.

Alterations in the regulation of gene expression are frequently associated with developmental diseases or cancer. Transcription activation is a key phenomenon in the regulation of gene expression. In all eukaryotes, mediator of RNA polymerase II transcription (Mediator), a large complex with modular organization, is generally required for transcription by RNA polymerase II, and it regulates various steps of this process.

Toward understanding of the mechanisms of Mediator function in vivo: Focus on the preinitiation complex assembly.

Mediator is a multisubunit complex conserved in eukaryotes that plays an essential coregulator role in RNA polymerase (Pol) II transcription. Despite intensive studies of the Mediator complex, the molecular mechanisms of its function in vivo remain to be fully defined. In this review, we will discuss the different aspects of Mediator function starting with its interactions with specific transcription factors, its recruitment to chromatin and how, as a coregulator, it contributes to the assembly of transcription machinery components within the preinitiation complex (PIC) in vivo and beyond the PIC formation.

Functional interplay between Mediator and TFIIB in preinitiation complex assembly in relation to promoter architecture.

Mediator is a large coregulator complex conserved from yeast to humans and involved in many human diseases, including cancers. Together with general transcription factors, it stimulates preinitiation complex (PIC) formation and activates RNA polymerase II (Pol II) transcription. In this study, we analyzed how Mediator acts in PIC assembly using in vivo, in vitro, and in silico approaches.

Mediator independently orchestrates multiple steps of preinitiation complex assembly in vivo.

Mediator is a large multiprotein complex conserved in all eukaryotes, which has a crucial coregulator function in transcription by RNA polymerase II (Pol II). However, the molecular mechanisms of its action in vivo remain to be understood. Med17 is an essential and central component of the Mediator head module.

PHD and TFIIS-Like domains of the Bye1 transcription factor determine its multivalent genomic distribution.

The BYpass of Ess1 (Bye1) protein is a putative S. cerevisiae transcription factor homologous to the human cancer-associated PHF3/DIDO family of proteins. Bye1 contains a Plant Homeodomain (PHD) and a TFIIS-like domain.

Mediator links transcription and DNA repair by facilitating Rad2/XPG recruitment.

Mediator is a large multiprotein complex conserved in all eukaryotes. The crucial function of Mediator in transcription is now largely established. However, we found that this complex also plays an important role by connecting transcription with DNA repair.

Genomic binding of Pol III transcription machinery and relationship with TFIIS transcription factor distribution in mouse embryonic stem cells.

RNA polymerase (Pol) III synthesizes the tRNAs, the 5S ribosomal RNA and a small number of untranslated RNAs. In vitro, it also transcribes short interspersed nuclear elements (SINEs). We investigated the distribution of Pol III and its associated transcription factors on the genome of mouse embryonic stem cells using a highly specific tandem ChIP-Seq method.

Direct interaction of RNA polymerase II and mediator required for transcription in vivo.

Gene transcription is highly regulated. Altered transcription can lead to cancer or developmental diseases. Mediator, a multisubunit complex conserved among eukaryotes, is generally required for RNA polymerase II (Pol II) transcription.

A gene-specific requirement of RNA polymerase II CTD phosphorylation for sexual differentiation in S. pombe.

Background: The switch from cellular proliferation to differentiation occurs to a large extent through specific programs of gene expression. In fission yeast, the master regulator of sexual differentiation, ste11, is induced by environmental conditions leading to mating and meiosis.

Results: We show that phosphorylation of serine 2 (S2P) in the C-terminal domain of the largest subunit of the RNA polymerase II (PolII) enzyme by the Lsk1 cyclin-dependent kinase has only a minor impact on global gene expression during vegetative growth but is critical for the induction of ste11 transcription during sexual differentiation.

Structure-function analysis of RNA polymerases I and III.

Recent advances in elucidating the structure of yeast Pol I and III are based on a combination of X-ray crystal analysis, electron microscopy and homology modelling. They allow a better comparison of the three eukaryotic nuclear RNA polymerases, underscoring the most obvious difference existing between the three enzymes, which lies in the existence of additional Pol-I-specific and Pol-III-specific subunits. Their location on the cognate RNA polymerases is now fairly well known, suggesting precise hypotheses as to their function in transcription during initiation, elongation, termination and/or reinitiation.

Mutations of RNA polymerase II activate key genes of the nucleoside triphosphate biosynthetic pathways.

The yeast URA2 gene, encoding the rate-limiting enzyme of UTP biosynthesis, is transcriptionally activated by UTP shortage. In contrast to other genes of the UTP pathway, this activation is not governed by the Ppr1 activator. Moreover, it is not due to an increased recruitment of RNA polymerase II at the URA2 promoter, but to its much more effective progression beyond the URA2 mRNA start site(s).

Mediator-dependent recruitment of TFIIH modules in preinitiation complex.

In vitro, without Mediator, the association of general transcription factors (GTF) and RNA polymerase II (Pol II) in preinitiation complexes (PIC) occurs in an orderly fashion. In this work, we explore the in vivo function of Mediator in GTF recruitment to PIC. A direct interaction between Med11 Mediator head subunit and Rad3 TFIIH subunit was identified.

Genome-wide location analysis reveals a role of TFIIS in RNA polymerase III transcription.

TFIIS is a transcription elongation factor that stimulates transcript cleavage activity of arrested RNA polymerase II (Pol II). Recent studies revealed that TFIIS has also a role in Pol II transcription initiation. To improve our understanding of TFIIS function in vivo, we performed genome-wide location analysis of this factor.