DEEPOMICS FFPE, a deep neural network model, identifies DNA sequencing artifacts from formalin fixed paraffin embedded tissue with high accuracy.

Formalin-fixed, paraffin-embedded (FFPE) tissue specimens are routinely used in pathological diagnosis, but their large number of artifactual mutations complicate the evaluation of companion diagnostics and analysis of next-generation sequencing data. Identification of variants with low allele frequencies is challenging because existing FFPE filtering tools label all low-frequency variants as artifacts. To address this problem, we aimed to develop DEEPOMICS FFPE, an AI model that can classify a true variant from an artifact.

Transcription and chromatin-based surveillance mechanism controls suppression of cryptic antisense transcription.

Phosphorylation of the RNA polymerase II C-terminal domain YSPTSPS consensus sequence coordinates key events during transcription, and its deregulation leads to defects in transcription and RNA processing. Here, we report that the histone deacetylase activity of the fission yeast Hos2/Set3 complex plays an important role in suppressing cryptic initiation of antisense transcription when RNA polymerase II phosphorylation is dysregulated due to the loss of Ssu72 phosphatase. Interestingly, although single Hos2 and Set3 mutants have little effect, loss of Hos2 or Set3 combined with ssu72Δ results in a synergistic increase in antisense transcription globally and correlates with elevated sensitivity to genotoxic agents.

Profiling RNA Polymerase II Phosphorylation Genome-Wide in Fission Yeast.

The RNA polymerase II carboxyl-terminal domain (CTD) consists of tandem repeats of consensus sequence Tyr-Ser-Pro-Thr-Ser-Pro-Ser. Dynamic posttranslational modifications of the CTD generate a CTD code crucial for the cotranscriptional recruitment of factors that control transcription, chromatin modification, and RNA processing. Analysis of CTD phosphorylation by ChIP (Chromatin ImmunoPrecipitation) coupled with high-throughput DNA sequencing (ChIP-seq) is a powerful tool to investigate the changes in CTD phosphorylation during the transcription cycle.

Elongation/Termination Factor Exchange Mediated by PP1 Phosphatase Orchestrates Transcription Termination.

Termination of RNA polymerase II (Pol II) transcription is a key step that is important for 3' end formation of functional mRNA, mRNA release, and Pol II recycling. Even so, the underlying termination mechanism is not yet understood. Here, we demonstrate that the conserved and essential termination factor Seb1 is found on Pol II near the end of the RNA exit channel and the Rpb4/7 stalk.

Histone deacetylation promotes transcriptional silencing at facultative heterochromatin.

It is important to accurately regulate the expression of genes involved in development and environmental response. In the fission yeast Schizosaccharomyces pombe, meiotic genes are tightly repressed during vegetative growth. Despite being embedded in heterochromatin these genes are transcribed and believed to be repressed primarily at the level of RNA.

The conserved protein Seb1 drives transcription termination by binding RNA polymerase II and nascent RNA.

Termination of RNA polymerase II (Pol II) transcription is an important step in the transcription cycle, which involves the dislodgement of polymerase from DNA, leading to release of a functional transcript. Recent studies have identified the key players required for this process and showed that a common feature of these proteins is a conserved domain that interacts with the phosphorylated C-terminus of Pol II (CTD-interacting domain, CID). However, the mechanism by which transcription termination is achieved is not understood.

Exosome Cofactors Connect Transcription Termination to RNA Processing by Guiding Terminated Transcripts to the Appropriate Exonuclease within the Nuclear Exosome.

The yeast Nrd1 interacts with the C-terminal domain (CTD) of RNA polymerase II (RNApII) through its CTD-interacting domain (CID) and also associates with the nuclear exosome, thereby acting as both a transcription termination and RNA processing factor. Previously, we found that the Nrd1 CID is required to recruit the nuclear exosome to the Nrd1 complex, but it was not clear which exosome subunits were contacted. Here, we show that two nuclear exosome cofactors, Mpp6 and Trf4, directly and competitively interact with the Nrd1 CID and differentially regulate the association of Nrd1 with two catalytic subunits of the exosome.

The RNA polymerase II C-terminal domain-interacting domain of yeast Nrd1 contributes to the choice of termination pathway and couples to RNA processing by the nuclear exosome.

The RNA polymerase II (RNApII) C-terminal domain (CTD)-interacting domain (CID) proteins are involved in two distinct RNApII termination pathways and recognize different phosphorylated forms of CTD. To investigate the role of differential CTD-CID interactions in the choice of termination pathway, we altered the CTD-binding specificity of Nrd1 by domain swapping. Nrd1 with the CID from Rtt103 (Nrd1(CID(Rtt103))) causes read-through transcription at many genes, but can also trigger termination where multiple Nrd1/Nab3-binding sites and the Ser(P)-2 CTD co-exist.

Cd2+ binds to Atx1 and affects the physical interaction between Atx1 and Ccc2 in Saccharomyces cerevisiae.

The ATX1 deletion strain of Saccharomyces cerevisiae is more resistant to Cd(2+) than the wild-type. To investigate the function of Atx1 in Cd(2+) toxicity, we used a metal-binding assay to study the interaction between Atx1 and Cd(2+) in vitro. Using circular dichroism and two-hybrid analyses, we found that Atx1 can bind Cd(2+) specifically and that Cd(2+) binding to Atx1 affects the physical interaction between Atx1 and Ccc2.

Cadmium regulates copper homoeostasis by inhibiting the activity of Mac1, a transcriptional activator of the copper regulon, in Saccharomyces cerevisiae.

Cadmium is a toxic metal and the mechanism of its toxicity has been studied in various model systems from bacteria to mammals. We employed Saccharomyces cerevisiae as a model system to study cadmium toxicity at the molecular level because it has been used to identify the molecular mechanisms of toxicity found in higher organisms. cDNA microarray and Northern blot analyses revealed that cadmium salts inhibited the expression of genes related to copper metabolism.

FgEnd1 is a putative component of the endocytic machinery and mediates ferrichrome uptake in F. graminearum.

The function of endocytic pathway in filamentous fungi has remained elusive. Recently, we have identified that FgEnd1, which has a 27% amino acid homology and shares specific EH3 domain with ScEnd3 of Saccharomyces cerevisiae, is a putative member of the endocytic machinery in Fusarium graminearum. The failure of the scend3 mutant to uptake Lucifer yellow (LY) was recovered by introducing FgEnd1 into S.

A novel function of Aft1 in regulating ferrioxamine B uptake: Aft1 modulates Arn3 ubiquitination in Saccharomyces cerevisiae.

Aft1 is a transcriptional activator in Saccharomyces cerevisiae that responds to iron availability and regulates the expression of genes in the iron regulon, such as FET3, FTR1 and the ARN family. Using a two-hybrid screen, we found that Aft1 physically interacts with the FOB (ferrioxamine B) transporter Arn3. This interaction modulates the ability of Arn3 to take up FOB.

A novel role for thioredoxin reductase in the iron metabolism of S. cerevisiae.

Intracellular levels of iron are tightly regulated. Saccharomyces cerevisiae uses well-defined pathways to extract iron molecules from the environment. Once inside the cell, the iron molecules must be transferred to target sites via an intracellular iron transporter.

Functional characterization of Hsp33 protein from Bacillus psychrosaccharolyticus; additional function of HSP33 on resistance to solvent stress.

Psychrophiles have been known as efficient organism to degrade organic solvent. To investigate the mechanism of solvent stress and identify the factors that affect the solvent stress in psychrophiles, we selected Bacillus psychrosaccharolyticus one of the psychrophiles and two-dimensional gel electrophoresis was performed. Among the protein spots analyzed by 2-DE, five spots induced in 3% IPA stress conditions were identified by MS/MS, and one of these spots was identified as a Hsp33 family.