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Genotoxic stress resulting from DNA damage is resolved through a signaling cascade known as the DNA Damage Response (DDR). The repair of damaged DNA is essential for cell survival, often requiring the DDR to attenuate other cellular processes such as the cell cycle, DNA replication, and transcription of genes not involved in DDR. The complex relationship between DDR and transcription has only recently been investigated. Transcription can facilitate the DDR in response to double-strand breaks (DSBs) and stimulate nucleotide excision repair (NER). However, transcription may need to be reduced to prevent potential interference with the repair machinery. In this review, we discuss various mechanisms that regulate transcription repression in response to different types of DNA damage, categorizing them by their range and duration of effect. Finally, we explore various models of transcription recovery following DNA damage-induced repression. [ABSTRACT FROM AUTHOR]

Transfer RNAs (tRNAs) are small non-coding RNAs (sncRNAs) essential for protein translation. Emerging evidence suggests that tRNAs can also be processed into smaller fragments, tRNA-derived small RNAs (tsRNAs), a novel class of sncRNAs with powerful applications and high biological relevance to cancer. tsRNAs biogenesis is heterogeneous and involves different ribonucleases, such as Angiogenin and Dicer. For many years, tsRNAs were thought to be just degradation products. However, accumulating evidence shows their roles in gene expression: either directly via destabilising the mRNA or the ribosomal machinery, or indirectly via regulating the expression of ribosomal components. Furthermore, tsRNAs participate in various biological processes linked to cancer, including apoptosis, cell cycle, immune response, and retroviral insertion into the human genome. It is emerging that tsRNAs have significant therapeutic potential. Endogenous tsRNAs can be used as cancer biomarkers, while synthetic tsRNAs and antisense oligonucleotides can be employed to regulate gene expression. In this review, we are recapitulating the regulatory roles of tsRNAs, with a focus on cancer biology. [ABSTRACT FROM AUTHOR]

MicroRNAs (miRNAs) are small regulatory noncoding RNAs that are generated in the canonical RNA interference (RNAi) pathway. Drosha, DiGeorge syndrome critical region 8 (DGCR8) and Dicer are key players in miRNA biogenesis. Argonaute (Ago) proteins bind to miRNAs and are guided by them to find messenger RNA targets and carry out post‐transcriptional silencing of protein‐coding genes. Recently, emerging evidence suggests that RNAi factors have a range of noncanonical functions that are beyond miRNA biogenesis. These functions pertain to various biological processes, such as development, transcriptional regulation, RNA processing and maintenance of genome integrity. Here, we review recent literature reporting miRNA‐independent, noncanonical functions of Drosha, DGCR8, Dicer and Ago proteins and discuss the importance of these functions. [ABSTRACT FROM AUTHOR]

Dicer is a key component of RNA interference (RNAi) and well-known for its role in biogenesis of micro (mi)RNA in the cytoplasm. Increasing evidence suggests that mammalian Dicer is also present and active in the nucleus. We have previously shown that phosphorylated human Dicer associates with chromatin in response to DNA damage and processes double-stranded (ds)RNA in the nucleus. However, a recent study by Much et al. investigated endogenously tagged HA-Dicer both in primary mouse embryonic fibroblasts (PMEFs) as well as adult homozygous viable and fertile HA-Dicer mice under physiological conditions and concluded that murine Dicer is exclusively cytoplasmic. The authors challenged several findings, reporting functions of Dicer in mammalian nuclei. We have re- investigated this issue by applying subcellular fractionation, super-resolution microscopy followed by 3D reconstitution, and phospho-Dicer-specific antibodies using the same HA- Dicer PMEF cell line. Our data show that a small fraction of the murine HA-Dicer pool, approximately 5%, localises in the nucleus and is phosphorylated upon DNA damage. We propose that Dicer localisation is dynamic and not exclusively cytoplasmic, particularly in cells exposed to DNA damage. [ABSTRACT FROM AUTHOR]

The RNase III enzymes Drosha and Dicer are essential for the production of small non-coding RNAs (ncRNAs). In canonical RNAi, microRNAs (miRNAs) regulate gene expression by post-transcriptional gene silencing. In non-canonical RNAi, nuclear RNAi factors generate small ncRNAs that are essential for transcriptional gene silencing. Recent evidence points to the existence of additional non-canonical nuclear RNAi functions in various organisms, including in genome maintenance and editing, as well as in DNA repair. Drosha and Dicer directly regulate gene expression and RNA metabolism at different stages, such as transcriptional initiation and termination, and the processing of various RNA species, including pre-mRNAs. Furthermore, Dicer isoforms were recently discovered and attributed with roles in apoptosis, development and disease. [ABSTRACT FROM AUTHOR]

Non-coding transcripts play an important role in gene expression regulation in all species, including budding and fission yeast. Such regulatory transcripts include intergenic ncRNA (non-coding RNA), 5 and 3 UTRs, introns and antisense transcripts. In the present review, we discuss advantages and limitations of recently developed sequencing techniques, such as ESTs, DNA microarrays, RNA-Seq (RNA sequencing), DRS (direct RNA sequencing) and TIF-Seq (transcript isoform sequencing). We provide an overview of methods applied in yeast and how each of them has contributed to our knowledge of gene expression regulation and transcription. [ABSTRACT FROM AUTHOR]

We show that convergent transcription induces transcriptional gene silencing (TGS) in trans for both fission yeast and mammalian cells. This method has advantages over existing strategies to induce gene silencing. Previous studies in fission yeast have characterized TGS as a cis-specific process involving RNA interference that maintains heterochromatic regions such as centromeres. In contrast, in mammalian cells, gene silencing is known to occur through a post-transcriptional mechanism that uses exogenous short interfering RNAs or endogenous microRNAs to inactivate mRNA. We now show that the introduction of convergent transcription plasmids into either Schizosaccharomyces pombe or mammalian cells allows the production of double-stranded RNA from inserted gene fragments, resulting in TGS of endogenous genes. We predict that using convergent transcription to induce gene silencing will be a generally useful strategy and allow for a fuller molecular understanding of the biology of TGS. [ABSTRACT FROM AUTHOR]

Summary: Transcription analyses reported in these studies reveal that convergent genes in S. pombe generate overlapping transcripts in the G1 phase of the cell cycle. We show that this double-strand (ds) RNA induces localized RNAi (Dicer and RITS) dependent transient heterochromatin structures including histone H3 lysine 9 trimethylation marks and Swi6 association. Consequently cohesin is recruited to these chromosomal positions through interaction with Swi6. In G2, localized cohesin is further concentrated into the intergenic regions of the convergent genes tested. This results in a block to further dsRNA formation by promoting gene-proximal transcription termination between the convergent genes. Cohesin release at mitosis leads to a new G1 phase with repeated dsRNA formation, transient heterochromatin, and cohesin recruitment. Our results uncover a hitherto unanticipated role for cohesin and further suggest a widespread role for the selective formation of dsRNA, heterochromatin, and subsequent cohesin recruitment in regulated transcriptional termination. [Copyright &y& Elsevier]

Non-coding RNA (ncRNA) has recently emerged as a vital component of the DNA damage response (DDR), which was previously believed to be solely regulated by proteins. Many species of ncRNA can directly or indirectly influence DDR and enhance DNA repair, particularly in response to double-strand DNA breaks, which may hold therapeutic potential in the context of cancer. These include long non-coding RNA (lncRNA), microRNA, damage-induced lncRNA, DNA damage response small RNA, and DNA:RNA hybrid structures, which can be categorised as cis or trans based on the location of their synthesis relative to DNA damage sites. Mechanisms of RNA-dependent DDR include the recruitment or scaffolding of repair factors at DNA break sites, the regulation of repair factor expression, and the stabilisation of repair intermediates. DDR can also be communicated intercellularly via exosomes, leading to bystander responses in healthy neighbour cells to generate a population-wide response to damage. Many microRNA species have been directly implicated in the propagation of bystander DNA damage, autophagy, and radioresistance, which may prove significant for enhancing cancer treatment via radiotherapy. Here, we review recent developments centred around ncRNA and their contributions to intracellular and intercellular DDR mechanisms. [ABSTRACT FROM AUTHOR]

There is extensive transcription throughout the eukaryotic genome resulting in both antisense transcripts from coding regions and cryptic unstable transcripts (CUTs) from intergenic regions. In this issue, demonstrate in the budding yeast that antisense transcripts, if stabilized by exosome impairment, are able to mediate gene silencing via the recruitment of histone deacetylases. [Copyright &y& Elsevier]

Double-strand breaks (DSBs) are the most lethal form of DNA damage. Transcriptional activity at DSBs, as well as transcriptional repression around DSBs, are both required for efficient DNA repair. The chromatin landscape defines and coordinates these two opposing events. However, how the open and condensed chromatin architecture is regulated remains unclear. Here, we show that the GATAD2B–NuRD complex associates with DSBs in a transcription- and DNA:RNA hybrid-dependent manner, to promote histone deacetylation and chromatin condensation. This activity establishes a spatio-temporal boundary between open and closed chromatin, which is necessary for the correct termination of DNA end resection. The lack of the GATAD2B–NuRD complex leads to chromatin hyperrelaxation and extended DNA end resection, resulting in homologous recombination (HR) repair failure. Our results suggest that the GATAD2B–NuRD complex is a key coordinator of the dynamic interplay between transcription and the chromatin landscape, underscoring its biological significance in the RNA-dependent DNA damage response. Synopsis: Regulation of open and condensed chromatin architecture is key for efficient DNA double-strand break (DSB) repair involving local transcriptional activation and repression. Here, the GATAD2B-NuRD complex is found to form a boundary between open and closed chromatin to prevent excessive DNA end resection and repair failure. The GATAD2B-NuRD complex associates with DSBs in a manner dependent on transcription and on DNA:RNA hybrids, and facilitates histone deacetylation and chromatin condensation. Lack of GATAD2B-NuRD results in chromatin hyper-relaxation and excessive DNA end resection, leading to failure in homologous recombination (HR) repair. The GATAD2B-NuRD complex acts as a key regulator in the interplay between transcription and chromatin dynamics during the RNA-dependent DNA damage response. Lack of the GATAD2B-NuRD complex results in chromatin hyper-relaxation and extended DNA end resection around DSBs, leading to failure of homologous recombination repair. [ABSTRACT FROM AUTHOR]

RNAi plays a central role in the regulation of eukaryotic genes. In Schizosaccharomyces pombe fission yeast, RNAi involves the formation of siRNA from dsRNA that acts to establish and maintain heterochromatin over centromeres, telomeres, and mating loci. We showed previously that transient heterochromatin also forms over S. pombe convergent genes (CGs). Remarkably, most RNAi genes are themselves convergent. We demonstrate here that transient heterochromatin formed by the RNAi pathway over RNAi CGs leads to their autoregulation in G1-S. Furthermore, the switching of RNAi gene orientation from convergent to tandem causes loss of their G1-S down-regulation. Surprisingly, yeast mutants with tandemized dcr1, ago1, or clr4 genes display aberrant centromeric heterochromatin, which results in abnormal cell morphology. Our results emphasize the significance of gene orientation for correct RNAi gene expression, and suggest a role for cell cycle-dependent formation of RNAi CG heterochromatin in cellular integrity. [ABSTRACT FROM AUTHOR]

Phosphorylation of the C-terminal domain (CTD) of RNA polymerase II (RNAP II) is a dynamic process that regulates transcription and coordinates it with pre-mRNA processing. We show here that Rct1, a nuclear multidomain cyclophilin from Schizosaccharomyces pombe, is encoded by an essential gene that interacts with the CTD and regulates its phosphorylation in vivo. Downregulation of Rct1 levels results in increased phosphorylation of the CTD at both Ser2 and Ser5 and in a commensurate decrease in RNAP II transcription. In contrast, overexpression of Rct1 decreases phosphorylation on both sites. The close association of Rct1 with transcriptionally active chromatin suggests a role in regulation of RNAP II transcriptional activity. These data, together with the pleiotropic phenotype upon Rct1 deregulation, suggest that this multidomain cyclophilin is an important player in maintaining the correct phosphorylation code of the CTD and thereby regulating CTD function. [ABSTRACT FROM AUTHOR]

Vascular aging is a major risk factor for age-related cardiovascular diseases, which have high rates of morbidity and mortality. It is characterized by changes in the blood vessels, such as macroscopically increased vascular diameter and intima-medial thickness, chronic inflammation, vascular calcification, arterial stiffening, and atherosclerosis. DNA damage and the subsequent various DNA damage response (DDR) pathways are important causative factors of vascular aging. Deficient DDR, which may result in the accumulation of unrepaired damaged DNA or mutations, can lead to vascular aging. On the other hand, over-activation of some DDR proteins, such as poly (ADP ribose) polymerase (PARP) and ataxia telangiectasia mutated (ATM), also can enhance the process of vascular aging, suggesting that DDR can have both positive and negative effects on vascular aging. Despite the evidence reviewed in this paper, the role of DDR in vascular aging and potential therapeutic targets remain poorly understood and require further investigation. • Vascular aging is the most important risk factor in the high morbidity and mortality of age-related cardiovascular diseases. • DNA damage and the subsequent various DNA damage response (DDR) pathways as important causative factors in vascular aging. • DDR can positively or negatively influence vascular aging. • Modulating DDR pathways pharmacologically or molecularly might have potential therapeutic benefits in managing age-related cardiovascular diseases (CVDs). [ABSTRACT FROM AUTHOR]

The endoribonuclease Dicer is a key component of the human RNA interference pathway and is known for its role in cytoplasmic microRNA production. Recent findings suggest that noncanonical Dicer generates small noncoding RNA to mediate the DNA damage response (DDR). Here, we show that human Dicer is phosphorylated in the platform-Piwi/Argonaute/Zwille-connector helix cassette (S1016) upon induction of DNA damage. Phosphorylated Dicer (p-Dicer) accumulates in the nucleus and is recruited to DNA double-strand breaks. We further demonstrate that turnover of damageinduced nuclear, double-stranded (ds) RNA requires additional phosphorylation of carboxy-terminal Dicer residues (S1728 and S1852). DNA damage-induced nuclear Dicer accumulation is conserved in mammals. Dicer depletion causes endogenous DNA damage and delays the DDR by impaired recruitment of repair factors MDC1 and 53BP1. Collectively, we place Dicer within the context of the DDR by demonstrating a DNA damage-inducible phosphoswitch that causes localized processing of nuclear dsRNA by p-Dicer to promote DNA repair. [ABSTRACT FROM AUTHOR]

Dicer is a central enzymatic player in RNA-interference pathways that acts to regulate gene expression in nearly all eukaryotes. Although the cytoplasmic function of Dicer is well documented in mammals, its nuclear function remains obscure. Here we show that Dicer is present in both the nucleus and cytoplasm, and its nuclear levels are tightly regulated. Dicer interacts with RNA polymerase II (Pol II) at actively transcribed gene loci. Loss of Dicer causes the appearance of endogenous double-stranded RNA (dsRNA), which in turn leads to induction of the interferon-response pathway and consequent cell death. Our results suggest that Pol II-associated Dicer restricts endogenous dsRNA formation from overlapping noncoding-RNA transcription units. Failure to do so has catastrophic effects on cell function. [ABSTRACT FROM AUTHOR]