The Emerging Roles of ATP-Dependent Chromatin Remodeling Enzymes in Nucleotide Excision Repair

DNA repair in eukaryotic cells takes place in the context of chromatin, where DNA, including damaged DNA, is tightly packed into nucleosomes and higher order chromatin structures. Chromatin intrinsically restricts accessibility of DNA repair proteins to the damaged DNA and impacts upon the overall rate of DNA repair. Chromatin is highly responsive to DNA damage and undergoes specific remodeling to facilitate DNA repair. How damaged DNA is accessed, repaired and restored to the original chromatin state, and how chromatin remodeling coordinates these processes in vivo, remains largely unknown. ATP-dependent chromatin remodelers (ACRs) are the master regulators of chromatin structure and dynamics. Conserved from yeast to humans, ACRs utilize the energy of ATP to reorganize packing of chromatin and control DNA accessibility by sliding, ejecting or restructuring nucleosomes. Several studies have demonstrated that ATP-dependent remodeling activity of ACRs plays important roles in coordination of spatio-temporal steps of different DNA repair pathways in chromatin. This review focuses on the role of ACRs in regulation of various aspects of nucleotide excision repair (NER) in the context of chromatin. We discuss current understanding of ATP-dependent chromatin remodeling by various subfamilies of remodelers and regulation of the NER pathway in vivo.     

Epigenetic Regulation of microRNAs in Cancer: Shortening the Distance from Bench to Bedside

Cancer is a complex disease involving alterations of multiple processes, with both genetic and epigenetic features contributing as core factors to the disease. In recent years, it has become evident that non-coding RNAs (ncRNAs), an epigenetic factor, play a key role in the initiation and progression of cancer. MicroRNAs, the most studied non-coding RNAs subtype, are key controllers in a myriad of cellular processes, including proliferation, differentiation, and apoptosis. Furthermore, the expression of miRNAs is controlled, concomitantly, by other epigenetic factors, such as DNA methylation and histone modifications, resulting in aberrant patterns of expression upon the occurrence of cancer. In this sense, aberrant miRNA landscape evaluation has emerged as a promising strategy for cancer management. In this review, we have focused on the regulation (biogenesis, processing, and dysregulation) of miRNAs and their role as modulators of the epigenetic machinery. We have also highlighted their potential clinical value, such as validated diagnostic and prognostic biomarkers, and their relevant role as chromatin modifiers in cancer therapy.     

Role and Regulation of Pif1 Family Helicases at the Replication Fork

Pif1 helicases are a multifunctional family of DNA helicases that are important for many aspects of genomic stability in the nucleus and mitochondria. Pif1 helicases are conserved from bacteria to humans. Pif1 helicases play multiple roles at the replication fork, including promoting replication through many barriers such as G-quadruplex DNA, the rDNA replication fork barrier, tRNA genes, and R-loops. Pif1 helicases also regulate telomerase and promote replication termination, Okazaki fragment maturation, and break-induced replication. This review highlights many of the roles and regulations of Pif1 at the replication fork that promote cellular health and viability.     

RIF1 Links Replication Timing with Fork Reactivation and DNA Double-Strand Break Repair

Replication timing (RT) is a cellular program to coordinate initiation of DNA replication in all origins within the genome. RIF1 (replication timing regulatory factor 1) is a master regulator of RT in human cells. This role of RIF1 is associated with binding G4-quadruplexes and changes in 3D chromatin that may suppress origin activation over a long distance. Many effects of RIF1 in fork reactivation and DNA double-strand (DSB) repair (DSBR) are underlined by its interaction with TP53BP1 (tumor protein p53 binding protein). In G1, RIF1 acts antagonistically to BRCA1 (BRCA1 DNA repair associated), suppressing end resection and homologous recombination repair (HRR) and promoting non-homologous end joining (NHEJ), contributing to DSBR pathway choice. RIF1 is an important element of intra-S-checkpoints to recover damaged replication fork with the involvement of HRR. High-resolution microscopic studies show that RIF1 cooperates with TP53BP1 to preserve 3D structure and epigenetic markers of genomic loci disrupted by DSBs. Apart from TP53BP1, RIF1 interact with many other proteins, including proteins involved in DNA damage response, cell cycle regulation, and chromatin remodeling. As impaired RT, DSBR and fork reactivation are associated with genomic instability, a hallmark of malignant transformation, RIF1 has a diagnostic, prognostic, and therapeutic potential in cancer. Further studies may reveal other aspects of common regulation of RT, DSBR, and fork reactivation by RIF1.     

Plant DNA Methylation: An Epigenetic Mark in Development,Environmental Interactions,and Evolution

DNA methylation is an epigenetic modification of the genome involved in the regulation of gene expression and modulation of chromatin structure. Plant genomes are widely methylated, and the methylation generally occurs on the cytosine bases through the activity of specific enzymes called DNA methyltransferases. On the other hand, methylated DNA can also undergo demethylation through the action of demethylases. The methylation landscape is finely tuned and assumes a pivotal role in plant development and evolution. This review illustrates different molecular aspects of DNA methylation and some plant physiological processes influenced by this epigenetic modification in model species, crops, and ornamental plants such as orchids. In addition, this review aims to describe the relationship between the changes in plant DNA methylation levels and the response to biotic and abiotic stress. Finally, we discuss the possible evolutionary implications and biotechnological applications of DNA methylation.     

Chromatin Profiling Techniques: Exploring the Chromatin Environment and Its Contributions to Complex Traits

The genetic architecture of complex traits is multifactorial. Genome-wide association studies (GWASs) have identified risk loci for complex traits and diseases that are disproportionately located at the non-coding regions of the genome. On the other hand, we have just begun to understand the regulatory roles of the non-coding genome, making it challenging to precisely interpret the functions of non-coding variants associated with complex diseases. Additionally, the epigenome plays an active role in mediating cellular responses to fluctuations of sensory or environmental stimuli. However, it remains unclear how exactly non-coding elements associate with epigenetic modifications to regulate gene expression changes and mediate phenotypic outcomes. Therefore, finer interrogations of the human epigenomic landscape in associating with non-coding variants are warranted. Recently, chromatin-profiling techniques have vastly improved our understanding of the numerous functions mediated by the epigenome and DNA structure. Here, we review various chromatin-profiling techniques, such as assays of chromatin accessibility, nucleosome distribution, histone modifications, and chromatin topology, and discuss their applications in unraveling the brain epigenome and etiology of complex traits at tissue homogenate and single-cell resolution. These techniques have elucidated compositional and structural organizing principles of the chromatin environment. Taken together, we believe that high-resolution epigenomic and DNA structure profiling will be one of the best ways to elucidate how non-coding genetic variations impact complex diseases, ultimately allowing us to pinpoint cell-type targets with therapeutic potential.     

A Fork Trap in the Chromosomal Termination Area Is Highly Conserved across All Escherichia coli Phylogenetic Groups

Termination of DNA replication, the final stage of genome duplication, is surprisingly complex, and failures to bring DNA synthesis to an accurate conclusion can impact genome stability and cell viability. In Escherichia coli, termination takes place in a specialised termination area opposite the origin. A ‘replication fork trap’ is formed by unidirectional fork barriers via the binding of Tus protein to genomic ter sites. Such a fork trap system is found in some bacterial species, but it appears not to be a general feature of bacterial chromosomes. The biochemical properties of fork trap systems have been extensively characterised, but little is known about their precise physiological roles. In this study, we compare locations and distributions of ter terminator sites in E. coli genomes across all phylogenetic groups, including Shigella. Our analysis shows that all ter sites are highly conserved in E. coli, with slightly more variability in the Shigella genomes. Our sequence analysis of ter sites and Tus proteins shows that the fork trap is likely to be active in all strains investigated. In addition, our analysis shows that the dif chromosome dimer resolution site is consistently located between the innermost ter sites, even if rearrangements have changed the location of the innermost termination area. Our data further support the idea that the replication fork trap has an important physiological role that provides an evolutionary advantage.     

Functional Coupling between DNA Replication and Sister Chromatid Cohesion Establishment

Several lines of evidence suggest the existence in the eukaryotic cells of a tight, yet largely unexplored, connection between DNA replication and sister chromatid cohesion. Tethering of newly duplicated chromatids is mediated by cohesin, an evolutionarily conserved hetero-tetrameric protein complex that has a ring-like structure and is believed to encircle DNA. Cohesin is loaded onto chromatin in telophase/G1 and converted into a cohesive state during the subsequent S phase, a process known as cohesion establishment. Many studies have revealed that down-regulation of a number of DNA replication factors gives rise to chromosomal cohesion defects, suggesting that they play critical roles in cohesion establishment. Conversely, loss of cohesin subunits (and/or regulators) has been found to alter DNA replication fork dynamics. A critical step of the cohesion establishment process consists in cohesin acetylation, a modification accomplished by dedicated acetyltransferases that operate at the replication forks. Defects in cohesion establishment give rise to chromosome mis-segregation and aneuploidy, phenotypes frequently observed in pre-cancerous and cancerous cells. Herein, we will review our present knowledge of the molecular mechanisms underlying the functional link between DNA replication and cohesion establishment, a phenomenon that is unique to the eukaryotic organisms.     

Theranostic Interpolation of Genomic Instability in Breast Cancer

Breast cancer is a diverse disease caused by mutations in multiple genes accompanying epigenetic aberrations of hazardous genes and protein pathways, which distress tumor-suppressor genes and the expression of oncogenes. Alteration in any of the several physiological mechanisms such as cell cycle checkpoints, DNA repair machinery, mitotic checkpoints, and telomere maintenance results in genomic instability. Theranostic has the potential to foretell and estimate therapy response, contributing a valuable opportunity to modify the ongoing treatments and has developed new treatment strategies in a personalized manner. “Omics” technologies play a key role while studying genomic instability in breast cancer, and broadly include various aspects of proteomics, genomics, metabolomics, and tumor grading. Certain computational techniques have been designed to facilitate the early diagnosis of cancer and predict disease-specific therapies, which can produce many effective results. Several diverse tools are used to investigate genomic instability and underlying mechanisms. The current review aimed to explore the genomic landscape, tumor heterogeneity, and possible mechanisms of genomic instability involved in initiating breast cancer. We also discuss the implications of computational biology regarding mutational and pathway analyses, identification of prognostic markers, and the development of strategies for precision medicine. We also review different technologies required for the investigation of genomic instability in breast cancer cells, including recent therapeutic and preventive advances in breast cancer.     

DNA Polymerase-Parental DNA Interaction Is Essential for Helicase-Polymerase Coupling during Bacteriophage T7 DNA Replication

DNA helicase and polymerase work cooperatively at the replication fork to perform leading-strand DNA synthesis. It was believed that the helicase migrates to the forefront of the replication fork where it unwinds the duplex to provide templates for DNA polymerases. However, the molecular basis of the helicase-polymerase coupling is not fully understood. The recently elucidated T7 replisome structure suggests that the helicase and polymerase sandwich parental DNA and each enzyme pulls a daughter strand in opposite directions. Interestingly, the T7 polymerase, but not the helicase, carries the parental DNA with a positively charged cleft and stacks at the fork opening using a β-hairpin loop. Here, we created and characterized T7 polymerases each with a perturbed β-hairpin loop and positively charged cleft. Mutations on both structural elements significantly reduced the strand-displacement synthesis by T7 polymerase but had only a minor effect on DNA synthesis performed against a linear DNA substrate. Moreover, the aforementioned mutations eliminated synergistic helicase-polymerase binding and unwinding at the DNA fork and processive fork progressions. Thus, our data suggested that T7 polymerase plays a dominant role in helicase-polymerase coupling and replisome progression.