Checkpoint kinase 1 in DNA damage response and cell cycle regulation

Originally identified as a mediator of DNA damage response (DDR), checkpoint kinase 1 (Chk1) has a broader role in checkpoint activation in DDR and normal cell cycle regulation. Chk1 activation involves phosphorylation at conserved sites. However, recent work has identified a splice variant of Chk1, which may regulate Chk1 in both DDR and normal cell cycle via molecular interaction. Upon activation, Chk1 phosphorylates a variety of substrate proteins, resulting in the activation of DNA damage checkpoints, cell cycle arrest, DNA repair, and/or cell death. Chk1 and its related signaling may be an effective therapeutic target in diseases such as cancer.     

Eukaryotic DNA damage checkpoint activation in response to double-strand breaks

Double-strand breaks (DSBs) are the most detrimental form of DNA damage. Failure to repair these cytotoxic lesions can result in genome rearrangements conducive to the development of many diseases, including cancer. The DNA damage response (DDR) ensures the rapid detection and repair of DSBs in order to maintain genome integrity. Central to the DDR are the DNA damage checkpoints. When activated by DNA damage, these sophisticated surveillance mechanisms induce transient cell cycle arrests, allowing sufficient time for DNA repair. Since the term “checkpoint” was coined over 20 years ago, our understanding of the molecular mechanisms governing the DNA damage checkpoint has advanced significantly. These pathways are highly conserved from yeast to humans. Thus, significant findings in yeast may be extrapolated to vertebrates, greatly facilitating the molecular dissection of these complex regulatory networks. This review focuses on the cellular response to DSBs in Saccharomyces cerevisiae, providing a comprehensive overview of how these signalling pathways function to orchestrate the cellular response to DNA damage and preserve genome stability in eukaryotic cells.     

Implication of the VRK1 chromatin kinase in the signaling responses to DNA damage: a therapeutic target?

DNA damage causes a local distortion of chromatin that triggers the sequential processes that participate in specific DNA repair mechanisms. This initiation of the repair response requires the involvement of a protein whose activity can be regulated by histones. Kinases are candidates to regulate and coordinate the connection between a locally altered chromatin and the response initiating signals that lead to identification of the type of lesion and the sequential steps required in specific DNA damage responses (DDR). This initiating kinase must be located in chromatin, and be activated independently of the type of DNA damage. We review the contribution of the Ser-Thr vaccinia-related kinase 1 (VRK1) chromatin kinase as a new player in the signaling of DNA damage responses, at chromatin and cellular levels, and its potential as a new therapeutic target in oncology. VRK1 is involved in the regulation of histone modifications, such as histone phosphorylation and acetylation, and in the formation of γH2AX, NBS1 and 53BP1 foci induced in DDR. Induction of DNA damage by chemotherapy or radiation is a mainstay of cancer treatment. Therefore, novel treatments can be targeted to proteins implicated in the regulation of DDR, rather than by directly causing DNA damage.     

Influenza infection induces host DNA damage and dynamic DNA damage responses during tissue regeneration

Influenza viruses account for significant morbidity worldwide. Inflammatory responses, including excessive generation of reactive oxygen and nitrogen species (RONS), mediate lung injury in severe influenza infections. However, the molecular basis of inflammation-induced lung damage is not fully understood. Here, we studied influenza H1N1 infected cells in vitro, as well as H1N1 infected mice, and we monitored molecular and cellular responses over the course of 2 weeks in vivo. We show that influenza induces DNA damage to both, when cells are directly exposed to virus in vitro (measured using the comet assay) and also when cells are exposed to virus in vivo (estimated via γH2AX foci). We show that DNA damage, as well as responses to DNA damage persist in vivo until long after virus has been cleared, at times when there are inflammation associated RONS (measured by xanthine oxidase activity and oxidative products). The frequency of lung epithelial and immune cells with increased γH2AX foci is elevated in vivo, especially for dividing cells (Ki-67-positive) exposed to oxidative stress during tissue regeneration. Additionally, we observed a significant increase in apoptotic cells as well as increased levels of DNA double strand break (DSB) repair proteins Ku70, Ku86 and Rad51 during the regenerative phase. In conclusion, results show that influenza induces DNA damage both in vitro and in vivo, and that DNA damage responses are activated, raising the possibility that DNA repair capacity may be a determining factor for tissue recovery and disease outcome.     

Sex chromosome inactivation in germ cells: emerging roles of DNA damage response pathways

Sex chromosome inactivation in male germ cells is a paradigm of epigenetic programming during sexual reproduction. Recent progress has revealed the underlying mechanisms of sex chromosome inactivation in male meiosis. The trigger of chromosome-wide silencing is activation of the DNA damage response (DDR) pathway, which is centered on the mediator of DNA damage checkpoint 1 (MDC1), a binding partner of phosphorylated histone H2AX (γH2AX). This DDR pathway shares features with the somatic DDR pathway recognizing DNA replication stress in the S phase. Additionally, it is likely to be distinct from the DDR pathway that recognizes meiosis-specific double-strand breaks. This review article extensively discusses the underlying mechanism of sex chromosome inactivation.     

The nuclear γ-H2AX apoptotic ring: implications for cancers and autoimmune diseases

Apoptosis is a fundamental process for metazoan development. It is also relevant to the pathophysiology of immune diseases and cancers and to the outcome of cancer chemotherapies, as well as being a target for cancer therapies. Apoptosis involves intrinsic pathways typically initiated by DNA damaging agents and engaging mitochondria, and extrinsic pathways typically initiated by “death receptors” and their ligands TRAIL and TNF at the cell surface. Recently, we discovered the apoptotic ring, which microscopically looks like a nuclear annular staining early in apoptosis. This ring is, in three-dimensional space, a thick intranuclear shell consisting of epigenetic modifications including histone H2AX and DNA damage response (DDR) proteins. It excludes the DNA repair factors usually associated with γ-H2AX in the DDR nuclear foci. Here, we summarize our knowledge of the apoptotic ring, and discuss its biological and pathophysiological relevance, as well as its value as a potential pharmacodynamic biomarker for anticancer therapies.     

DNA damage repair and transcription

Double-strand breaks arise frequently in the course of endogenous - normal and pathological - cellular DNA metabolism or can result from exogenous agents such as ionizing radiation. It is generally accepted that these lesions represent one of the most severe types of DNA damage with respect to preservation of genomic integrity. Therefore, cells have evolved complex mechanisms that include cell-cycle arrest, activation of various genes, including those associated with DNA repair, and in certain cases induction of the apoptotic pathway to respond to double-strand breaks. In this review we discuss recent progress in our understanding of cellular responses to DNA double-strand breaks. In addition to an analysis of the current paradigms of detection, signaling and repair, insights into the significance of chromatin remodeling in the double-strand break-response pathways are provided.     

Protecting DNA from errors and damage: an overview of DNA repair mechanisms in plants compared to mammals

The genome integrity of all organisms is constantly threatened by replication errors and DNA damage arising from endogenous and exogenous sources. Such base pair anomalies must be accurately repaired to prevent mutagenesis and/or lethality. Thus, it is not surprising that cells have evolved multiple and partially overlapping DNA repair pathways to correct specific types of DNA errors and lesions. Great progress in unraveling these repair mechanisms at the molecular level has been made by several talented researchers, among them Tomas Lindahl, Aziz Sancar, and Paul Modrich, all three Nobel laureates in Chemistry for 2015. Much of this knowledge comes from studies performed in bacteria, yeast, and mammals and has impacted research in plant systems. Two plant features should be mentioned. Plants differ from higher eukaryotes in that they lack a reserve germline and cannot avoid environmental stresses. Therefore, plants have evolved different strategies to sustain genome fidelity through generations and continuous exposure to genotoxic stresses. These strategies include the presence of unique or multiple paralogous genes with partially overlapping DNA repair activities. Yet, in spite (or because) of these differences, plants, especially Arabidopsis thaliana, can be used as a model organism for functional studies. Some advantages of this model system are worth mentioning: short life cycle, availability of both homozygous and heterozygous lines for many genes, plant transformation techniques, tissue culture methods and reporter systems for gene expression and function studies. Here, I provide a current understanding of DNA repair genes in plants, with a special focus on A. thaliana. It is expected that this review will be a valuable resource for future functional studies in the DNA repair field, both in plants and animals.     

Mechanisms regulating immune surveillance of cellular stress in cancer

The purpose of this review is to explore immune-mediated mechanisms of stress surveillance in cancer, with particular emphasis on the idea that all cancers have classical hallmarks (Hanahan and Weinberg in Cell 100:57–70, 67; Cell 144:646–674, 68) that could be interrelated. We postulate that hallmarks of cancer associated with cellular stress pathways (Luo et al. in Cell 136:823–837, 101) including oxidative stress, proteotoxic stress, mitotic stress, DNA damage, and metabolic stress could define and modulate the inflammatory component of cancer. As such, the overarching goal of this review is to define the types of cellular stress that cancer cells undergo, and then to explore mechanisms by which immune cells recognize, respond to, and are affected by each stress response.     

Are common fragile sites merely structural domains or highly organized “functional” units susceptible to oncogenic stress?

Common fragile sites (CFSs) are regions of the genome with a predisposition to DNA double-strand breaks in response to intrinsic (oncogenic) or extrinsic replication stress. CFS breakage is a common feature in carcinogenesis from its earliest stages. Given that a number of oncogenes and tumor suppressors are located within CFSs, a question that emerges is whether fragility in these regions is only a structural “passive” incident or an event with a profound biological effect. Furthermore, there is sparse evidence that other elements, like non-coding RNAs, are positioned with them. By analyzing data from various libraries, like miRbase and ENCODE, we show a prevalence of various cancer-related genes, miRNAs, and regulatory binding sites, such as CTCF within CFSs. We propose that CFSs are not only susceptible structural domains, but highly organized “functional” entities that when targeted, severe repercussion for cell homeostasis occurs.     

ATM protein kinase: the linchpin of cellular defenses to stress

ATM is the most significant molecule involved in monitoring the genomic integrity of the cell. Any damage done to DNA relentlessly challenges the cellular machinery involved in recognition, processing and repair of these insults. ATM kinase is activated early to detect and signal lesions in DNA, arrest the cell cycle, establish DNA repair signaling and faithfully restore the damaged chromatin. ATM activation plays an important role as a barrier to tumorigenesis, metabolic syndrome and neurodegeneration. Therefore, studies of ATM-dependent DNA damage signaling pathways hold promise for treatment of a variety of debilitating diseases through the development of new therapeutics capable of modulating cellular responses to stress. In this review, we have tried to untangle the complex web of ATM signaling pathways with the purpose of pinpointing multiple roles of ATM underlying the complex phenotypes observed in AT patients.     

Base excision DNA repair

DNA repair is a collection of several multienzyme, multistep processes keeping the cellular genome intact against genotoxic insults. One of these processes is base excision repair, which deals with the most ubiquitous lesions in DNA: oxidative base damage, alkylation, deamination, sites of base loss and single-strand breaks, etc. Individual enzymes acting in base excision repair have been identified. The recent years were marked with many advances in understanding of their structure and many interactions that make base excision repair a functional, versatile system. This review describes the current knowledge of structural biology and biochemistry of individual steps of base excision repair, several subpathways of the common base excision repair pathway, and interactions of the repair process with other cellular processes.     

The impact of HIV-1 on neurogenesis: implications for HAND

HIV-1 infection, in addition to its destructive effects on the immune system, plays a role in the development of neurocognitive deficits. Indeed up to 50 % of long-term HIV infected patients suffer from HIV-associated neurocognitive disorders (HAND). These deficits have been well characterized and defined clinically according to a number of cognitive parameters. HAND is often accompanied by atrophy of the brain including inhibition of neurogenesis, especially in the hippocampus.  Many mechanisms have been proposed as contributing factors to HAND including induction of oxidative stress in the central nervous system (CNS), chronic microglial-mediated neuroinflammation, amyloid-beta (Aβ) deposition, hyperphosphorylated tau protein, and toxic effects of combination antiretroviral therapy (cART). In these review we focus solely on recent experimental evidence suggesting that disturbance by HIV-1 results in impairment of neurogenesis as one contributing factor to HAND. Impaired neurogenesis has been linked to cognitive deficits and other neurodegenerative disorders. This article will highlight recently identified pathological mechanisms which potentially contribute to the development of impaired neurogenesis by HIV-1 or HIV-1-associated proteins from both animal and human studies.     

EGFR-dependent mechanisms in glioblastoma: towards a better therapeutic strategy

Glioblastoma is a particularly resilient cancer, and while therapies may be able to reach the brain by crossing the blood–brain barrier, they then have to deal with a highly invasive tumor that is very resistant to DNA damage. It seems clear that in order to kill aggressive glioma cells more efficiently and with fewer side effects on normal tissue, there must be a shift from classical cytotoxic chemotherapy to more targeted therapies. Since the epidermal growth factor receptor (EGFR) is altered in almost 50 % of glioblastomas, it currently represents one of the most promising therapeutic targets. In fact, it has been associated with several distinct steps in tumorigenesis, from tumor initiation to tumor growth and survival, and also with the regulation of cell migration and angiogenesis. However, inhibitors of the EGFR kinase have produced poor results with this type of cancer in clinical trials, with no clear explanation for the tumor resistance observed. Here we will review what we know about the expression and function of EGFR in cancer and in particular in gliomas. We will also evaluate which are the possible molecular and cellular escape mechanisms. As a result, we hope that this review will help improve the design of future EGFR-targeted therapies for glioblastomas.