Non-complementary DNA helical structure induced by positive torsional stress.

We have induced a local conformational transition by positive torsional stress in small synthetic circular DNA molecules containing cruciforms with immobile or tetramobile branched junctions. The immobile species correspond to the extruded and intruded extrema of the tetramobile junction. Under normal conditions the sequences of all the branched species prevent them from being re-absorbed into the circle. We have induced positive stress by addition of ethidium to the circle, in a low ionic strength medium. Alterations in gel electrophoretic mobility under increasing concentrations of ethidium suggest that the cruciforms undergo a transition under torsional stress. The product of this transition contains mispaired nucleotides, but interwound backbones. By comparing the electrophoretic mobilities of circles containing these structures with that of a completely complementary circle of the same length, we conclude that the twist in the mispairing region is similiar to that of completely paired species.     

The global dynamic of DNA methylation in response to heat stress revealed epigenetic mechanism of heat acclimation in Saccharina japonica

Saccharina japonica is an ecologically and economically important kelp in cold-temperate regions. When it is cultivated on a large scale in the temperate and even subtropical zones, heat stress is a frequent abiotic stress. This study is the first attempt to reveal the regulatory mechanism of the response to heat stress from the perspective of DNA methylation in S. japonica. We firstly obtained the characteristics of variation in the methylome under heat stress, and observed that heat stress caused a slight increase in the overall methylation level and methylation rate, especially in the non-coding regions of the genome. Secondly, we noted that methylation was probably one of factors affecting the expression of genes, and that methylation within the gene body was positively correlated with the gene expression (rho = 0.0784). Moreover, it was found that among the differentially expressed genes regulated by methylation, many genes were related to heat stress response, such as HSP gene family, genes of antioxidant enzymes, genes related to proteasome-ubiquitination pathway, and plant cell signaling pathways. This study demonstrated that DNA methylation is involved in regulating the response to heat stress, laying a foundation for studying the acclimation and adaptation of S. japonica to heat stress from an epigenetic perspective.     

The torsional state of DNA within the chromosome

Virtually all processes of the genome biology affect or are affected by the torsional state of DNA. Torsional energy associated with an altered twist facilitates or hinders the melting of the double helix, its molecular interactions, and its spatial folding in the form of supercoils. Yet, understanding how the torsional state of DNA is modulated remains a challenging task due to the multiplicity of cellular factors involved in the generation, transmission, and dissipation of DNA twisting forces. Here, an overview of the implication of DNA topoisomerases, DNA revolving motors, and other DNA interactions that determine local levels of torsional stress in bacterial and eukaryotic chromosomes is provided. Particular emphasis is made on the experimental approaches being developed to assess the torsional state of intracellular DNA and its organization into topological domains.     

The role of the Fanconi anemia network in the response to DNA replication stress

Fanconi anemia is a genetically heterogeneous disorder associated with chromosome instability and a highly elevated risk for developing cancer. The mutated genes encode proteins involved in the cellular response to DNA replication stress. Fanconi anemia proteins are extensively connected with DNA caretaker proteins, and appear to function as a hub for the coordination of DNA repair with DNA replication and cell cycle progression. At a molecular level, however, the raison d’être of Fanconi anemia proteins still remains largely elusive. The thirteen Fanconi anemia proteins identified to date have not been embraced into a single and defined biological process. To help put the Fanconi anemia puzzle into perspective, we begin this review with a summary of the strategies employed by prokaryotes and eukaryotes to tolerate obstacles to the progression of replication forks. We then summarize what we know about Fanconi anemia with an emphasis on biochemical aspects, and discuss how the Fanconi anemia network, a late acquisition in evolution, may function to permit the faithful and complete duplication of our very large vertebrate chromosomes.     

Plasticity of repetitive DNA in response to metal stress in Bryophytes

Abstract

The relationship between environmental stresses and the genome was investigated by examining the behaviour of repetitive DNA in response to lead or cadmium in two Bryophytes, differing from a physiological and an ecological point of view, namely the aquatic moss L. riparium and the terricolous moss F. hygrometrica. Using different experimental approaches, a direct relationship was shown to exist in these two mosses between the metal-induced stress and repetitive DNA. In fact, in both organisms, metal treatment was accompanied by a selective amplification of some GC-rich repetitive DNA sequences forming peculiar agglomerates inside the nucleus; this amplification is quantitatively proportional to the time of exposure of the plants to the metals and stops upon removal of the metal from the culture medium. Results show that ribosomal DNA sequences are involved in this metal-induced repetitive DNA agglomerate formation, although they are not the only repetitive sequences present within the heterochromatic DNA agglomerates. The plasticity of the genome of the Bryophytes in response to external stimuli, and the fact that repetitive DNA is involved in this plasticity are discussed.     

The checkpoint response to replication stress

Genome instability is a hallmark of cancer cells, and defective DNA replication, repair and recombination have been linked to its etiology. Increasing evidence suggests that proteins influencing S-phase processes such as replication fork movement and stability, repair events and replication completion, have significant roles in maintaining genome stability. DNA damage and replication stress activate a signal transduction cascade, often referred to as the checkpoint response. A central goal of the replication checkpoint is to maintain the integrity of the replication forks while facilitating replication completion and DNA repair and coordinating these events with cell cycle transitions. Progression through the cell cycle in spite of defective or incomplete DNA synthesis or unrepaired DNA lesions may result in broken chromosomes, genome aberrations, and an accumulation of mutations. In this review we discuss the multiple roles of the replication checkpoint during replication and in response to replication stress, as well as the enzymatic activities that cooperate with the checkpoint pathway to promote fork resumption and repair of DNA lesions thereby contributing to genome integrity.     

FBH1 promotes DNA double-strand breakage and apoptosis in response to DNA replication stress

Proper resolution of stalled replication forks is essential for genome stability. Purification of FBH1, a UvrD DNA helicase, identified a physical interaction with replication protein A (RPA), the major cellular single-stranded DNA (ssDNA)–binding protein complex. Compared with control cells, FBH1-depleted cells responded to replication stress with considerably fewer double-strand breaks (DSBs), a dramatic reduction in the activation of ATM and DNA-PK and phosphorylation of RPA2 and p53, and a significantly increased rate of survival. A minor decrease in ssDNA levels was also observed. All these phenotypes were rescued by wild-type FBH1, but not a FBH1 mutant lacking helicase activity. FBH1 depletion had no effect on other forms of genotoxic stress in which DSBs form by means that do not require ssDNA intermediates. In response to catastrophic genotoxic stress, apoptosis prevents the persistence and propagation of DNA lesions. Our findings show that FBH1 helicase activity is required for the efficient induction of DSBs and apoptosis specifically in response to DNA replication stress.     

DNA endonuclease- and active oxygen-associated degradation of chloroplast DNA in response to paraquat-induced oxidative stress

Activity of chloroplast-localized DNA endonuclease was observed in detached tobacco leaves that had been treated with paraquat and light The DNA endonuclease was able to cleave the chloroplast, plasmid, and single-stranded DNA, as estimated on an agarose gel. Activity was sensitive to two endonuclease inhibitors: aurintricarboxylic acid and ZnSO₄. The time course for activity showed a peak 4 h after the stress treatment These results suggest that this enzyme plays a specific physiological role during oxidative stress. Probable roles for this enzyme are also discussed.     

Autophagy-dependent senescence in response to DNA damage and chronic apoptotic stress

Autophagy regulates cell survival and cell death upon various cellular stresses, yet the molecular signaling events involved are not well defined. Here, we established the function of a proteolytic Cyclin E fragment (p18-CycE) in DNA damage-induced autophagy, apoptosis, and senescence. p18-CycE was identified in hematopoietic cells undergoing DNA damage-induced apoptosis. In epithelial cells exposed to DNA damage, chronic but not transient expression of p18-CycE leads to higher turnover of LC3 I/II and increased emergence of autophagosomes and autolysosomes. Levels of p18-CycE, which was generated by proteolytic cleavage of endogenous Cyclin E, were greatly increased by chloroquine and correlated with LC 3II conversion. Preventing p18-CycE genesis blocked conversion of LC3 I to LC3 II. Upon DNA damage, cytoplasmic ataxia-telangiectasia-mutated (ATM) was phosphorylated in p18-CycE-expressing cells resulting in sustained activation of the adenosine-mono-phosphate-dependent kinase (AMPK). These lead to sustained activation of mammalian autophagy-initiating kinase ULK1, which was abrogated upon inhibiting ATM and AMPK phosphorylation. Moreover, p18-CycE was degraded via autophagy followed by induction of senescence. Both autophagy and senescence were prevented by inhibiting autophagy, which leads to increased apoptosis in p18-CycE-expressing cells by stabilizing p18-CycE expression. Senescence was further associated with cytoplasmic co-localization and degradation of p18-CycE and Ku70. In brief, chronic p18-CycE expression-induced autophagy leads to clearance of p18-CycE following DNA damage and induction of senescence. Autophagy inhibition stabilized the cytoplasmic p18-CycE-Ku70 complex leading to apoptosis. Thus, our findings define how chronic apoptotic stress and DNA damage initiate autophagy and regulate cell survival through senescence and/or apoptosis.     

Proapoptotic Bid mediates the Atr-directed DNA damage response to replicative stress

Proapoptotic BH3 interacting domain death agonist (Bid), a BH3-only Bcl-2 family member, is situated at the interface between the DNA damage response and apoptosis, with roles in death receptor-induced apoptosis as well as cell cycle checkpoints following DNA damage.(1, 2, 3) In this study, we demonstrate that Bid functions at the level of the sensor complex in the Atm and Rad3-related (Atr)-directed DNA damage response. Bid is found with replication protein A (RPA) in nuclear foci and associates with the Atr/Atr-interacting protein (Atrip)/RPA complex following replicative stress. Furthermore, Bid-deficient cells show an impaired response to replicative stress manifest by reduced accumulation of Atr and Atrip on chromatin and at DNA damage foci, reduced recovery of DNA synthesis following replicative stress, and decreased checkpoint kinase 1 activation and RPA phosphorylation. These results establish a direct role for the BH3-only Bcl-2 family member, Bid, acting at the level of the damage sensor complex to amplify the Atr-directed cellular response to replicative DNA damage.     

The dynamic interplay in chromatin remodeling factors polycomb and trithorax proteins in response to DNA damage

The dynamic interplay in polycomb group (PcG) and trithorax group (TrxG) proteins in response to DNA damage directly involves in the DNA double strand breaks (DSBs) sites and potentially function in both homologous recombination (HR) and nonhomologous end joining (NHEJ) pathways. The process includes chromatin remodeling that is a major mechanism used by cells to relax chromatin in DNA damage response (DDR) and repair. PcGs show resistance ability to the process while, some tumor suppressor genes involves in the DDR and repair by interacting with TrxGs. Understanding how the dynamic interplay in PcGs and TrxGs impacts on DDR will shed light on the mechanisms of carcinogenesis and develop a new target from anti-DDR related drugs.