Molecular mechanisms of the repair of UV-induced DNA damages in plants

Solar UV-B radiation is an environmental factor which damage and destabilize genomes. UV-B-induced DNA lesions have cytotoxic and genotoxic effects on the cells of pro- and eucaryotes including plants. In addition, such lesions can cause gene mutations in plants. The products of the damages of cellular DNA caused by UV are examined in the present review and plant reparative pathways including photoreactivation, base excision repair and nucleotide excision repair are analyzed. The review deals as well with the mechanisms of plant DNA damage tolerance which allow to reduce the toxic effects of UV-B radiation.     

Genotoxic stress in plants: shedding light on DNA damage, repair and DNA repair helicases

Plant cells are constantly exposed to environmental agents and endogenous processes that inflict damage to DNA and cause genotoxic stress, which can reduce plant genome stability, growth and productivity. Plants are most affected by solar UV-B radiation, which damage the DNA by inducing the formation of two main UV photoproducts such as cyclobutane pyrimidine dimers (CPDs) and pyrimidine (6-4) pyrimidone photoproducts (6-4PPs). Reactive oxygen species (ROS) are also generated extra- or intra-cellularly, which constitute yet another source of genotoxic stress. As a result of this stress, the cellular DNA-damage responses (DDR) are activated, which transiently arrest the cell cycle and allow cells to repair DNA before proceeding into mitosis. DDR requires the activation of Ataxia telangiectasia-mutated (ATM) and Rad3-related (ATR) genes, which regulate the cell cycle and transmit the damage signals to downstream effectors of cell-cycle progression. Since genomic protection and stability are fundamental to ensure and sustain plant diversity and productivity, therefore, repair of DNA damages is essential. In plants the bulky DNA lesions, CPDs and 6-4PPs, are repaired by a simple and error-free mechanism: photoreactivation, which is a light-dependent mechanism and requires CPD or 6-4PP specific photolyases. In addition to this direct repair process, the plants also have sophisticated light-independent general repair mechanisms, such as the nucleotide excision repair (NER) and base excision repair (BER). The completed plant genome sequences reveal that most of the genes involved in NER and BER are present in higher plants, which suggests that the network of in-built DNA-damage repair mechanisms is conserved. This article describes the insight underlying the DNA damage and repair pathways in plants. The comet assay to measure the DNA damage and the role of DNA repair helicases such as XPD and XPB are also covered.     

Molecular mechanisms of DNA mismatch repair.

DNA mismatch repair (MMR) safeguards the integrity of the genome. In its role in postreplicative repair, this repair pathway corrects base-base and insertion/deletion (I/D) mismatches that have escaped the proofreading function of replicative polymerases. In its absence, cells assume a mutator phenotype in which the rate of spontaneous mutation is greatly elevated. The discovery that defects in mismatch repair segregate with certain cancer predisposition syndromes highlights its essential role in mutation avoidance. Recently, three-dimensional structures of MutS, a key repair protein that recognizes mismatches, have been determined by X-ray crystallography. This article provides an overview of the structural features of MutS proteins and discusses how the structural data together with biochemical and genetic studies reveal new insights into the molecular mechanisms of mismatch repair.     

Molecular mechanisms of DNA double-strand break repair

DNA double-strand breaks (DSBs) are major threats to the genomic integrity of cells. If not taken care of properly, they can cause chromosome fragmentation, loss and translocation, possibly resulting in carcinogenesis. Upon DSB formation, cell-cycle checkpoints are triggered and multiple DSB repair pathways can be activated. Recent research on the Nijmegen breakage syndrome, which predisposes patients to cancer, suggests a direct link between activation of cell-cycle checkpoints and DSB repair. Furthermore, the biochemical activities of proteins involved in the two major DSB repair pathways, homologous recombination and DNA end-joining, are now beginning to emerge. This review discusses these new findings and their implications for the mechanisms of DSB repair.     

Checkpoint mechanisms at the intersection between DNA damage and repair

In response to genomic insults cells trigger a signal transduction pathway, known as DNA damage checkpoint, whose role is to help the cell to cope with the damage by coordinating cell cycle progression, DNA replication and DNA repair mechanisms. Accumulating evidence suggests that activation of the first checkpoint kinase in the cascade is not due to the lesion itself, but it requires recognition and initial processing of the lesion by a specific repair mechanism. Repair enzymes likely convert a variety of physically and chemically different lesions to a unique common structure, a ssDNA region, which is the checkpoint triggering signal. Checkpoint kinases can modify the activity of repair mechanisms, allowing for efficient repair, on one side, and modulating the generation of the ssDNA signal, on the other. This strategy may be important to allow the most effective repair and a prompt recovery from the damage condition. Interestingly, at least in some cases, if the damage level is low enough the cell can deal with the lesions and it does not need to activate the checkpoint response. On the other hand if damage level is high or if the lesions are not rapidly repairable, checkpoint mechanisms become important for cell survival and preservation of genome integrity.     

Dynamics of heat-shock induced DNA damage and repair in senescent tobacco plants

Oxidative stress plays an important role in plant ageing and in response to different stresses. Oxidative DNA damage, unless repaired, may have detrimental consequences and increase genetic instability. Therefore, we determined the role of heat-shock induced oxidative stress on induction and repair of DNA damage in relation to oxidative stress tolerance in senescent tobacco plants. One-month-old (young) and three-month-old (senescent) plants were exposed to 42 °C for 2 and 4 h and left to recover at 26 °C for 24 and 72 h. The progression of senescence was characterized by the lower soluble protein and malondialdehyde content compared to young plants. Immediately after the heat shock, an increase in lipid peroxidation and guaiacol peroxidase activity, as well as DNA damage measured by the Comet assay were induced to higher extent in the young plants than in the senescent ones compared to their respective controls. Moreover, after 24-h recovery, the DNA damage further increased in the young plants whereas tendency of DNA repair was observed in the senescent plants. Upon 72-h recovery, no significant differences were noticed in all parameters studied (regardless of plant age) compared to the controls. The random amplified polymorphic DNA (RAPD) analysis confirmed genetic stability of the tobacco plants during the heat-shock exposures as well as the subsequent recovery periods.     

PNKP在DNA损伤修复中的作用

多聚核苷酸激酶/磷酸酶(polynucleotide kinase/phosphatase,PNKP)是一种DNA末端修复酶,同时具有激酶和磷酸酶活性,在DNA单链断裂修复途径、碱基切除修复途径以及DNA双链断裂修复中的非同源末端连接途径中发挥着至关重要的作用。近年来,由于一种与PNKP相关的常染色体隐性遗传病——MCSZ综合征的发现,使得人们对PNKP的关注度进一步增加。笔者从与PNKP相互作用的X射线交叉互补修复基因1(X-ray repair cross-complementing group 1,XRCC1)、X射线交叉互补修复基因4(X-ray repair cross-complementing group 4,XRCC4)和毛细血管扩张性共济失调突变基因(ataxia-telangiectasia mutated,ATM)入手,对PNKP在DNA损伤修复中的作用进行概述。     

DNA repair mechanisms in plants: crucial sensors and effectors for the maintenance of genome integrity

As obligate phototrophs, plants harness energy from sunlight to split water, producing oxygen and reducing power. This lifestyle exposes plants to particularly high levels of genotoxic stress that threatens genomic integrity, leading to mutation, developmental arrest and cell death. Plants, which with algae are the only photosynthetic eukaryotes, have evolved very effective pathways for DNA damage signalling and repair, and this review summarises our current understanding of these processes in the responses of plants to genotoxic stress. We also identify how the use of new and emerging technologies can complement established physiological and ecological studies to progress the application of this knowledge in biotechnology.     

DNA repair in higher plants

Numerous studies have demonstrated a requirement in plants for repair of DNA damage arising from either intrinsic or extrinsic sources. Investigations also have revealed a capacity for repair types of DNA damage, and conversely, identified mutants apparently defective in such repair. This article provides a concise overview of nuclear DNA repair mechanisms in higher plants, particularly those processes concerned with the repair of UV-induced lesions, and includes surveys of UV-sensitive mutants and genes implicated in DNA repair.     

Cohesin and DNA damage repair

Replicated DNA molecules are physically connected by cohesin complexes from the time of their synthesis in S-phase until they are segregated during anaphase of the subsequent mitosis or meiosis. This sister chromatid cohesion is essential for the biorientation of chromosomes on the mitotic or meiotic spindle. In addition, cohesion is also essential during G2-phase of the cell cycle to allow repair of DNA double-strand breaks by homologous recombination. Although cohesion can normally only be established during S-phase, recent work in yeast has shown that DNA double-strand breaks induce the recruitment of cohesin to the damage site and lead to the de novo formation of cohesion at this site. It is unknown if similar mechanisms operate in higher eukaryotes, but in mammalian cells phosphorylation of the cohesin subunit Smc1 by the protein kinase Atm has been shown to be important for DNA repair. We discuss how cohesin and sister chromatid cohesion might facilitate the repair of damaged DNA.     

Molecular mechanisms of urea transport in plants

Urea is a soil nitrogen form available to plant roots and a secondary nitrogen metabolite liberated in plant cells. Based on growth complementation of yeast mutants and “in-silico analysis”, two plant families have been identified and partially characterized that mediate membrane transport of urea in heterologous expression systems. AtDUR3 is a single Arabidopsis gene belonging to the sodium solute symporter family that cotransports urea with protons at high affinity, while members of the tonoplast intrinsic protein (TIP) subfamily of aquaporins transport urea in a channel-like manner. The following review summarizes current knowledge on the membrane localization, energetization and regulation of these two types of urea transporters and discusses their possible physiological roles in planta.     

Molecular mechanisms of phosphate transport in plants

Membrane-spanning transport proteins are responsible for the selective passage of most mineral nutrients and metabolites across cellular and intracellular membranes. This review's focus is on summarising the current state of research covering the molecular regulation and biochemical mechanisms involved in the transport of phosphorus, an often growth-limiting nutrient, in vascular plants. Physiological data illustrating the tight control of Pi homeostasis on the cellular as well as on the organism's level are discussed together with the recent results on molecular transport mechanisms.     

Having a direct look: Analysis of DNA damage and repair mechanisms by next generation sequencing

Genetic information is under constant attack from endogenous and exogenous sources, and the use of model organisms has provided important frameworks to understand how genome stability is maintained and how various DNA lesions are repaired. The advance of high throughput next generation sequencing (NGS) provides new inroads for investigating mechanisms needed for genome maintenance. These emerging studies, which aim to link genetic toxicology and mechanistic analyses of DNA repair processes in vivo, rely on defining mutational signatures caused by faulty replication, endogenous DNA damaging metabolites, or exogenously applied genotoxins; the analysis of their nature, their frequency and distribution. In contrast to classical studies, where DNA repair deficiency is assessed by reduced cellular survival, the localization of DNA repair factors and their interdependence as well as limited analysis of single locus reporter assays, NGS based approaches reveal the direct, quantal imprint of mutagenesis genome-wide, at the DNA sequence level. As we will show, such investigations require the analysis of DNA derived from single genotoxin treated cells, or DNA from cell populations regularly passaged through single cell bottlenecks when naturally occurring mutation accumulation is investigated. We will argue that the life cycle of the nematode Caenorhabditis elegans, its genetic malleability combined with whole genome sequencing provides an exciting model system to conduct such analysis.