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Results for “Genome Maintenance”

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At least 109 records · Page 6Linked to original sources
PubMed2000

Implication of the G2 checkpoint in the maintenance of genome integrity.

Checkpoints are surveillance mechanisms that block cell cycle transitions, for instance in response to DNA damage. We summarise here recent progress in the molecular characterisation of the G2 checkpoint which controls the entry into mitosis, and review new evidence which implicates deregulated expression of checkpoint proteins and proteins involved in DNA damage repair in cancer development. There now exists good evidence that individuals who have inherited mutations in genes involved in G2 checkpoint and DNA damage repair are predisposed to the development of various types of cancer, their cells having a strong tendency to accumulate additional mutations. However, the occurrence of mutations of most of these genes in sporadic tumors has yet to be analysed more accurately.

Animals↗
PubMed2003

ATM and related protein kinases: safeguarding genome integrity.

Maintenance of genome stability is essential for avoiding the passage to neoplasia. The DNA-damage response--a cornerstone of genome stability--occurs by a swift transduction of the DNA-damage signal to many cellular pathways. A prime example is the cellular response to DNA double-strand breaks, which activate the ATM protein kinase that, in turn, modulates numerous signalling pathways. ATM mutations lead to the cancer-predisposing genetic disorder ataxia-telangiectasia (A-T). Understanding ATM's mode of action provides new insights into the association between defective responses to DNA damage and cancer, and brings us closer to resolving the issue of cancer predisposition in some A-T carriers.

Animals↗
PubMed2005

Reconstitution of papillomavirus E2-mediated plasmid maintenance in Saccharomyces cerevisiae by the Brd4 bromodomain protein.

The papillomavirus E2 protein functions in viral transcriptional regulation, DNA replication, and episomal genome maintenance. Viral genomes are maintained in dividing cells by attachment to mitotic chromosomes by means of the E2 protein. To investigate the chromosomal tethering function of E2, plasmid stability assays were developed in Saccharomyces cerevisiae to determine whether the E2 protein could maintain plasmids containing the yeast autonomous replication sequence replication element but with the centromeric element replaced by E2-binding sites. E2 expression was not sufficient to maintain such plasmids, but plasmid stability could be rescued by expression of the mammalian protein Brd4. In the presence of both Brd4 and E2 proteins, plasmids with multiple E2-binding sites were stable without selection. S. cerevisiae encodes a homolog of Brd4 named Bdf1 that does not contain the C-terminal domain that interacts with the E2 protein. A fusion protein of Bdf1 and the Brd4 C-terminal "tail" could support E2-mediated plasmid maintenance in yeast. Using a panel of mutated E2 proteins, we determined that plasmid stability required the ability of E2 to bind DNA and to interact with Brd4 and mammalian mitotic chromosomes but did not require its replication initiation and transactivation functions. The S. cerevisiae-based plasmid maintenance assays described here are invaluable tools for dissecting mechanisms of episomal viral genome replication and screening for additional host protein factors involved in plasmid maintenance.

Binding Sites↗
PubMed2003

Telomere maintenance in fission yeast requires an Est1 ortholog.

Telomerase regulation is critical to genome maintenance yet remains poorly understood. Without telomerase's ability to synthesize telomere repeats, chromosome ends shorten progressively, as conventional DNA polymerases cannot fully replicate the ends of linear molecules. In Saccharomyces cerevisiae, telomerase activity in vivo absolutely depends on a set of telomerase accessory proteins that includes Est1p, which appears to recruit or activate telomerase at the site of polymerization. Thus, est1Delta cells have the same cellular senescence phenotype as cells lacking either the catalytic protein subunit of telomerase or its template-containing RNA subunit. While the telomerase protein is highly conserved among eukaryotes, the apparent lack of Est1p homologs has frustrated efforts to describe a common mechanism of telomerase recruitment and activation. Here, we describe SpEst1p, a homolog of Est1p from the evolutionarily distant Schizosaccharomyces pombe. Like ScEst1p, SpEst1p is required for telomerase activity in vivo. Coupled with the identification of an orthologous Est1 protein in humans [10], this suggests a much wider conservation of telomerase regulation than was previously known. Strikingly, in cells with compromised telomere function (taz1Delta), SpEst1p loss confers a lethal germination phenotype, while telomerase loss does not, indicating that SpEst1p plays an unexpected additional role in chromosome end protection.

Animals↗
PubMed1997

Genomic deletion of an imprint maintenance element abolishes imprinting of both insulin-like growth factor II and H19.

Insulin-like growth factor II (Igf2) is maternally imprinted in normal tissues with only the paternal copy of the gene being transcribed, whereas the contiguous gene H19 is paternally imprinted. Dysregulation of IGF2 imprinting is commonly observed in Wilms' tumor and other human tumors. Previous work comparing promoter-specific imprinting of human and mouse Igf2 suggested the presence of a cis element upstream of Igf2 that regulates or maintains the imprinting of three downstream promoters. To explore the molecular mechanism of maintenance of genomic imprinting, we targeted the region between insulin 2 and Igf2, where the cis imprint maintenance element (IME) resides in mouse fibroblasts. In those clones in which the targeting vector was randomly integrated into the genome, mouse Igf2 remained imprinted. However, when the targeted region containing the IME was deleted by homologous recombination, whether from the paternal or maternal allele, activation of the imprinted maternal allele of Igf2 was observed. In addition, there was a loss of H19 imprinting when the IME was deleted. The requirement of IME from both parental alleles for the maintenance of genomic imprinting thus suggests the importance of a spatial structure of DNA around Igf2 and H19. Modifications in the IME, like abnormal methylation in Wilms' tumors, may represent a novel mechanism for loss of genomic imprinting.

Animals↗
PubMed2025

Efficiency of DNA repair mechanisms of domestic dog primary fibroblasts isolated from small and large breeds of different ages in response to double stranded breaks (DSB).

Aging is associated with increased genomic instability, a phenomenon largely driven by the accumulation of DNA damage over time, and large species of mammals seem to have more robust DNA repair systems associated with longer lives. Among DNA lesions, double-strand breaks (DSBs) are particularly deleterious and have been implicated in age-related functional decline and disease. In this study, we investigated how age and body mass affect the efficiency of DSB repair (DSBr) in primary fibroblast cells isolated from domestic dogs, a species that exhibits significant intraspecies variation in lifespan and body mass. Primary fibroblast cells were isolated from puppies and senior dogs of both large and small breeds. Cells were treated with 100 µM etoposide to induce DSBs and subsequently analyzed at two post-treatment recovery intervals (2 and 24 h) to correlated with the two pathways associated with DSBr, the fast, non-homologous end joining (NHEJ) and the much slower, homologous recombination (HR). Cells were stained for γ-H2AX foci and images were collected using confocal microscopy. We found that mean fluorescence per cell was higher in older dogs of both size classes in the 2 h recovery, indicating higher amounts of DNA damage but suggesting similar efficiencies through the NHEJ repair mechanism in older dogs despite size class. We also show that mean fluorescence per cell was higher in the older large breed dogs in the 24 h recovery, suggesting that the slower phase associated with HR seems to be deficient in cells from older, larger breeds of dogs. These findings support the broader theory that aging is associated with impaired genomic maintenance and establish domestic dogs as a valuable model for studying the cellular mechanisms of age-related genomic instability.

Animals↗
PubMed2005

RNA repair--the latest addition to the toolbox for macromolecular maintenance.

All organisms must keep their DNA intact and free of lesions to obtain faithful transmission of genetic information, and many cellular proteins are involved in genome maintenance. However, also other macromolecules may be subject to repair. Deleterious lesions in proteins can be reversed by several mechanisms, and this appears to be particularly important in cell types where protein turnover is low. In addition, recent studies indicate that RNA is also repaired. AlkB proteins, which were initially found to repair chemically alkylated DNA, can also remove alkyl lesions from RNA, and, moreover, provide functional reactivation of damaged tRNA and mRNA. Here, recent findings on RNA repair are discussed.

Active Transport, Cell Nucleus↗
PubMed2003

High-resolution structure of the E.coli RecQ helicase catalytic core.

RecQ family helicases catalyze critical genome maintenance reactions in bacterial and eukaryotic cells, playing key roles in several DNA metabolic processes. Mutations in recQ genes are linked to genome instability and human disease. To define the physical basis of RecQ enzyme function, we have determined a 1.8 A resolution crystal structure of the catalytic core of Escherichia coli RecQ in its unbound form and a 2.5 A resolution structure of the core bound to the ATP analog ATPgammaS. The RecQ core comprises four conserved subdomains; two of these combine to form its helicase region, while the others form unexpected Zn(2+)-binding and winged-helix motifs. The structures reveal the molecular basis of missense mutations that cause Bloom's syndrome, a human RecQ-associated disease. Finally, based on findings from the structures, we propose a mechanism for RecQ activity that could explain its functional coordination with topoisomerase III.

Adenosine Triphosphatases↗
PubMed2006

Taking it off: regulation of H3 K56 acetylation by Hst3 and Hst4.

Histone modifications have been implicated in both DNA repair and checkpoint-mediated responses to DNA damage. Recently much attention has focused on the acetylation of H3 K56. Indeed, this modification is cell cycle-regulated, maintained upon replicative damage in a checkpoint-dependent manner, and is essential for surviving DNA damage. We and others have discovered that two members of the HDAC Sirtuin family, Hst3 and Hst4, negatively regulate H3 K56 acetylation in budding yeast. Additionally, we have shown that these two HDACs are targeted for repression by the DNA damage checkpoint, which is vital for DNA damage tolerance. Discovery that two HDACs are negative regulators of the cellular response to DNA damage and that they target the acetylation of H3 K56 reveals a complex relationship between histone modifications, HDACs, and the DNA damage response. Here, we discuss the recent reports of the regulation of H3 K56-Ac by Hst3 and Hst4 and put forth the critical questions that remain for understanding the intimate, though poorly characterized, connection between chromatin states and genomic maintenance.

Acetylation↗
PubMed2026

Integrative Genomic and Transcriptomic Insights into High-Altitude Adaptation in Changthangi Goats.

The Changthangi goat, native to the high-altitude Ladakh Plateau in northern India, thrives in oxygen-deficient environments above 4,000 m. This study investigated the genetic basis of high-altitude adaptation in Changthangi goats by integrating comparative genomics and transcriptomics, using the tropical lowland Jamunapari goat as a comparative model. Whole-genome sequence data from 15 individuals per breed were analyzed using complementary selection sweep metrics, including nucleotide diversity, Tajima's D, iHS, CLR, XP-EHH, and FST. These analyses identified candidate genomic regions under strong selective pressure, encompassing genes involved in hypoxia sensing (HIF-1α, HIF-2α/EPAS1, EGLN1), angiogenesis (VEGFA, AGGF1, ZEB1), cardiovascular regulation (PRKCB, ESR1, RYR2), mitochondrial and energy metabolism (ACADSB, ACSS3, ACSL1), cellular stress tolerance (BCL2, ATM), and thermogenesis (UCP1, FGF21). Unlike previous caprine studies that primarily infer hypoxia adaptation from genomic signals alone, our study integrates cardiac transcriptomics to demonstrate that genomic selection in Changthangi goats is accompanied by coordinated transcriptional remodeling across interconnected physiological systems in a physiologically relevant tissue. Comparative cardiac transcriptomic profiling revealed concordant expression divergence in genes associated with oxygen transport, vascular remodeling, mitochondrial function, substrate utilization, redox balance, and genome maintenance. This integrative multi-omics framework provides a mechanistic view of caprine high-altitude adaptation and highlights the value of combining genomic selection analyses with tissue-specific transcriptional profiling to resolve complex adaptive traits.

Animals↗
PubMed2004

Novel and stress-regulated microRNAs and other small RNAs from Arabidopsis.

MicroRNAs (miRNAs) and short interfering RNAs (siRNAs) are small noncoding RNAs that have recently emerged as important regulators of mRNA degradation, translational repression, and chromatin modification. In Arabidopsis thaliana, 43 miRNAs comprising 15 families have been reported thus far. In an attempt to identify novel and abiotic stress regulated miRNAs and siRNAs, we constructed a library of small RNAs from Arabidopsis seedlings exposed to dehydration, salinity, or cold stress or to the plant stress hormone abscisic acid. Sequencing of the library and subsequent analysis revealed 26 new miRNAs from 34 loci, forming 15 new families. Two of the new miRNAs from three loci are members of previously reported miR171 and miR319 families. Some of the miRNAs are preferentially expressed in specific tissues, and several are either upregulated or downregulated by abiotic stresses. Ten of the miRNAs are highly conserved in other plant species. Fifty-one potential targets with diverse function were predicted for the newly identified miRNAs based on sequence complementarity. In addition to miRNAs, we identified 102 other novel endogenous small RNAs in Arabidopsis. These findings suggest that a large number of miRNAs and other small regulatory RNAs are encoded by the Arabidopsis genome and that some of them may play important roles in plant responses to environmental stresses as well as in development and genome maintenance.

Animals↗
PubMed2003

Chk1 and Chk2 kinases in checkpoint control and cancer.

Accumulation of mutations and chromosomal aberrations is one of the hallmarks of cancer cells. This enhanced genetic instability is fueled by defects in the genome maintenance mechanisms including DNA repair and cell cycle checkpoint pathways. Here, we discuss the emerging roles of the mammalian Chk1 and Chk2 kinases as key signal transducers within the complex network of genome integrity checkpoints, as candidate tumor suppressors disrupted in sporadic as well as some hereditary malignancies and as potential targets of new anticancer therapies.

Animals↗
PubMed2002

Defending genome integrity during S-phase: putative roles for RecQ helicases and topoisomerase III.

The maintenance of genome stability is important not only for cell viability, but also for the suppression of neoplastic transformation in higher eukaryotes. It has long been recognised that a common feature of cancer cells is genomic instability. Although the so-called three 'Rs' of genome maintenance, DNA replication, recombination and repair, have historically been studied in isolation, a wealth of recent evidence indicates that these processes are intimately interrelated and interdependent. In this article, we will focus on challenges to the maintenance of genome integrity that arise during the S-phase of the cell cycle, and the possible roles that RecQ helicases and topoisomerase III play in the maintenance of genome integrity during the process of DNA replication.

Adenosine Triphosphatases↗
PubMed2025

Donor transcription suppresses D-loops in cis and promotes genome stability.

DNA is a substrate for competing protein-mediated activities. Whether and how transcription and the synaptic steps of recombination collide or are coordinated has not been investigated. Here, using a controlled break induction system and physical detection of D-loop DNA joint molecules in S. cerevisiae, we show that donor transcription by RNA polymerase II strongly and acutely suppresses D-loops in cis. The extent of this suppression depends on the orientation of transcription, suggesting the preferential usage of one end for the repair of DNA break in transcribed regions. Transcription-mediated D-loop suppression does not rely on endogenous transcription factors, the RNA product, or RNA:DNA hybrids. It is independent of, and can be more potent than the conserved trans D-loop-disruption factors Sgs1-Top3-Rmi1BLM-TOPO3α-RMI1/2, Mph1FANCM, and Srs2. This transcription-mediated control promotes genome maintenance by inhibiting ectopic recombination and multi-invasion-induced rearrangements, while authorizing allelic inter-homolog repair. These findings reveal the prioritization between two universal DNA-dependent processes and its role in promoting genome stability.

Genomic Instability↗
PubMed2006

DDB1 maintains genome integrity through regulation of Cdt1.

DDB1, a component of a Cul4A ubiquitin ligase complex, promotes nucleotide excision repair (NER) and regulates DNA replication. We have investigated the role of human DDB1 in maintaining genome stability. DDB1-depleted cells accumulate DNA double-strand breaks in widely dispersed regions throughout the genome and have activated ATM and ATR cell cycle checkpoints. Depletion of Cul4A yields similar phenotypes, indicating that an E3 ligase function of DDB1 is important for genome maintenance. In contrast, depletion of DDB2, XPA, or XPC does not cause activation of DNA damage checkpoints, indicating that defects in NER are not involved. One substrate of DDB1-Cul4A that is crucial for preventing genome instability is Cdt1. DDB1-depleted cells exhibit increased levels of Cdt1 protein and rereplication, despite containing other Cdt1 regulatory mechanisms. The rereplication, accumulation of DNA damage, and activation of checkpoint responses in DDB1-depleted cells require entry into S phase and are partially, but not completely, suppressed by codepletion of Cdt1. Therefore, DDB1 prevents DNA lesions from accumulating in replicating human cells, in part by regulating Cdt1 degradation.

Animals↗
PubMed1999

The yeast dynamin-like protein, Mgm1p, functions on the mitochondrial outer membrane to mediate mitochondrial inheritance.

The mdm17 mutation causes temperature-dependent defects in mitochondrial inheritance, mitochondrial morphology, and the maintenance of mitochondrial DNA in the yeast Saccharomyces cerevisiae. Defects in mitochondrial transmission to daughter buds and changes in mitochondrial morphology were apparent within 30 min after shifting cells to 37 degrees C, while loss of the mitochondrial genome occurred after 4-24 h at the elevated temperature. The mdm17 lesion mapped to MGM1, a gene encoding a dynamin-like GTPase previously implicated in mitochondrial genome maintenance, and the cloned MGM1 gene complements all of the mdm17 mutant phenotypes. Cells with an mgm1-null mutation displayed aberrant mitochondrial inheritance and morphology. A version of mgm1 mutated in a conserved residue in the putative GTP-binding site was unable to complement any of the mutant defects. It also caused aberrant mitochondrial distribution and morphology when expressed at high levels in cells that also contained a wild-type copy of the gene. Mgm1p was localized to the mitochondrial outer membrane and fractionated as a component of a high molecular weight complex. These results indicate that Mgm1p is a mitochondrial inheritance and morphology component that functions on the mitochondrial surface.

Alleles↗
PubMed1999

DNA repair systems in archaea: mementos from the last universal common ancestor?

DNA repair in the Archaea is relevant to the consideration of genome maintenance and replication fidelity in the last universal common ancestor (LUCA) from two perspectives. First, these prokaryotes embody a mix of bacterial and eukaryal molecular features. Second, DNA repair proteins would have been essential in LUCA to maintain genome integrity, regardless of the environmental temperature. Yet we know very little of the basic molecular mechanisms of DNA damage and repair in the Archaea in general. Many studies on DNA repair in archaea have been conducted with hyperthermophiles because of the additional stress imposed on their macromolecules by high temperatures. In addition, of the six complete archaeal genome sequences published so far, five are thermophilic archaea. We have recently shown that the hyperthermophile Pyrococcus furiosus has an extraordinarily high capacity for repair of radiation-induced double-strand breaks and we have identified and sequenced several genes involved in DNA repair in P. furiosus. At the sequence level, only a few genes share homology with known bacterial repair genes. For instance, our phylogenetic analysis indicates that archaeal recombinases occur in two paralogous gene families, one of which is very deeply branched, and both recombinases are more closely related to the eukaryotic RAD51 and Dmc1 gene families than to the Escherichia coli recA gene. We have also identified a gene encoding a repair endo/exonuclease in the genomes of several Archaea. The archaeal sequences are highly homologous to those of the eukaryotic Rad2 family and they cluster with genes of the FEN-1 subfamily, which are known to be involved in DNA replication and repair in eukaryotes. We argue that there is a commonality of mechanisms and protein sequences, shared between prokaryotes and eukaryotes for several modes of DNA repair, reflecting diversification from a minimal set of genes thought to represent the genome of the LUCA.

Amino Acid Sequence↗