Ultraconserved region-containing Transformer 2β4 controls senescence of colon cancer cells

Ultraconserved regions (UCRs) are 4200 bp genomic segments with perfect human-to-rodent sequence identity. Transcribed UCRs constitute a new category of noncoding RNAs whose functions remain poorly understood. The human transformer 2β (TRA2B) gene contains a 419-bp UCR spanning the 276-bp exon 2 and its neighboring introns. TRA2B exon 2 has premature stop codons, whereas an exon 2-containing splice variant (TRA2β4) was expressed preferentially in the nuclei of human colon cancer cells. TRA2β4 knockdown p53-independently stimulated CDKN1A transcription and increased p21, resulting in the appearance of senescent cells. Biotin pull-down and RNA immunoprecipitation assays revealed that TRA2β4 interacted with Sp1 through a Sp1-binding sequence (485-GGGG-488) in a stem-loop structure of exon 2. Mutation of this sequence (485-AAGG-488) disrupted the stem-loop structure, blocked the interaction with Sp1 and increased CDKN1A transcription. Overexpression of TRA2β4 significantly decreased CDKN1A mRNA levels and accelerated cell growth, but the introduction of the mutation in the Sp1-binding sequence completely canceled these effects. Taken together, TRA2β4 may sequester Sp1 from occupying promoters of target genes including CDKN1A, promoting cell growth by interrupting the senescence-related gene expression program. This novel function of TRA2β4 may uncover an oncogenic function of transcribed UCRs

DNA damage induces reactive oxygen species generation through the H2AX-Nox1/Rac1 pathway

The DNA damage response (DDR) cascade and ROS (reactive oxygen species) signaling are both involved in the induction of cell death after DNA damage, but a mechanistic link between these two pathways has not been clearly elucidated. This study demonstrates that ROS induction after treatment of cells with neocarzinostatin (NCS), an ionizing radiation mimetic, is at least partly mediated by increasing histone H2AX. Increased levels of ROS and cell death induced by H2AX overexpression alone or DNA damage leading to H2AX accumulation are reduced by treating cells with the antioxidant N-Acetyl-L-Cysteine (NAC), the NADP(H) oxidase (Nox) inhibitor DPI, expression of Rac1N17, and knockdown of Nox1, but not Nox4, indicating that induction of ROS by H2AX is mediated through Nox1 and Rac1 GTPase. H2AX increases Nox1 activity partly by reducing the interaction between a Nox1 activator NOXA1 and its inhibitor 14-3-3zeta. These results point to a novel role of histone H2AX that regulates Nox1-mediated ROS generation after DNA damage

Down-regulation of HSP70 sensitizes gastric epithelial cells to apoptosis and growth retardation triggered by H. pylori

<p>Abstract</p> <p>Background</p> <p><it>H. pylori </it>infection significantly attenuated the expression of HSP70 in gastric mucosal cells. However, the role of HSP70 cancellation in <it>H. pylori</it>-associated cell damages is largely unclear.</p> <p>Methods</p> <p>Small interfering RNA (siRNA) was used to down-regulate HSP70 in gastric epithelial cell lines AGS. The transfected cells were then incubated with <it>H. pylori </it>and the functions of HSP70 suppression were observed by viability assay, cell cycle analyses and TUNEL assay. HSP70 target apoptotic proteins were further identified by Western blot.</p> <p>Results</p> <p>The inhibition of HSP70 has further increased the effect of growth arrest and apoptosis activation triggered by <it>H. pylori </it>in gastric epithelial cells. The anti-proliferation function of HSP70 depletion was at least by up-regulating p21 and cell cycle modulation with S-phase accumulation. An increase of apoptosis-inducing factor (AIF) and cytosolic cytochrome C contributes to the activation of apoptosis following down-regulation of intracellular HSP70. Extracellular HSP70 increased cellular resistance to apoptosis by suppression the release of AIF and cytochrome c from mitochondria, as well as inhibition of p21 expression.</p> <p>Conclusions</p> <p>The inhibition of HSP70 aggravated gastric cellular damages induced by <it>H. pylori</it>. Induction of HSP70 could be a potential therapeutic target for protection gastric mucosa from <it>H. pylori</it>-associated injury.</p

<em>Enterococcus faecalis</em> Infection Causes Inflammation, Intracellular Oxphos-Independent ROS Production, and DNA Damage in Human Gastric Cancer Cells

Background: Achlorhydria caused by e.g. atrophic gastritis allows for bacterial overgrowth, which induces chronic inflammation and damage to the mucosal cells of infected individuals driving gastric malignancies and cancer. Enterococcus faecalis (E. faecalis) can colonize achlohydric stomachs and we therefore wanted to study the impact of E. faecalis infection on inflammatory response, reactive oxygen species (ROS) formation, mitochondrial respiration, and mitochondrial genetic stability in gastric mucosal cells. Methods: To separate the changes induced by bacteria from those of the inflammatory cells we established an in vitro E. faecalis infection model system using the gastric carcinoma cell line MKN74. Total ROS and superoxide was measured by fluorescence microscopy. Cellular oxygen consumption was characterized non-invasively using XF24 microplate based respirometry. Gene expression was examined by microarray, and response pathways were identified by Gene Set Analysis (GSA). Selected gene transcripts were verified by quantitative real-time polymerase chain reaction (qRT-PCR). Mitochondrial mutations were determined by sequencing. Results: Infection of MKN74 cells with E. faecalis induced intracellular ROS production through a pathway independent of oxidative phosphorylation (oxphos). Furthermore, E. faecalis infection induced mitochondrial DNA instability. Following infection, genes coding for inflammatory response proteins were transcriptionally up-regulated while DNA damage repair and cell cycle control genes were down-regulated. Cell growth slowed down when infected with viable E. faecalis and responded in a dose dependent manner to E. faecalis lysate. Conclusions: Infection by E. faecalis induced an oxphos-independent intracellular ROS response and damaged the mitochondrial genome in gastric cell culture. Finally the bacteria induced an NF-kappa B inflammatory response as well as impaired DNA damage response and cell cycle control gene expression

Pan-cancer analysis of whole genomes

Cancer is driven by genetic change, and the advent of massively parallel sequencing has enabled systematic documentation of this variation at the whole-genome scale(1-3). Here we report the integrative analysis of 2,658 whole-cancer genomes and their matching normal tissues across 38 tumour types from the Pan-Cancer Analysis of Whole Genomes (PCAWG) Consortium of the International Cancer Genome Consortium (ICGC) and The Cancer Genome Atlas (TCGA). We describe the generation of the PCAWG resource, facilitated by international data sharing using compute clouds. On average, cancer genomes contained 4-5 driver mutations when combining coding and non-coding genomic elements; however, in around 5% of cases no drivers were identified, suggesting that cancer driver discovery is not yet complete. Chromothripsis, in which many clustered structural variants arise in a single catastrophic event, is frequently an early event in tumour evolution; in acral melanoma, for example, these events precede most somatic point mutations and affect several cancer-associated genes simultaneously. Cancers with abnormal telomere maintenance often originate from tissues with low replicative activity and show several mechanisms of preventing telomere attrition to critical levels. Common and rare germline variants affect patterns of somatic mutation, including point mutations, structural variants and somatic retrotransposition. A collection of papers from the PCAWG Consortium describes non-coding mutations that drive cancer beyond those in the TERT promoter(4); identifies new signatures of mutational processes that cause base substitutions, small insertions and deletions and structural variation(5,6); analyses timings and patterns of tumour evolution(7); describes the diverse transcriptional consequences of somatic mutation on splicing, expression levels, fusion genes and promoter activity(8,9); and evaluates a range of more-specialized features of cancer genomes(8,10-18).Peer reviewe

Iron Behaving Badly: Inappropriate Iron Chelation as a Major Contributor to the Aetiology of Vascular and Other Progressive Inflammatory and Degenerative Diseases

The production of peroxide and superoxide is an inevitable consequence of aerobic metabolism, and while these particular "reactive oxygen species" (ROSs) can exhibit a number of biological effects, they are not of themselves excessively reactive and thus they are not especially damaging at physiological concentrations. However, their reactions with poorly liganded iron species can lead to the catalytic production of the very reactive and dangerous hydroxyl radical, which is exceptionally damaging, and a major cause of chronic inflammation. We review the considerable and wide-ranging evidence for the involvement of this combination of (su)peroxide and poorly liganded iron in a large number of physiological and indeed pathological processes and inflammatory disorders, especially those involving the progressive degradation of cellular and organismal performance. These diseases share a great many similarities and thus might be considered to have a common cause (i.e. iron-catalysed free radical and especially hydroxyl radical generation). The studies reviewed include those focused on a series of cardiovascular, metabolic and neurological diseases, where iron can be found at the sites of plaques and lesions, as well as studies showing the significance of iron to aging and longevity. The effective chelation of iron by natural or synthetic ligands is thus of major physiological (and potentially therapeutic) importance. As systems properties, we need to recognise that physiological observables have multiple molecular causes, and studying them in isolation leads to inconsistent patterns of apparent causality when it is the simultaneous combination of multiple factors that is responsible. This explains, for instance, the decidedly mixed effects of antioxidants that have been observed, etc...Comment: 159 pages, including 9 Figs and 2184 reference

Synthetic lethal therapies for cancer: what's next after PARP inhibitors?

The genetic concept of synthetic lethality has now been validated clinically through the demonstrated efficacy of poly(ADP-ribose) polymerase (PARP) inhibitors for the treatment of cancers in individuals with germline loss-of-function mutations in either BRCA1 or BRCA2. Three different PARP inhibitors have now been approved for the treatment of patients with BRCA-mutant ovarian cancer and one for those with BRCA-mutant breast cancer; these agents have also shown promising results in patients with BRCA-mutant prostate cancer. Here, we describe a number of other synthetic lethal interactions that have been discovered in cancer. We discuss some of the underlying principles that might increase the likelihood of clinical efficacy and how new computational and experimental approaches are now facilitating the discovery and validation of synthetic lethal interactions. Finally, we make suggestions on possible future directions and challenges facing researchers in this field

Comprehensive analysis of chromothripsis in 2,658 human cancers using whole-genome sequencing

Chromothripsis is a mutational phenomenon characterized by massive, clustered genomic rearrangements that occurs in cancer and other diseases. Recent studies in selected cancer types have suggested that chromothripsis may be more common than initially inferred from low-resolution copy-number data. Here, as part of the Pan-Cancer Analysis of Whole Genomes (PCAWG) Consortium of the International Cancer Genome Consortium (ICGC) and The Cancer Genome Atlas (TCGA), we analyze patterns of chromothripsis across 2,658 tumors from 38 cancer types using whole-genome sequencing data. We find that chromothripsis events are pervasive across cancers, with a frequency of more than 50% in several cancer types. Whereas canonical chromothripsis profiles display oscillations between two copy-number states, a considerable fraction of events involve multiple chromosomes and additional structural alterations. In addition to non-homologous end joining, we detect signatures of replication-associated processes and templated insertions. Chromothripsis contributes to oncogene amplification and to inactivation of genes such as mismatch-repair-related genes. These findings show that chromothripsis is a major process that drives genome evolution in human cancer

Retrospective evaluation of whole exome and genome mutation calls in 746 cancer samples

Funder: NCI U24CA211006Abstract: The Cancer Genome Atlas (TCGA) and International Cancer Genome Consortium (ICGC) curated consensus somatic mutation calls using whole exome sequencing (WES) and whole genome sequencing (WGS), respectively. Here, as part of the ICGC/TCGA Pan-Cancer Analysis of Whole Genomes (PCAWG) Consortium, which aggregated whole genome sequencing data from 2,658 cancers across 38 tumour types, we compare WES and WGS side-by-side from 746 TCGA samples, finding that ~80% of mutations overlap in covered exonic regions. We estimate that low variant allele fraction (VAF < 15%) and clonal heterogeneity contribute up to 68% of private WGS mutations and 71% of private WES mutations. We observe that ~30% of private WGS mutations trace to mutations identified by a single variant caller in WES consensus efforts. WGS captures both ~50% more variation in exonic regions and un-observed mutations in loci with variable GC-content. Together, our analysis highlights technological divergences between two reproducible somatic variant detection efforts