Mitochondrial protein-linked DNA breaks perturb mitochondrial gene transcription and trigger free radical–induced DNA damage

Breakage of one strand of DNA is the most common form of DNA damage. Most damaged DNA termini require end-processing in preparation for ligation. The importance of this step is highlighted by the association of de- fects in the 3′-end processing enzyme tyrosyl DNA phosphodiesterase 1 (TDP1) and neurodegeneration and by the cytotoxic induction of protein-linked DNA breaks (PDBs) and oxidized nucleic acid intermediates during chemotherapy and radiotherapy. Although much is known about the repair of PDBs in the nucleus, little is known about this process in the mitochondria. We reveal that TDP1 resolves mitochondrial PDBs (mtPDBs), thereby promoting mitochondrial gene transcription. Overexpression of a toxic form of mitochondrial topo- isomerase I (TOP1mt*), which generates excessive mtPDBs, results in a TDP1-dependent compensatory up- regulation of mitochondrial gene transcription. In the absence of TDP1, the imbalance in transcription of mitochondrial- and nuclear-encoded electron transport chain (ETC) subunits results in misassembly of ETC complex III. Bioenergetics profiling further reveals that TDP1 promotes oxidative phosphorylation under both basal and high energy demands. It is known that mitochondrial dysfunction results in free radical leakage and nuclear DNA damage; however, the detection of intermediates of radical damage to DNA is yet to be shown. Consequently, we report an increased accumulation of carbon-centered radicals in cells lacking TDP1, using electron spin resonance spectroscopy. Overexpression of the antioxidant enzyme superoxide dismutase 1 (SOD1) reduces carbon-centered adducts and protects TDP1-deficient cells from oxidative stress. Conversely, overexpression of the amyotrophic lateral sclerosis–associated mutant SOD1G93A leads to marked sensitivity. Whereas Tdp1 knockout mice develop normally, overexpression of SOD1G93A suggests early embryonic lethal- ity. Together, our data show that TDP1 resolves mtPDBs, thereby regulating mitochondrial gene transcription and oxygen consumption by oxidative phosphorylation, thus conferring cellular protection against reactive oxygen species–induced damage

Analyse der Dynamik zellulärer Calcium Signale mit spezifisch lokalisierten Sensorproteinen

Calcium (Ca$^{2+}$) is an essential second messenger that regulates and modulates a variety of cellular functions. In particular, local changes of the calcium concentration play a key role in physiologically relevant processes. The elucidation of temporal and spatial dynamics underlying Ca$^{2+}$ signaling will improve knowledge to understand intracellular signaling mechanisms and the impact of Ca$^{2+}$ on such mechanisms. Therefore, imaging of Ca$^{2+}$ signals is an important field of current investigation. Until recently, synthetic Ca$^{2+}$-sensitive dyes were the only available agents to visualize Ca$^{2+}$ dynamics in living cells. Nowadaysgenetically encoded Ca$^{2+}$ indicators, e.g. GCaMP3.0, are often used as an alternative to monitor calcium signaling. The expression of GCaMP3.0 in eukaryotic cells mainly leads to a homogenous, cytosolic distribution of the protein. To enable the measurement of localized Ca$^{2+}$ signals in different subcellular compartments, the GCaMP3.0 encoding gene was genetically modified as a part of this thesis. Adding short targeting motifs to GCaMP3.0 led to specific subcellular distribution of the protein either in the plasma membrane, the mitochondrial matrix or the mitochondrial outer membrane as well as the golgi apparatus. Additionally, another sensor variant was targeted to the nucleus. Any impairment of the fluorescence properties of the modified sensor proteins was ruled out by in vitro biophysical characterization. Furthermore, HEK293 cells were used as a model system to investigate the spatial localization and functionality of modified as well as non-modified GCaMP3.0. Apart from that, the targeted GCaMP3.0 variants were designed to study temporal and spatial Ca$^{2+}$ dynamics in neurons. Therefore the genes encoding the modified GCaMP3.0 variants were used to generate recombinant Adeno-associated viruses. These viruses were used as gene ferries to deliver GCaMP3.0 variants to primary cultures derived from rat cortex. In both cell types, HEK293 cells and rat cortical neurons, the targeted sensor variants allowed time resolved measurements of localized Ca$^{2+}$ signals

Teaching Peace along with routine education: A simple and easy to implement strategy for a peaceful happy world

In this paper the authors propose ways of implementing peace education andintegrating it in the routine curriculum. Unlike the teaching of academic and vocational skills, which are based on the transmission of knowledge and skills alone, peace education is based on understanding of human motives and, above all, on example.The key individuals in the effort of peace education are teachers in primary, secondary and higher education, and the point at which peace education has to start is in the training of future teachers. Peace education has to communicate both values supportive of peace and non-violence, and the reasoning and interpersonal skills required for non-violent conflict resolution. Rather than introducing formal instruction in”peace science”, teachers may find it more effective to introduce the subject informally with examples that illustrate the value of non-violence, and examples of the ways in which violent conflict can be avoided or resolved. Herein, it is emphasized that peace education can be introduced effectively by merging it with the existing education system. The current paper outlines the concept for introducing peace education and provides an implementation strategy, contributes unique methods, and offers novel suggestions regarding its why, where and how

TDP1/TOP1 ratio as a promising indicator for the response of small cell lung cancer to topotecan

BACKGROUND AND OBJECTIVE Small cell lung cancer (SCLC) is one of the most challenging tumors to treat due to high proliferation rate, early metastatic dissemination and rapid development of chemotherapy resistance. The current treatment protocols involve the use of topoisomerase 1 (TOP1) poisons such as irinotecan and topotecan in combination with platinum-based compounds. TOP1 poisons kill cancer cells by trapping TOP1 on DNA, generating lethal DNA double-strand breaks. A potential mechanism employed by cancer cells to resist killing by TOP1 poisons is to overexpress enzymes involved in the repair of TOP1-DNA breaks. Tyrosyl DNA phosphodiesterase 1 (TDP1) is a key player in this process and despite its importance, no data is currently available to correlate TDP1 protein and mRNA levels with catalytic activity in SCLC. In addition, it is not known if TDP1 and TOP1 protein levels correlate with the cellular response of SCLC to TOP1 based therapies. METHODS AND RESULTS We report a remarkable variation in TDP1 and TOP1 protein levels in a panel of SCLC cell lines. TDP1 protein level correlates well with TDP1 mRNA and TDP1 catalytic activity, as measured by two newly developed independent activity assays, suggesting the potential utility of immunohistochemistry in assessing TDP1 levels in SCLC tissues. We further demonstrate that whilst TDP1 protein level alone does not correlate with topotecan sensitivity, TDP1/TOP1 ratio correlates well with sensitivity in 8 out of 10 cell lines examined. CONCLUSION This study provides the first cellular analyses of TDP1 and TOP1 in SCLC and suggests the potential utility of TDP1/TOP1 ratio to assess the response of SCLC to topotecan. The establishment and validation of an easy-to-use TDP1 enzymatic assay in cell extracts could be exploited as a diagnostic tool in the clinic. These findings may help in stratifying patients that are likely to benefit from TOP1 poisons and TDP1 inhibitors currently under development

Repression of transcription at DNA breaks requires cohesin throughout interphase and prevents genome instability

Cohesin subunits are frequently mutated in cancer, but how they function as tumor suppressors is unknown. Cohesin mediates sister chromatid cohesion, but this is not always perturbed in cancer cells. Here, we identify a previously unknown role for cohesin. We find that cohesin is required to repress transcription at DNA double-strand breaks (DSBs). Notably, cohesin represses transcription at DSBs throughout interphase, indicating that this is distinct from its known role in mediating DNA repair through sister chromatid cohesion. We identified a cancer-associated SA2 mutation that supports sister chromatid cohesion but is unable to repress transcription at DSBs. We further show that failure to repress transcription at DSBs leads to large-scale genome rearrangements. Cancer samples lacking SA2 display mutational patterns consistent with loss of this pathway. These findings uncover a new function for cohesin that provides insights into its frequent loss in cancer

TDP1 deficiency sensitizes human cells to base damage via distinct topoisomerase I and PARP mechanisms with potential applications for cancer therapy

Base damage and topoisomerase I (Top1)-linked DNA breaks are abundant forms of endogenous DNA breakage, contributing to hereditary ataxia and underlying the cytotoxicity of a wide range of anti-cancer agents. Despite their frequency, the overlapping mechanisms that repair these forms of DNA breakage are largely unknown. Here, we report that depletion of Tyrosyl DNA phosphodiesterase 1 (TDP1) sensitizes human cells to alkylation damage and the additional depletion of apurinic/apyrimidinic endonuclease I (APE1) confers hypersensitivity above that observed for TDP1 or APE1 depletion alone. Quantification of DNA breaks and clonogenic survival assays confirm a role for TDP1 in response to base damage, independently of APE1. The hypersensitivity to alkylation damage is partly restored by depletion of Top1, illustrating that alkylating agents can trigger cytotoxic Top1-breaks. Although inhibition of PARP activity does not sensitize TDP1-deficient cells to Top1 poisons, it confers increased sensitivity to alkylation damage, highlighting partially overlapping roles for PARP and TDP1 in response to genotoxic challenge. Finally, we demonstrate that cancer cells in which TDP1 is inherently deficient are hypersensitive to alkylation damage and that TDP1 depletion sensitizes glioblastoma-resistant cancer cells to the alkylating agent temozolomide

IQ and the Weight of Nations

The antecedents of the ‘weights of nations’ have been in the center of theoretical and empirical discussion over the past decades. In this research, we contribute to extant studies by investigating the effect of general intelligence, measured by nation IQ, on the Body Mass Indices (BMI) of male and female populations for 187 countries of the world. Our results suggest an inverted U-shaped link between intelligence and BMI. Even after controlling for an alternative set of control variables such as trade openness, urbanization and others, our results remain intact. This paper documents a turning point of 80.8 for female and 83.7 IQ score for male BMI. The results remain robust to a number of robustness checks

Epigenetic changes in histone acetylation underpin resistance to the topoisomerase I inhibitor irinotecan.

The topoisomerase I (TOP1) inhibitor irinotecan triggers cell death by trapping TOP1 on DNA, generating cytotoxic protein-linked DNA breaks (PDBs). Despite its wide application in a variety of solid tumors, the mechanisms of cancer cell resistance to irinotecan remains poorly understood. Here, we generated colorectal cancer (CRC) cell models for irinotecan resistance and report that resistance is neither due to downregulation of the main cellular target of irinotecan TOP1 nor upregulation of the key TOP1 PDB repair factor TDP1. Instead, the faster repair of PDBs underlies resistance, which is associated with perturbed histone H4K16 acetylation. Subsequent treatment of irinotecan-resistant, but not parental, CRC cells with histone deacetylase (HDAC) inhibitors can effectively overcome resistance. Immunohistochemical analyses of CRC tissues further corroborate the importance of histone H4K16 acetylation in CRC. Finally, the resistant clones exhibit cross-resistance with oxaliplatin but not with ionising radiation or 5-fluoruracil, suggesting that the latter two could be employed following loss of irinotecan response. These findings identify perturbed chromatin acetylation in irinotecan resistance and establish HDAC inhibitors as potential therapeutic means to overcome resistance

In vitro digestibility of dietary carbohydrates: toward a standardized methodology beyond amylolytic and microbial enzymes

This work has been financed by the Spanish Ministry of Economy, Industry and Competitiveness (Project AGL2017-84614-C2-1-R).Peer reviewe