The XRCC1 phosphate-binding pocket binds poly (ADP-ribose) and is required for XRCC1 function

Poly (ADP-ribose) is synthesized at DNA single-strand breaks and can promote the recruitment of the scaffold protein, XRCC1. However, the mechanism and importance of this process has been challenged. To address this issue, we have characterized the mechanism of poly (ADP-ribose) binding by XRCC1 and examined its importance for XRCC1 function. We show that the phosphate-binding pocket in the central BRCT1 domain of XRCC1 is required for selective binding to poly (ADP-ribose) at low levels of ADP-ribosylation, and promotes interaction with cellular PARP1. We also show that the phosphate-binding pocket is required for EGFP-XRCC1 accumulation at DNA damage induced by UVA laser, H2O2, and at sites of sub-nuclear PCNA foci, suggesting that poly (ADP-ribose) promotes XRCC1 recruitment both at single-strand breaks globally across the genome and at sites of DNA replication stress. Finally, we show that the phosphate-binding pocket is required following DNA damage for XRCC1-dependent acceleration of DNA single-strand break repair, DNA base excision repair, and cell survival. These data support the hypothesis that poly (ADP-ribose) synthesis promotes XRCC1 recruitment at DNA damage sites and is important for XRCC1 function

Energy levels of light nuclei. III

Recent years have seen the accumulation of a considerable amount of new information, both experimental and theoretical, pertaining to the location and character of the excited states in the light nuclei. We have attempted in the following sections to summarize the pertinent facts appearing in the literature and to present this material in a systematic way. To a large extend the organization of the present article follows the pattern of the two previous papers on this subject.(1,2) The present article contains a summary of the most important information appearing in the earlier papers and is intended to supersede them entirely

Efficient single-strand break repair requires binding to both poly(ADP-ribose) and DNA by the central BRCT domain of XRCC1

XRCC1 accelerates repair of DNA single-strand breaks by acting as a scaffold protein for the recruitment of Pol-beta, LigIII-alpha and end-processing factors such as PNKP and APTX. XRCC1 itself is recruited to DNA damage through interaction of its central BRCT domain with poly-(ADP-ribose) chains generated by PARP1 or PARP2. XRCC1 is believed to interact directly with DNA at sites of damage, but the molecular basis for this interaction within XRCC1 remains unclear. We now show that the central BRCT domain simultaneously mediates interaction of XRCC1 with poly-(ADP-ribose) and DNA, through separate and non-overlapping binding sites on opposite faces of the domain. Mutation of residues within the DNA binding site, which includes the site of a common disease-associated human polymorphism, affects DNA binding of this XRCC1 domain in vitro, and impairs XRCC1 recruitment and retention at DNA damage and repair of single-strand breaks in vivo

Tests of electron flavor conservation with the Sudbury Neutrino Observatory

We analyze tests of electron flavor conservation that can be performed at the Sudbury Neutrino Observatory (SNO). These tests, which utilize $^8$B solar neutrinos interacting with deuterium, measure: 1) the shape of the recoil electron spectrum in charged-current (CC) interactions (the CC spectrum shape); and 2) the ratio of the number of charged current to neutral current (NC) events (the CC/NC ratio). We determine standard model predictions for the CC spectral shape and for the CC/NC ratio, together with realistic estimates of their errors and the correlations between errors. We consider systematic uncertainties in the standard neutrino spectrum and in the charged-current and neutral current cross-sections, the SNO energy resolution and absolute energy scale, and the SNO detection efficiencies. Assuming that either matter-enhanced or vacuum neutrino oscillations solve the solar neutrino problems, we calculate the confidence levels with which electron flavor non-conservation can be detected using either the CC spectrum shape or the CC/NC ratio, or both. If the SNO detector works as expected, the neutrino oscillation solutions that best-fit the results of the four operating solar neutrino experiments can be distinguished unambiguously from the standard predictions of electron flavor conservation.Comment: 31 pages (RevTeX) + 10 figures (postscript). Requires epsfig.sty. Gzipped figures also available at ftp://ftp.sns.ias.edu/pub/lisi/snopaper . To appear in Phys. Rev.

Mode of action of DNA-competitive small molecule inhibitors of tyrosyl DNA phosphodiesterase 2

TDP2 is a 5’-tyrosyl DNA phosphodiesterase important for the repair of DNA adducts generated by non-productive (abortive) activity of topoisomerase II. TDP2 facilitates therapeutic resistance to topoisomerase poisons, which are widely used in the treatment of a range of cancer types. Consequently, TDP2 is an interesting target for the development of small molecule inhibitors that could restore sensitivity to topoisomerase-directed therapies. Previous studies identified a class of deazaflavin-based molecules that showed inhibitory activity against TDP2 at therapeutically useful concentrations, but their mode of action was uncertain. We have confirmed that the deazaflavin series inhibits TDP2 enzyme activity in a fluorescence-based assay, suitable for HTS-screening. We have gone on to determine crystal structures of these compounds bound to a ‘humanised’ form of murine TDP2. The structures reveal their novel mode of action as competitive ligands for the binding site of an incoming DNA substrate, and point the way to generating novel and potent inhibitors of TDP2

Is poetry therapy an appropriate intervention for clients recovering from anorexia? A critical review of the literature and client report

© 2017 Informa UK Limited, trading as Taylor & Francis Group. Poetry therapy is an arts-based psychotherapeutic intervention, often delivered in groups. This paper argues that the process and benefits of poetry therapy may be particularly suited to clients recovering from anorexia, as an adjunct to other treatments. Poetry therapy and its history are described briefly, and the relevance of poetry therapy for clients recovering from anorexia is outlined. After one client contributes her experience of this treatment for illustration, the paper offers a review of the evidence base for poetry therapy for eating disorders, and argues that, while research is limited, further research is warranted. Finally, a description of one form of clinical application is offered, to enable replication

XRCC1 mutation is associated with PARP1 hyperactivation and cerebellar ataxia

XRCC1 is a molecular scaffold protein that assembles multi-protein complexes involved in DNA single-strand break repair1,2. Here we show that biallelic mutations in the human XRCC1 gene are associated with ocular motor apraxia, axonal neuropathy, and progressive cerebellar ataxia. Cells from a patient with mutations in XRCC1 exhibited not only reduced rates of single-strand break repair but also elevated levels of protein ADP-ribosylation. This latter phenotype is recapitulated in a related syndrome caused by mutations in the XRCC1 partner protein PNKP3,4,5 and implicates hyperactivation of poly(ADP-ribose) polymerase/s as a cause of cerebellar ataxia. Indeed, remarkably, genetic deletion of Parp1 rescued normal cerebellar ADP-ribose levels and reduced the loss of cerebellar neurons and ataxia in Xrcc1-defective mice, identifying a molecular mechanism by which endogenous single-strand breaks trigger neuropathology. Collectively, these data establish the importance of XRCC1 protein complexes for normal neurological function and identify PARP1 as a therapeutic target in DNA strand break repair-defective disease

MiTF links Erk1/2 kinase and p21CIP1/WAF1 activation after UVC radiation in normal human melanocytes and melanoma cells

As a survival factor for melanocytes lineage cells, MiTF plays multiple roles in development and melanomagenesis. What role MiTF plays in the DNA damage response is currently unknown. In this report we observed that MiTF was phosphorylated at serine 73 after UVC radiation, which was followed by proteasome-mediated degradation. Unlike after c-Kit stimulation, inhibiting p90RSK-1 did not abolish the band shift of MiTF protein, nor did it abolish the UVC-mediated MiTF degradation, suggesting that phosphorylation on serine 73 by Erk1/2 is a key event after UVC. Furthermore, the MiTF-S73A mutant (Serine 73 changed to Alanine via site-directed mutagenesis) was unable to degrade and was continuously expressed after UVC exposure. Compared to A375 melanoma cells expressing wild-type MiTF (MiTF-WT), cells expressing MiTF-S73A mutant showed less p21WAF1/CIP1 accumulation and a delayed p21WAF1/CIP1 recovery after UVC. Consequently, cells expressing MiTF-WT showed a temporary G1 arrest after UVC, but cells expressing MiTF-S73A mutant or lack of MiTF expression did not. Finally, cell lines with high levels of MiTF expression showed higher resistance to UVC-induced cell death than those with low-level MiTF. These data suggest that MiTF mediates a survival signal linking Erk1/2 activation and p21WAF1/CIP1 regulation via phosphorylation on serine 73, which facilitates cell cycle arrest. In addition, our data also showed that exposure to different wavelengths of UV light elicited different signal pathways involving MiTF