Dynamic self-assembly of DNA minor groove-binding ligand DB921 into nanotubes triggered by an alkali halide.

We describe a novel self-assembling supramolecular nanotube system formed by a heterocyclic cationic molecule which was originally designed for its potential as an antiparasitic and DNA sequence recognition agent. Our structural characterisation work indicates that the nanotubes form via a hierarchical assembly mechanism that can be triggered and tuned by well-defined concentrations of simple alkali halide salts in water. The nanotubes assembled in NaCl have inner and outer diameters of ca. 22 nm and 26 nm respectively, with lengths that reach into several microns. Our results suggest the tubes consist of DB921 molecules stacked along the direction of the nanotube long axis. The tubes are stabilised by face-to-face π-π stacking and ionic interactions between the charged amidinium groups of the ligand and the negative halide ions. The assembly process of the nanotubes was followed using small-angle X-ray and neutron scattering, transmission electron microscopy and ultraviolet/visible spectroscopy. Our data demonstrate that assembly occurs through the formation of intermediate ribbon-like structures that in turn form helices that tighten and compact to form the final stable filament. This assembly process was tested using different alkali-metal salts, showing a strong preference for chloride or bromide anions and with little dependency on the type of cation. Our data further demonstrates the existence of a critical anion concentration above which the rate of self-assembly is greatly enhanced

PARP14 is a PARP with both ADP-ribosyl transferase and hydrolase activities

This is the final version. Available on open access from the American Association for the Advancement of Science via the DOI in this recordData availability: The MS proteomics data have been deposited to the ProteomeXchange Consortium via the PRIDE (92) partner repository with the dataset identifier PXD043452.PARP14 is a mono-ADP-ribosyl transferase involved in the control of immunity, transcription, and DNA replication stress management. However, little is known about the ADP-ribosylation activity of PARP14, including its substrate specificity or how PARP14-dependent ADP-ribosylation is reversed. We show that PARP14 is a dual-function enzyme with both ADP-ribosyl transferase and hydrolase activity acting on both protein and nucleic acid substrates. In particular, we show that the PARP14 macrodomain 1 is an active ADP-ribosyl hydrolase. We also demonstrate hydrolytic activity for the first macrodomain of PARP9. We reveal that expression of a PARP14 mutant with the inactivated macrodomain 1 results in a marked increase in mono(ADP-ribosyl)ation of proteins in human cells, including PARP14 itself and antiviral PARP13, and displays specific cellular phenotypes. Moreover, we demonstrate that the closely related hydrolytically active macrodomain of SARS2 Nsp3, Mac1, efficiently reverses PARP14 ADP-ribosylation in vitro and in cells, supporting the evolution of viral macrodomains to counteract PARP14-mediated antiviral response.Biotechnology and Biological Sciences Research Council (BBSRC)Wellcome TrustOxford University Challenge Seed FundEdward Penley Abraham Research FundOvarian Cancer Research AllianceResearch Council of NorwaySwedish Research CouncilMedical Research Council (MRC)Novo Nordisk Foundation Center for Protein ResearchDanish Council of Independent ResearchDanish Cancer Societ

Reversible mono-ADP-ribosylation of DNA breaks.

ADP-ribosylation is a chemical modification of macromolecules that plays an important role in regulation of quintessential biological processes such as DNA repair, transcription, chromatin remodelling, stress response, apoptosis, bacterial metabolism and many others. ADP-ribosylation is carried out by ADP-ribosyltransferase proteins, such as poly(ADP-ribose) polymerases (PARPs) that transfer either monomer or polymers of ADP-ribose onto the molecular targets by using nicotinamide adenine dinucleotide (NAD(+) ) as a cofactor. Traditionally, proteins have been described as primary targets of ADP-ribosylation; however, there has been growing evidence that DNA may be a common target as well. Here, we show using biochemical studies that PARP3, a DNA damage activated ADP-ribosyltransferase, can mono ADP-ribosylate double stranded DNA ends. ADP-ribosylation of DNA mediated by PARP3 attaches a single mono-ADP-ribose moiety to the phosphate group at the terminal ends of DNA. We further show that mono ADP-ribosylation at DNA ends can be efficiently reversed by several cellular hydrolases (PARG, MACROD2, TARG1 and ARH3). This suggests that mono ADP-ribosylated DNA adducts can be efficiently removed in cells by several mechanisms. This article is protected by copyright. All rights reserved

Reversible mono-ADP-ribosylation of DNA breaks.

ADP-ribosylation is a chemical modification of macromolecules that plays an important role in regulation of quintessential biological processes such as DNA repair, transcription, chromatin remodelling, stress response, apoptosis, bacterial metabolism and many others. ADP-ribosylation is carried out by ADP-ribosyltransferase proteins, such as poly(ADP-ribose) polymerases (PARPs) that transfer either monomer or polymers of ADP-ribose onto the molecular targets by using nicotinamide adenine dinucleotide (NAD(+) ) as a cofactor. Traditionally, proteins have been described as primary targets of ADP-ribosylation; however, there has been growing evidence that DNA may be a common target as well. Here, we show using biochemical studies that PARP3, a DNA damage activated ADP-ribosyltransferase, can mono ADP-ribosylate double stranded DNA ends. ADP-ribosylation of DNA mediated by PARP3 attaches a single mono-ADP-ribose moiety to the phosphate group at the terminal ends of DNA. We further show that mono ADP-ribosylation at DNA ends can be efficiently reversed by several cellular hydrolases (PARG, MACROD2, TARG1 and ARH3). This suggests that mono ADP-ribosylated DNA adducts can be efficiently removed in cells by several mechanisms. This article is protected by copyright. All rights reserved

Pathogenesis and virulence of flavivirus infections

The Flavivirus genus consists of >70 members including several that are considered significant human pathogens. Flaviviruses display a broad spectrum of diseases that can be roughly categorised into two phenotypes – systemic disease involving haemorrhage exemplified by dengue and yellow Fever virus, and neurological complications associated with the likes of West Nile and Zika viruses. Attempts to develop vaccines have been variably successful against some. Besides, mosquito-borne flaviviruses can be vertically transmitted in the arthropods, enabling long term persistence and the possibility of re-emergence. Therefore, developing strategies to combat disease is imperative even if vaccines become available. The cellular interactions of flaviviruses with their human hosts are key to establishing the viral lifecycle on the one hand, and activation of host immunity on the other. The latter should ideally eradicate infection, but often leads to immunopathological and neurological consequences. In this review, we use Dengue and Zika viruses to discuss what we have learned about the cellular and molecular determinants of the viral lifecycle and the accompanying immunopathology, while highlighting current knowledge gaps which need to be addressed in future studies

MacroD1 Is a Promiscuous ADP-Ribosyl Hydrolase Localized to Mitochondria

MacroD1 is a macrodomain containing protein that has mono-ADP-ribose hydrolase enzymatic activity toward several ADP-ribose adducts. Dysregulation of MacroD1 expression has been shown to be associated with the pathogenesis of several forms of cancer. To date, the physiological functions and sub-cellular localization of MacroD1 are unclear. Previous studies have described nuclear and cytosolic functions of MacroD1. However, in this study we show that endogenous MacroD1 protein is highly enriched within mitochondria. We also show that MacroD1 is highly expressed in human and mouse skeletal muscle. Furthermore, we show that MacroD1 can efficiently remove ADP-ribose from 5' and 3'-phosphorylated double stranded DNA adducts in vitro. Overall, we have shown that MacroD1 is a mitochondrial protein with promiscuous enzymatic activity that can target the ester bonds of ADP-ribosylated phosphorylated double-stranded DNA ends. These findings have exciting implications for MacroD1 and ADP-ribosylation within the regulation of mitochondrial function and DNA-damage in vivo

Targeting telomerase and telomeres: a click chemistry approach towards highly selective G-quadruplex ligands.

Maintenance of telomeres--specialized complexes that protect the ends of chromosomes, is undertaken by the enzyme complex telomerase, which is a key factor that is activated in more than 80% of cancer cells, but is absent in most normal cells. Targeting telomere maintenance mechanisms could potentially halt tumour growth across a broad spectrum of cancer types, with little cytotoxic effect outside cancer cells. Here, we describe in detail a new class of G-quadruplex binding ligands synthesized using a click chemistry approach. These ligands comprise a 1,3-di(1,2,3-triazol-4-yl)benzene pharmacophore, and display high levels of selectivity for interaction with G-quadruplex DNA vs. duplex DNA. The ability of these ligands to inhibit the enzymatic activity of telomerase correlates with their ability to stabilize quadruplex DNA, and with estimates of affinity calculated by molecular modeling

MacroD1 is a promiscuous ADP-ribosyl hydrolase localized to mitochondria

MacroD1 is a macrodomain containing protein that has mono-ADP-ribose hydrolase enzymatic activity toward several ADP-ribose adducts. Dysregulation of MacroD1 expression has been shown to be associated with the pathogenesis of several forms of cancer. To date, the physiological functions and sub-cellular localization of MacroD1 are unclear. Previous studies have described nuclear and cytosolic functions of MacroD1. However, in this study we show that endogenous MacroD1 protein is highly enriched within mitochondria. We also show that MacroD1 is highly expressed in human and mouse skeletal muscle. Furthermore, we show that MacroD1 can efficiently remove ADP-ribose from 5' and 3'-phosphorylated double stranded DNA adducts in vitro. Overall, we have shown that MacroD1 is a mitochondrial protein with promiscuous enzymatic activity that can target the ester bonds of ADP-ribosylated phosphorylated double-stranded DNA ends. These findings have exciting implications for MacroD1 and ADP-ribosylation within the regulation of mitochondrial function and DNA-damage in vivo

Relationship between epicardial fat and quantitative coronary artery plaque progression: insights from computer tomography coronary angiography

First online: 03 September 2015Epicardial fat volume (EFV) has been suggested to promote atherosclerotic plaque development in coronary arteries, and has been correlated with both coronary stenosis and acute coronary events. Although associated with progression of coronary calcification burden, a relationship with progression of coronary atheroma volume has not been previously tested. We studied patients who had clinically indicated serial 320-row multi-detector computer tomography coronary angiography with a median 25-month interval. EFV was measured at baseline and follow-up. In vessels with coronary stenosis, quantitative analysis was performed to measure atherosclerotic plaque burden, volume and aggregate plaque volume at baseline and follow-up. The study comprised 64 patients (58.4 ± 12.2 years, 27 males, 192 vessels, 193 coronary segments). 79 (41 %) coronary segments had stenosis at baseline. Stenotic segments were associated with greater baseline EFV than those without coronary stenosis (117.4 ± 45.1 vs. 102.3 ± 51.6 cm(3), P = 0.046). 46 (24 %) coronary segments displayed either new plaque formation or progression of adjusted plaque burden at follow-up. These were associated with higher baseline EFV than segments without stenosis or those segments that had stenoses that did not progress (128.7 vs. 101.0 vs. 106.7 cm(3) respectively, P = 0.006). On multivariate analysis, baseline EFV was the only independent predictor of coronary atherosclerotic plaque progression or new development (P = 0.014). High baseline EFV is associated with the presence of coronary artery stenosis and plaque volume progression. Accumulation of EFV may be implicated in the evolution and progression of coronary atheroma.Peter J. Psaltis, Andrew H. Talman, Kiran Munnur, James D. Cameron, Brian S. H. Ko, Ian T. Meredith, Sujith K. Seneviratne, Dennis T. L. Won