DNA double-strand breaks (DSBs) are toxic DNA lesions, which, if not properly repaired, may lead to genomic instability, cell death and senescence. Damage-induced long non-coding RNAs (dilncRNAs) are transcribed from broken DNA ends and contribute to DNA damage response (DDR) signaling. Here we show that dilncRNAs play a role in DSB repair by homologous recombination (HR) by contributing to the recruitment of the HR proteins BRCA1, BRCA2, and RAD51, without affecting DNA-end resection. In S/G2-phase cells, dilncRNAs pair to the resected DNA ends and form DNA:RNA hybrids, which are recognized by BRCA1. We also show that BRCA2 directly interacts with RNase H2, mediates its localization to DSBs in the S/G2 cell-cycle phase, and controls DNA:RNA hybrid levels at DSBs. These results demonstrate that regulated DNA:RNA hybrid levels at DSBs contribute to HR-mediated repair.

The copper-catalyzed cycloaddition reaction between a propargyl-appended europium complex and azidomethylferrocene yields a d-f dyad whose photophysical properties can be reversibly switched by varying the oxidation state of the ferrocene chromophore.

The most frequently mutated oncogene in cancer is KRAS, which utilizes alternative fourth exons to generate two gene products, KRAS4A and KRAS4B, that differ only in their C-terminal membrane-targeting region 1. Because oncogenic mutations occur in exons 2 or 3, when KRAS is activated by mutation two constitutively active KRAS proteins are encoded, each capable of transforming cells 2. No functional distinctions among the splice variants have been established. Oncogenic KRAS alters tumor metabolism 3. Among these alterations is increased glucose uptake and glycolysis, even in the presence of abundant oxygen 4 (the Warburg Effect). Whereas these metabolic effects of oncogenic KRAS have been explained by transcriptional upregulation of glucose transporters and glycolytic enzymes 3-5 , direct regulation of metabolic enzymes has not been examined. We report a direct, GTP-dependent interaction between KRAS4A and hexokinase 1 (HK1) that alters the activity of the kinase, establishing HK1 as an effector of KRAS4A. The interaction is unique to KRAS4A because the palmitoylation/depalmitoylation cycle of this RAS isoform permits co-localization with HK1 on the outer mitochondrial membrane (OMM). KRAS4A expression in cancer may drive unique metabolic vulnerabilities that can be exploited therapeutically. Users may view, print, copy, and download text and data-mine the content in such documents, for the purposes of academic research, subject always to the full Conditions of use:

It is clear that a crowded environment influences the structure, dynamics, and interactions of biological molecules, but the complexity of this phenomenon demands the development of new experimental and theoretical approaches. Here we use two complementary single-molecule FRET techniques to show that the kinetics of DNA base pairing and unpairing, which are fundamental to both the biological role of DNA and its technological applications, are strongly modulated by a crowded environment. We directly observed single DNA hairpins, which are excellent model systems for studying hybridization, either freely diffusing in solution or immobilized on a surface under crowding conditions. The hairpins followed two-state folding dynamics with a closing rate increasing by 4-fold and the opening rate decreasing 2-fold, for only modest concentrations of crowder [10% (w/w) polyethylene glycol (PEG)]. These experiments serve both to unambiguously highlight the impact of a crowded environment on a fundamental biological process, DNA base pairing, and to illustrate the benefits of single-molecule approaches to probing the structure and dynamics of complex biomolecular systems.

One of the most central questions about the repair of a double-strand DNA break (DSB) concerns how the two free DNA ends are brought together — a step called synapsis. Using single-molecule FRET (smFRET), we show here that both Ku plus XRCC4:DNA ligase IV are necessary and sufficient to achieve a flexible synapsis of blunt DNA ends, whereas either alone is not. Addition of XLF causes a transition to a close synaptic state, and maximum efficiency of close synapsis is achieved within 20 min. The promotion of close synapsis by XLF indicates a role that is independent of a filament structure, with action focused at the very ends of each duplex. DNA-PKcs is not required for the formation of either the flexible or close synaptic states. This model explains in biochemical terms the evolutionarily central synaptic role of Ku, X4L4, and XLF in NHEJ for all eukaryotes.