Project description:PurposeVariations in the radiosensitivity of tumor cells within and between tumors impact tumor response to radiation, including the dose required to achieve permanent local tumor control. The increased expression of DNA-PKcs, a key component of a major DNA damage repair pathway in tumors treated by radiation, suggests that DNA-PKcs-dependent repair is likely a cause of tumor cell radioresistance. This study evaluates the relative biological effect of spread-out Bragg-peak protons in DNA-PKcs-deficient cells and the same cells transfected with a functional DNA-PKcs gene.Materials and methodsA cloned radiation-sensitive DNA-PKcs-deficient tumor line and its DNA-PKcs-transfected resistant counterpart were used in this study. The presence of functional DNA-PKcs was evaluated by DNA-PKcs autophosphorylation. Cells to be proton irradiated or x-irradiated were obtained from the same single cell suspension and dilution series to maximize precision. Cells were concurrently exposed to 6-MV x-rays or mid 137-MeV spread-out Bragg peak protons and cultured for colony formation.ResultsThe surviving fraction data were well fit by the linear-quadratic model for each of 8 survival curves. The results suggest that the relative biological effectiveness of mid spread-out Bragg peak protons is approximately 6% higher in DNA-PKcs-mediated resistant tumor cells than in their DNA-PKcs-deficient and radiation-sensitive counterpart.ConclusionDNA-PKcs-dependent repair of radiation damage is less capable of repairing mid spread-out Bragg peak proton lesions than photon-induced lesions, suggesting protons may be more efficient at sterilizing DNA-PKcs-expressing cells that are enriched in tumors treated by conventional fractionated dose x-irradiation.
Project description:Exosomes are complexes containing 3' --> 5' exoribonucleases that have important roles in processing, decay and quality control of various RNA molecules. Archaeal exosomes consist of a hexameric core of three active RNase PH subunits (ribosomal RNA processing factor (Rrp)41) and three inactive RNase PH subunits (Rrp42). A trimeric ring of subunits with putative RNA-binding domains (Rrp4/cep1 synthetic lethality (Csl)4) is positioned on top of the hexamer on the opposite side to the RNA degrading sites. Here, we present the 1.6 A resolution crystal structure of the nine-subunit exosome of Sulfolobus solfataricus and the 2.3 A structure of this complex bound to an RNA substrate designed to be partly trimmed rather than completely degraded. The RNA binds both at the active site on one side of the molecule and on the opposite side in the narrowest constriction of the central channel. Multiple substrate-binding sites and the entrapment of the substrate in the central channel provide a rationale for the processive degradation of extended RNAs and the stalling of structured RNAs.
Project description:PurposeElectrocardiogram-gated computed tomography with coronary angiography can be used for cardiac substructure sparing (CSS) optimization, which identifies and improves avoidance of cardiac substructures when treating with intensity modulated radiotherapy (IMRT). We investigated whether intensity modulated proton therapy (IMPT) would further reduce dose to cardiac substructures for patients with mediastinal lymphoma.Patients and methodsTwenty-one patients with mediastinal lymphoma were enrolled and underwent electrocardiogram-gated computed tomography angiography during or shortly after simulation for radiotherapy planning. Thirteen patients with delineated cardiac substructures underwent comparative planning with both IMPT and IMRT. Plans were normalized for equivalent (95%) target volume coverage for treatment comparison.ResultsThirteen patients met criteria for this study. The median size of the mediastinal lymphadenopathy was 7.9 cm at the greatest diameter. Compared with IMRT-CSS, IMPT-CSS significantly reduced mean dose to all cardiac substructures, including 3 coronary arteries and 4 cardiac valves. Use of IMPT significantly reduced average whole-heart dose from 9.6 to 4.9 Gy (P < .0001), and average mean lung dose was 9.7 vs 5.8 Gy (P < .0001). Prospectively defined clinically meaningful improvement was observed in at least 1 coronary artery in 9 patients (69%), at least 1 cardiac valve in 10 patients (77%), and whole heart in all 13 patients.ConclusionsFor patients with mediastinal lymphoma, IMPT-CSS treatment planning significantly reduced radiation dose to cardiac substructures. The significant improvements outlined in this study for proton therapy suggest possible clinical improvement in alignment with previous analyses of CSS optimization.
Project description:Gasdermins were recently identified as the mediators of pyroptosis - inflammatory cell death triggered by cytosolic sensing of invasive infection and danger signals. Upon activation, gasdermins form cell membrane pores, which release pro-inflammatory cytokines and alarmins and damage the integrity of the cell membrane. Roles for gasdermins in autoimmune and inflammatory diseases, infectious diseases, deafness and cancer are emerging, revealing potential novel therapeutic avenues. Here, we review current knowledge of the family of gasdermins, focusing on their mechanisms of action and roles in normal physiology and disease. Efforts to develop drugs to modulate gasdermin activity to reduce inflammation or activate more potent immune responses are highlighted.
Project description:Despite species-specific differences in the pathways of respiratory metabolism are remarkably conserved across the kingdoms of life with glycolysis, the tricarboxylic acid cycle, and mitochondrial electron transport chain representing the major components of the process in the vast majority of organisms. In addition to being of critical importance in fueling life itself these pathways serve as interesting case studies for substrate channelling with research on this theme having been carried out for over 40 years. Here we provide a cross-kingdom review of the ample evidence for protein-protein interaction and enzyme assemblies within the three component pathways as well as describing the scarcer available evidence for substrate channelling itself.
Project description:Here we report crystal structures of dimethylglycine oxidase (DMGO) from the bacterium Arthrobacter globiformis, a bifunctional enzyme that catalyzes the oxidation of N,N-dimethyl glycine and the formation of 5,10-methylene tetrahydrofolate. The N-terminal region binds FAD covalently and oxidizes dimethylglycine to a labile iminium intermediate. The C-terminal region binds tetrahydrofolate, comprises three domains arranged in a ring-like structure and is related to the T-protein of the glycine cleavage system. The complex with folinic acid indicates that this enzyme selectively activates the N10 amino group for initial attack on the substrate. Dead-end reactions with oxidized folate are avoided by the strict stereochemical constraints imposed by the folate-binding funnel. The active sites in DMGO are approximately 40 A apart, connected by a large irregular internal cavity. The tetrahydrofolate-binding funnel serves as a transient entry-exit port, and access to the internal cavity is controlled kinetically by tetrahydrofolate binding. The internal cavity enables sequestration of the reactive iminium intermediate prior to reaction with tetrahydrofolate and avoids formation of toxic formaldehyde. This mode of channelling in DMGO is distinct from other channelling mechanisms.
Project description:Channelling single-photon emission in multiple well-defined directions and simultaneously controlling its polarization characteristics is highly desirable for numerous quantum technology applications. We show that this can be achieved by using quantum emitters (QEs) nonradiatively coupled to surface plasmon polaritons (SPPs), which are scattered into outgoing free-propagating waves by appropriately designed metasurfaces. The QE-coupled metasurface design is based on the scattering holography approach with radially diverging SPPs as reference waves. Using holographic metasurfaces fabricated around nanodiamonds with single Ge vacancy centres, we experimentally demonstrate on-chip integrated efficient generation of two well-collimated single-photon beams propagating along different 15° off-normal directions with orthogonal linear polarizations.
Project description:Emerging evidence indicates that mitochondrial flashes (mitoflashes) are highly conserved elemental mitochondrial signaling events. However, which signal controls their ignition and how they are integrated with other mitochondrial signals and functions remain elusive. In this study, we aimed to further delineate the signal components of the mitoflash and determine the mitoflash trigger mechanism. Using multiple biosensors and chemical probes as well as label-free autofluorescence, we found that the mitoflash reflects chemical and electrical excitation at the single-organelle level, comprising bursting superoxide production, oxidative redox shift, and matrix alkalinization as well as transient membrane depolarization. Both electroneutral H(+)/K(+) or H(+)/Na(+) antiport and matrix proton uncaging elicited immediate and robust mitoflash responses over a broad dynamic range in cardiomyocytes and HeLa cells. However, charge-uncompensated proton transport, which depolarizes mitochondria, caused the opposite effect, and steady matrix acidification mildly inhibited mitoflashes. Based on a numerical simulation, we estimated a mean proton lifetime of 1.42 ns and diffusion distance of 2.06 nm in the matrix. We conclude that nanodomain protons act as a novel, to our knowledge, trigger of mitoflashes in energized mitochondria. This finding suggests that mitoflash genesis is functionally and mechanistically integrated with mitochondrial energy metabolism.