Atomic structures and functional implications of the archaeal RecQ-like helicase Hjm

<p>Abstract</p> <p>Background</p> <p><it>Pyrococcus furiosus </it>Hjm (<it>Pfu</it>Hjm) is a structure-specific DNA helicase that was originally identified by <it>in vitro </it>screening for Holliday junction migration activity. It belongs to helicase superfamily 2, and shares homology with the human DNA polymerase Θ (PolΘ), HEL308, and <it>Drosophila </it>Mus308 proteins, which are involved in DNA repair. Previous biochemical and genetic analyses revealed that <it>Pfu</it>Hjm preferentially binds to fork-related Y-structured DNAs and unwinds their double-stranded regions, suggesting that this helicase is a functional counterpart of the bacterial RecQ helicase, which is essential for genome maintenance. Elucidation of the DNA unwinding and translocation mechanisms by <it>Pfu</it>Hjm will require its three-dimensional structure at atomic resolution.</p> <p>Results</p> <p>We determined the crystal structures of <it>Pfu</it>Hjm, in two apo-states and two nucleotide bound forms, at resolutions of 2.0–2.7 Å. The overall structures and the local conformations around the nucleotide binding sites are almost the same, including the side-chain conformations, irrespective of the nucleotide-binding states. The architecture of Hjm was similar to that of <it>Archaeoglobus fulgidus </it>Hel308 complexed with DNA. An Hjm-DNA complex model, constructed by fitting the five domains of Hjm onto the corresponding Hel308 domains, indicated that the interaction of Hjm with DNA is similar to that of Hel308. Notably, sulphate ions bound to Hjm lie on the putative DNA binding surfaces. Electron microscopic analysis of an Hjm-DNA complex revealed substantial flexibility of the double stranded region of DNA, presumably due to particularly weak protein-DNA interactions. Our present structures allowed reasonable homology model building of the helicase region of human PolΘ, indicating the strong conformational conservation between archaea and eukarya.</p> <p>Conclusion</p> <p>The detailed comparison between our DNA-free <it>Pfu</it>Hjm structure and the structure of Hel308 complexed with DNA suggests similar DNA unwinding and translocation mechanisms, which could be generalized to all of the members in the same family. Structural comparison also implied a minor rearrangement of the five domains during DNA unwinding reaction. The unexpected small contact between the DNA duplex region and the enzyme appears to be advantageous for processive helicase activity.</p

Novel Conserved-region T-cell Mosaic Vaccine With High Global HIV-1 Coverage Is Recognized by Protective Responses in Untreated Infection

An effective human immunodeficiency virus type 1 (HIV-1) vaccine is the best solution for halting the acquired immune deficiency syndrome epidemic. Here, we describe the design and preclinical immunogenicity of T-cell vaccine expressing novel immunogens tHIVconsvX, vectored by DNA, simian (chimpanzee) adenovirus, and poxvirus modified vaccinia virus Ankara (MVA), a combination highly immunogenic in humans. The tHIVconsvX immunogens combine the three leading strategies for elicitation of effective CD8+ T cells: use of regions of HIV-1 proteins functionally conserved across all M group viruses (to make HIV-1 escape costly on viral fitness), inclusion of bivalent complementary mosaic immunogens (to maximize global epitope matching and breadth of responses, and block common escape paths), and inclusion of epitopes known to be associated with low viral load in infected untreated people (to induce field-proven protective responses). tHIVconsvX was highly immunogenic in two strains of mice. Furthermore, the magnitude and breadth of CD8+ T-cell responses to tHIVconsvX-derived peptides in treatment-naive HIV-1+ patients significantly correlated with high CD4+ T-cell count and low viral load. Overall, the tHIVconsvX design, combining the mosaic and conserved-region approaches, provides an indisputably better coverage of global HIV-1 variants than previous T-cell vaccines. These immunogens delivered in a highly immunogenic framework of adenovirus prime and MVA boost are ready for clinical development

Comparison of effects of bezafibrate and fenofibrate on circulating proprotein convertase subtilisin/kexin type 9 and adipocytokine levels in dyslipidemic subjects with impaired glucose tolerance or type 2 diabetes mellitus: Results from a crossover study

金沢大学医学系研究科Background: Bezafibrate and fenofibrate show different binding properties against peroxisome proliferator-activated receptor subtypes, which could cause different clinical effects on circulating proprotein convertase subtilisin/kexin type 9 (PCSK9) levels and on various metabolic markers. Methods: An open, randomized, four-phased crossover study using 400 mg of bezafibrate or 200 mg of fenofibrate was performed. Study subjects were 14 dyslipidemia with impaired glucose tolerance or type 2 diabetes mellitus (61 ± 16 years, body mass index (BMI) 26 ± 3 kg/m2, total cholesterol (TC) 219 ± 53 mg/dL, triglyceride (TG) 183 ± 83 mg/dL, high-density lipoprotein-cholesterol (HDL-C) 46 ± 8 mg/dL, fasting plasma glucose 133 ± 31 mg/dL and HbA1c 6.2 ± 0.8%). Subjects were given either bezafibrate or fenofibrate for 8 weeks, discontinued for 4 weeks and then switched to the other fibrate for 8 weeks. Circulating PCSK9 levels and other metabolic parameters, including adiponectin, leptin and urine 8-hydroxy-2′-deoxyguanosine (8-OHdG) were measured at 0, 8, 12 and 20 weeks. Results: Plasma PCSK9 concentrations were significantly increased (+39.7% for bezafibrate and +66.8% for fenofibrate, p < 0.001) in all patients except for one subject when treated with bezafibrate. Both bezafibrate and fenofibrate caused reductions in TG (-38.3%, p < 0.001 vs. -32.9%, p < 0.01) and increases in HDL-C (+18.0%, p < 0.001 vs. +11.7%, p < 0.001). Fenofibrate significantly reduced serum cholesterol levels (TC, -11.2%, p < 0.01; non-HDL-C, -17.3%, p < 0.01; apolipoprotein B, -15.1%, p < 0.01), whereas bezafibrate significantly improved glucose tolerance (insulin, -17.0%, p < 0.05) and metabolic markers (γ-GTP, -38.9%, p < 0.01; adiponectin, +15.4%, p < 0.05; urine 8-OHdG/Cre, -9.5%, p < 0.05). Conclusion: Both bezafibrate and fenofibrate increased plasma PCSK9 concentrations. The addition of a PCSK9 inhibitor to each fibrate therapy may achieve beneficial cholesterol lowering along with desirable effects of respective fibrates. © 2011 Elsevier Ireland Ltd. All rights reserved

スマートフォン ト インターネット オ モチイタ トクシマ ケンリツ カイフ ビョウイン エンカク イリョウ シエン システム k-support ノ ドウニュウ

Because a specialist in general medical treatment lacked the Kaifu area of South Tokushima absolutely, we forced a limited doctor to many burdens and performed medical treatment while I always carried risks on my back for the disease except the specialty domain. A stroke specialist in particular is an absent medical depopulated area, and it is difficult to perform the rt-PA IV therapy that is a standard therapy for a stroke for the immediate nature period. Using remote video diagnosis treatment supporting system SYNAPSE ERm that was the smartphone application that FUJIFILM developed for the purpose of canceling these, we introduced smartphone and Tokushima Prefectural Kaifu Hospital remoteness medical treatment support system （k-support） by the Internet as area medical treatment support in February, 2013. This system can provide image information and patient information such as CT or the MRI to a tablet phone and the smartphone of Tokushima Prefectural Kaifu Hospital full-time employment doctors and the doctors who support it, and work in a House in real time. In other words, we can obtain necessary information without asking the when and where and can send appropriate instructions, advice to the Tokushima Prefectural Kaifu Hospital medical attendant from a specialist for it. After introduction, the treatment with this system in 58 emergency patients was carried out in seven months until August 31. The example letting the wide area present the smartphone such as this system and a remote medical treatment support system using the Internet in the medical depopulated area is the first trial in this country

Dual Functions of ASCIZ in the DNA Base Damage Response and Pulmonary Organogenesis

Zn2+-finger proteins comprise one of the largest protein superfamilies with diverse biological functions. The ATM substrate Chk2-interacting Zn2+-finger protein (ASCIZ; also known as ATMIN and ZNF822) was originally linked to functions in the DNA base damage response and has also been proposed to be an essential cofactor of the ATM kinase. Here we show that absence of ASCIZ leads to p53-independent late-embryonic lethality in mice. Asciz-deficient primary fibroblasts exhibit increased sensitivity to DNA base damaging agents MMS and H2O2, but Asciz deletion or knock-down does not affect ATM levels and activation in mouse, chicken, or human cells. Unexpectedly, Asciz-deficient embryos also exhibit severe respiratory tract defects with complete pulmonary agenesis and severe tracheal atresia. Nkx2.1-expressing respiratory precursors are still specified in the absence of ASCIZ, but fail to segregate properly within the ventral foregut, and as a consequence lung buds never form and separation of the trachea from the oesophagus stalls early. Comparison of phenotypes suggests that ASCIZ functions between Wnt2-2b/ß-catenin and FGF10/FGF-receptor 2b signaling pathways in the mesodermal/endodermal crosstalk regulating early respiratory development. We also find that ASCIZ can activate expression of reporter genes via its SQ/TQ-cluster domain in vitro, suggesting that it may exert its developmental functions as a transcription factor. Altogether, the data indicate that, in addition to its role in the DNA base damage response, ASCIZ has separate developmental functions as an essential regulator of respiratory organogenesis

Recent Results from LHD Experiment with Emphasis on Relation to Theory from Experimentalist’s View

he Large Helical Device (LHD) has been extending an operational regime of net-current free plasmas towardsthe fusion relevant condition with taking advantage of a net current-free heliotron concept and employing a superconducting coil system. Heating capability has exceeded 10 MW and the central ion and electron temperatureshave reached 7 and 10 keV, respectively. The maximum value of β and pulse length have been extended to 3.2% and 150 s, respectively. Many encouraging physical findings have been obtained. Topics from recent experiments, which should be emphasized from the aspect of theoretical approaches, are reviewed. Those are (1) Prominent features in the inward shifted configuration, i.e., mitigation of an ideal interchange mode in the configuration with magnetic hill, and confinement improvement due to suppression of both anomalous and neoclassical transport, (2) Demonstration ofbifurcation of radial electric field and associated formation of an internal transport barrier, and (3) Dynamics of magnetic islands and clarification of the role of separatrix

DNA損傷応答タンパク質ASCIZは塩基除去修復と遺伝子変換をリンクする

京都大学0048新制・課程博士博士(医学)甲第15576号医博第3479号新制||医||982(附属図書館)28097京都大学大学院医学研究科分子医学系専攻(主査)教授 渡邉 正己, 教授 平岡 眞寛, 教授 小松 賢志学位規則第4条第1項該当Doctor of Medical ScienceKyoto UniversityDA