九州大学 研究者情報
研究者情報 (研究者の方へ)入力に際してお困りですか?
基本情報 研究活動 教育活動 社会活動
石野 良純(いしの よしずみ) データ更新日:2018.06.14

教授 /  農学研究院 生命機能科学部門 生物機能分子化学講座


主な研究テーマ
メタゲノム解析手法を用いた海の環境モニタリング
キーワード:メタゲノム解析、次世代シークエンシング、海洋環境
2011.10~2018.03.
天然変性タンパク質の分子認識機構と機能発現
キーワード:天然変性蛋白質、分子認識、蛋白質構造、
2009.11~2014.03.
DNA複製の分子機構解析
キーワード:DNA複製、分子機構、アーキア、古細菌、極限環境微生物
1990.01.
組換え修復の分子機構解析
キーワード:DNA repair, DNA recombination, endonucelase, homologous recombination, Holliday junction
1996.04.
遺伝子工学技術開発
キーワード:遺伝子工学, 遺伝子増幅, 組換えDNA, 核酸関連酵素
1983.04.
従事しているプロジェクト研究
極限環境下におけるアーキアの遺伝情報維持機構の解明 (基盤研究 A)
2014.04~2017.03, 代表者:石野良純, 九州大学
本研究計画は、極限環境下における生命維持現象の解明を目指すものである。すなわち、100℃の熱水中で生息する超好熱性アーキア(古細菌)がどのようにして、厳しい環境下に適応して遺伝情報を維持し、子孫に伝達していくのかという仕組みを明らかにする。アーキアは真正細菌とも真核生物とも異なる進化的に独立した第三の生物であり、太古の地球から現在まで生息してきた。本研究は、代表的な超好熱性アーキアである Thermococcal目のアーキア株を用いて、遺伝学的手法と生化学的手法を融合させながら、さらにオミックス手法を加えて、遺伝情報維持と正確な伝達機構をシステマティックに解明するという、世界初の研究提案である。本提案の研究は、超高温という特殊環境での生命現象の解明と同時に、他の生物ドメインとの比較生物学によって、生物が獲得した遺伝情報複製、修復、転写、翻訳装置の作動原理に迫ることができる。.
農林水産技術会議委託プロジェクト研究
2011.04~2016.03, 代表者:五條堀 孝(JV1)、佐野 元彦 (JV2), 国立遺伝学研究所 教授(JV1), 中央水産研究所 (JV2), 農林水産省 技術会議.
Digital DNA Chip による生物多様性評価手法の開発
2011.01~2017.03, 代表者:五條堀 孝, 国立遺伝学研究所 教授, JST CREST研究.
超高次複合体解析に基づくゲノム動態研究プロジェクト
2009.10~2011.03, 代表者:片山 勉, 九州大学, 九州大学
九州大学P&Pとして20年度に採択された研究・教育プロジェクトである。代表者は片山教授(薬学研究院)
.
天然変性タンパク質の分子認識機構と機能発現
2009.10~2014.09, 代表者:佐藤  衛, 横浜市立大学, 文部科学省(日本)
文部科学省、新学術領域研究として20年度採択された研究プロジェクトであり、領域代表は佐藤  衛教授(横浜市立大)で、この中の計画代表を務める。.
実践による超分子複合体モデリングシステムの開発
2005.10, 代表者:白井 剛, 長浜バイオ大学, 科学技術振興機構
バイオインフォーマティクス推進事業.
特定領域研究「染色体サイクルの制御ネットワーク」
2005.10, 代表者:正井久雄, 東京都臨床医学総合研究所, 文部科学省.
特定領域研究「ゲノムホメオスタシスの分子機構」
2001.10~2006.03, 代表者:品川日出夫, 大阪大学, 文部科学省.
研究業績
主要著書
1. 石野良純、跡見晴幸監修, アーキア生物学, 共立出版, 2017.10, [URL],  「地球上の生物は大きく3つのグループに分けられる」という3ドメイン説は、最近では生物学の専門家だけではなく、多くの人が認識するようになってきた。しかし、それを正しく説明できる人はまだまだ少ないであろう。本誌は、“第3の生物”と呼ばれるアーキアに関する生物学の教科書であり、地球上の3種の生物のうちの一つとして進化してきたアーキアについての基本的な知識を供給するものである。
 筆者がアーキアを知ったのは1990年代の初めであり、好熱菌の生命現象に興味を持ち始めた時であった。80℃以上の高温で生育することができる超好熱菌と呼ばれる生物のほとんどが細菌(真正細菌)とは区別されるアーキアであることを知ってたいへん驚いた。筆者はDNA鎖を合成することで知られるDNAポリメラーゼに興味を持っていた。 当時、DNAポリメラーゼは大きく分けて、細菌が持つPol I型(大腸菌DNA Polymerase I に代表される)と真核生物が有するα型(ヒト DNA polymerase α に代表される)と呼ばれていたが、アーキアが真核生物と類似したα型の酵素を有することに大変興味を引かれた。複製装置の分子進化とその根本原理を理解するために、アーキアを研究材料としてDNA複製研究を行いたいと思い現在まで続けてきたが、この30年の間にアーキア研究によって多くの興奮を味わってきた。
 アメリカ合衆国イリノイ大学のカールウース (Carl R Woese)によって、アーキアが第3の生物として提唱されたのは1977年のことであるが、本年はちょうど40周年になる。10年前の2007年11月にイリノイ大学において、カールウースを讃えて、 “Hidden before our eyes” という、アーキア発見30周年を祝う記念シンポジウムが開催された。筆者はこのシンポジウムに参加し、ウースがアーキアを発見した時の研究室を訪ねた。そこには30年前にリボソームRNAの解析をしたオートラジオグラフィーのフィルムがきれいに整理されたX線フィルムの箱がずらっと並んでいた(巻頭写真 1)。そこから取り出された一枚のフィルムは独特のスポット模様を示しており、それが、アーキアが第3の生物であることを示す生データであった(巻頭写真 2)。それから5年後、カールウースは2012年12月30日にこの世を去った。現在我々は、地球上の生物が3つのドメインに分けられるということを知っており、バクテリア、ユーカリアと共に、アーキアの生物学は日々発展している。
 我が国におけるアーキアの研究は、1988年に設立された「日本アーキバクテリア研究会」が中心になって発展してきた。本会は学会ではなく、年会費も集めないで、アーキアが好きな研究者が年に一度集まって講演会を開催する。2002年からは「日本アーキア研究会」と改名し、活動を続けている。アーキアを主題にした英語のテキストブックは数冊出版されてはいるものの、日本語で書かれたアーキアの教科書的書物は2冊しかなく、1998年に出版された「古細菌の生物学」(東大出版会)から約20年が経過している。カールウースによるアーキアの提唱から40周年に当たる本年、アーキア研究会で活動している研究者が分担して、アーキアについての現在までの知見をとりまとめることによって、アーキアに興味を持たれる高校理科の教諭、学部、大学院学生、また生物学に興味を持たれる一般の方々に、アーキアについてより理解していただこうと考えた。各章は、それぞれの内容を専門としている現役のアーキア研究者に執筆いただいたので、日進月歩であるアーキア研究の最先端内容までを含んでいる。専門家でないと、少々難しい内容まで踏み込んでいる部分もあるが、興味を持って読んでいただけることを願っている。本書が多くの人々のアーキアという生物の理解につながり、また、一人でも多くの若者へアーキア生物学の面白さを伝えられて、アーキアの研究をしてみたいという志を持ってもらえるきっかけとなれば至福の喜びである。

.
2. 石野 良純, 「進化するゲノム編集技術」, 株式会社エヌティーエス, 17-28ページ, 2015.10, ある遺伝子の機能を知りたい時には、その遺伝子が破壊された変異体を人工的に作成し、その表現型が野生型生物と比較してどのように変化するかを調べる遺伝学的手法が有効である。生物が有する相同的DNA組換え能を利用して標的の遺伝子を改変するジーンターゲティング技術は、それが比較的容易な微生物の一部のモデル生物では分子生物学の発展とともに、日常的に用いられてきた。1989年にノックアウトマウス作製技術が開発されて以来、高等真核生物においてもこの手法が利用され始め、遺伝子欠損生物作製までに煩雑な操作と長時間を要するにも.
3. Yoshizumi Ishino, Sonoko Iahino, DNA replication in thermophilic microorganisms, In Thermophilic Microorganisms. Horizon Scientific Press, Norwich, UK, Horizon Scientific Press, Norwich, UK, 2015.05, DNA replication is essential for maintaining genetic information and transferring it from ancestor to descendant. To protect the genetic information from mutations, biological organisms have acquired several types of DNA repair systems. The extreme thermophiles living on the earth are microorganisms from the Bacteria and Archaea domains. The structures of the bacterial and archaeal genomes are circular, and the mechanism of replication initiation, by the binding of the initiator protein to the replication origin (oriC), is conserved in the two domains. The elongation process is also conserved, because similar primase, helicase, polymerase, and ligase functions are observed in the two domains. However, the proteins involved in the DNA replication process are quite different between Bacteria and Archaea, and their replication machineries seem to have evolved independently. The DNA repair system in extreme thermophiles should work efficiently to maintain their genome integrity at high temperatures. However, the DNA repair systems are diverse and still not comprehensively understood yet, although research in this field has been actively pursued in the systems from Escherichia coli to human. In this chapter, we focus on DNA replication in the thermophilic bacteria and archaea, and summarize the current understanding of the molecular mechanisms of replication in thermophilic microorganisms.

.
4. Yoshizumi Ishino, Sonoko Iahino, The Mechanisms of DNA Replication
DNA Replication in Archaea, the Third Domain of Life
, INTECH d.o.o., Rijeka, Croatia, pp. 91-126, 2013.03.
5. Sonoko Iahino, Yoshizumi Ishino, Thermophilic microbes in environmental and industrial biotechnology
DNA polymerases and DNA ligases
, Springer, 429-457, 2013.03.
6. Sonoko Iahino, Yoshizumi Ishino, Microorganisms in Sustainable Agriculture and Biotechnology.
Application of environmental DNA resources to create useful DNA polymerases with different properties.
, Springer, pp. 663-678, 2012.03.
主要原著論文
1. Ishino, S., Skouloubris, S., Kudo, H., l'Hermitte-Stead, C., Es-Sadik, A., Lambry, J.C., Ishino, Y., and Myllykallio, H. , Activation of the mismatch-specific endonuclease EndoMS/NucS by the replication clamp is required for high fidelity DNA replication. , doi: 10.1093/nar/gky460. , 45, 印刷中, 2018.06, The mismatch repair (MMR) system, exemplified by the MutS/MutL proteins, is widespread in Bacteria and Eukarya. However, molecular mechanisms how numerous archaea and bacteria lacking the mutS/mutL genes maintain high replication fidelity and genome stability have remained elusive. EndoMS is a recently discovered hyperthermophilic mismatch-specific endonuclease encoded by nucS in Thermococcales. We deleted the nucS from the actinobacterium Corynebacterium glutamicum and demonstrated a drastic increase of spontaneous transition mutations in the nucS deletion strain. The observed spectra of these mutations were consistent with the enzymatic properties of EndoMS in vitro. The robust mismatch-specific endonuclease activity was detected with the purified C. glutamicum EndoMS protein but only in the presence of the β-clamp (DnaN). Our biochemical and genetic data suggest that the frequently occurring G/T mismatch is efficiently repaired by the bacterial EndoMS-β-clamp complex formed via a carboxy-terminal sequence motif of EndoMS proteins. Our study thus has great implications for understanding how the activity of the novel MMR system is coordinated with the replisome and provides new mechanistic insight into genetic diversity and mutational patterns in industrially and clinically (e.g. Mycobacteria) important archaeal and bacterial phyla previously thought to be devoid of the MMR system..
2. Miyazono, K.I., Ishino, S., Makita, N., Ito, T., Ishino, Y., and Tanokura, M. , Crystal structure of the novel lesion-specific endonuclease PfuEndoQ from Pyrococcus furiosus. , Nucleic Acids Res., doi: 10.1093/nar/gky261., 46, 9, 4807 -4818, 2018.05, Because base deaminations, which are promoted by high temperature, ionizing radiation, aerobic respiration and nitrosative stress, produce mutations during replication, deaminated bases must be repaired quickly to maintain genome integrity. Recently, we identified a novel lesion-specific endonuclease, PfuEndoQ, from Pyrococcus furiosus, and PfuEndoQ may be involved in the DNA repair pathway in Thermococcales of Archaea. PfuEndoQ recognizes a deaminated base and cleaves the phosphodiester bond 5' of the lesion site. To elucidate the structural basis of the substrate recognition and DNA cleavage mechanisms of PfuEndoQ, we determined the structure of PfuEndoQ using X-ray crystallography. The PfuEndoQ structure and the accompanying biochemical data suggest that PfuEndoQ recognizes a deaminated base using a highly conserved pocket adjacent to a Zn2+-binding site and hydrolyses a phosphodiester bond using two Zn2+ ions. The PfuEndoQ-DNA complex is stabilized by a Zn-binding domain and a C-terminal helical domain, and the complex may recruit downstream proteins in the DNA repair pathway..
3. Daimon, K., Ishino, S., Imai, N., Nagumo, S., Yamagami, T., Matsukawa, H. and Ishino, Y. , Two Family B DNA Polymerases From Aeropyrum pernix, Based on Revised Translational Frames. , Frontiers in Molecular Biosciences , doi: 10.3389/fmolb.2018.00037. , 16, 5, 37, 2018.04, Living organisms are divided into three domains, Bacteria, Eukarya, and Archaea. Comparative studies in the three domains have provided useful information to understand the evolution of the DNA replication machinery. DNA polymerase is the central enzyme of DNA replication. The presence of multiple family B DNA polymerases is unique in Crenarchaeota, as compared with other archaeal phyla, which have a single enzyme each for family B (PolB) and family D (PolD). We analyzed PolB1 and PolB3 in the hyperthermophilic crenarchaeon, Aeropyrum pernix, and found that they are larger proteins than those predicted from the coding regions in our previous study and from public database annotations. The recombinant larger PolBs exhibited the same DNA polymerase activities as previously reported. However, the larger PolB3 showed remarkably higher thermostability, which made this enzyme applicable to PCR. In addition, the high tolerance to salt and heparin suggests that PolB3 will be useful for amplification from the samples with contaminants, and therefore it has a great potential for diagnostic use in the medical and environmental field..
4. Nagata, M., Ishino, S., Yamagami, T., Simons, J-R., Kanai, T., Atomi, H., and Ishino, Y. , Possible function of the second RecJ-like protein in stalled replication fork repair by interacting with Hef. , Sci Rep. , doi: 10.1038/s41598-017-17306-0., 7, 1, 16949, 2017.12, RecJ was originally identified in Escherichia coli and plays an important role in the DNA repair and recombination pathways. Thermococcus kodakarensis, a hyperthermophilic archaeon, has two RecJ-like nucleases. These proteins are designated as GAN (GINS-associated nuclease) and HAN (Hef-associated nuclease), based on the protein they interact with. GAN is probably a counterpart of Cdc45 in the eukaryotic CMG replicative helicase complex. HAN is considered mainly to function with Hef for restoration of the stalled replication fork. In this study, we characterized HAN to clarify its functions in Thermococcus cells. HAN showed single-strand specific 3' to 5' exonuclease activity, which was stimulated in the presence of Hef. A gene disruption analysis revealed that HAN was non-essential for viability, but the ΔganΔhan double mutant did not grow under optimal conditions at 85 °C. This deficiency was not fully recovered by introducing the mutant han gene, encoding the nuclease-deficient HAN protein, back into the genome. These results suggest that the unstable replicative helicase complex without GAN performs ineffective fork progression, and thus the stalled fork repair system including HAN becomes more important. The nuclease activity of HAN is required for the function of this protein in T. kodakarensis..
5. Nagata, M., Ishino, S., Yamagami, T., Ogino, H., Simons, J-R., Kanai, T., Atomi, H., and Ishino, Y. , The Cdc45/RecJ-like protein forms a complex with GINS and MCM, and is important for DNA replication in Thermococcus kodakarensis. , Nucleic Acids Res., doi: 10.1093/nar/gkx740., 45, 18, 10693. -10705. , 2017.10, The archaeal minichromosome maintenance (MCM) has DNA helicase activity, which is stimulated by GINS in several archaea. In the eukaryotic replicative helicase complex, Cdc45 forms a complex with MCM and GINS, named as CMG (Cdc45-MCM-GINS). Cdc45 shares sequence similarity with bacterial RecJ. A Cdc45/RecJ-like protein from Thermococcus kodakarensis shows a bacterial RecJ-like exonuclease activity, which is stimulated by GINS in vitro. Therefore, this archaeal Cdc45/RecJ is designated as GAN, from GINS-associated nuclease. In this study, we identified the CMG-like complex in T. kodakarensis cells. The GAN·GINS complex stimulated the MCM helicase, but MCM did not affect the nuclease activity of GAN in vitro. The gene disruption analysis showed that GAN was non-essential for its viability but the Δgan mutant did not grow at 93°C. Furthermore, the Δgan mutant showed a clear retardation in growth as compared with the parent cells under optimal conditions at 85°C. These deficiencies were recovered by introducing the gan gene encoding the nuclease deficient GAN protein back to the genome. These results suggest that the replicative helicase complex without GAN may become unstable and ineffective in replication fork progression. The nuclease activity of GAN is not related to the growth defects of the Δgan mutant cells.

.
6. Antranikian, G., Suleiman, M., Schafers, C., Adams, MWW., Bartolucci, S., Blamey, JM., Birkeland, NK., Bonch-Osmolovskaya, E., da Costa, MS., Cowan, D., Danson, M., Forterre, P., Kelly, R., Ishino, Y., Littlechild, J., Moracci, M., Noll, K., Oshima, T., Robb, F., Rossi, M., Santos, H., Schonheit, P., Sterner, R., Thauer, R.,Thomm, M., Wiegel, J., and Stetter, KO., Diversity of bacteria and archaea from two shallow marine hydrothermal vents from Vulcano Island., Extremophiles., doi: 10.1007/s00792-017-0938-y., 21, 733.-742., 2017.07, To obtain new insights into community compositions of hyperthermophilic microorganisms, defined as having optimal growth temperatures of 80 °C and above, sediment and water samples were taken from two shallow marine hydrothermal vents (I and II) with temperatures of 100 °C at Vulcano Island, Italy. A combinatorial approach of denaturant gradient gel electrophoresis (DGGE) and metagenomic sequencing was used for microbial community analyses of the samples. In addition, enrichment cultures, growing anaerobically on selected polysaccharides such as starch and cellulose, were also analyzed by the combinatorial approach. Our results showed a high abundance of hyperthermophilic archaea, especially in sample II, and a comparable diverse archaeal community composition in both samples. In particular, the strains of the hyperthermophilic anaerobic genera Staphylothermus and Thermococcus, and strains of the aerobic hyperthermophilic genus Aeropyrum, were abundant. Regarding the bacterial community, ε-Proteobacteria, especially the genera Sulfurimonas and Sulfurovum, were highly abundant. The microbial diversity of the enrichment cultures changed significantly by showing a high dominance of archaea, particularly the genera Thermococcus and Palaeococcus, depending on the carbon source and the selected temperature..
7. Liu, S., Ishino, S., Ishino, Y., Pehau-Arnaudet, G., Krupovic, M., and Prangishvili, D. , A novel type of polyhedral viruses infecting hyperthermophilic archaea., J. Virol. , doi: 10.1128/JVI.00589-17., 91, e00589-17. , 2017.06, Encapsidation of genetic material into polyhedral particles is one of the most common structural solutions employed by viruses infecting hosts in all three domains of life. Here, we describe a new virus of hyperthermophilic archaea, Sulfolobus polyhedral virus 1 (SPV1), which condenses its circular double-stranded DNA genome in a manner not previously observed for other known viruses. The genome complexed with virion proteins is wound up sinusoidally into a spherical coil which is surrounded by an envelope and further encased by an outer polyhedral capsid apparently composed of the 20-kDa virion protein. Lipids selectively acquired from the pool of host lipids are integral constituents of the virion. None of the major virion proteins of SPV1 show similarity to structural proteins of known viruses. However, minor structural proteins, which are predicted to mediate host recognition, are shared with other hyperthermophilic archaeal viruses infecting members of the order Sulfolobales The SPV1 genome consists of 20,222 bp and contains 45 open reading frames, only one-fifth of which could be functionally annotated.IMPORTANCE Viruses infecting hyperthermophilic archaea display a remarkable morphological diversity, often presenting architectural solutions not employed by known viruses of bacteria and eukaryotes. Here we present the isolation and characterization of Sulfolobus polyhedral virus 1, which condenses its genome into a unique spherical coil. Due to the original genomic and architectural features of SPV1, the virus should be considered a representative of a new viral family, "Portogloboviridae.".
8. Ogino, H., Ishino, S., Kohda, D., and Ishino, Y., The RecJ2 protein in the thermophilic archaeon Thermoplasma acidophilum is a 3'-5' exonuclease and associates with a DNA replication complex., J. Biol. Chem., doi: 10.1074/jbc.M116.767921., 292, 7921.-7931., 2017.05, RecJ/cell division cycle 45 (Cdc45) proteins are widely conserved in the three domains of life, i.e. in bacteria, Eukarya, and Archaea. Bacterial RecJ is a 5'-3' exonuclease and functions in DNA repair pathways by using its 5'-3' exonuclease activity. Eukaryotic Cdc45 has no identified enzymatic activity but participates in the CMG complex, so named because it is composed of Cdc45, minichromosome maintenance protein complex (MCM) proteins 2-7, and GINS complex proteins (Sld5, Psf11-3). Eukaryotic Cdc45 and bacterial/archaeal RecJ share similar amino acid sequences and are considered functional counterparts. In Archaea, a RecJ homolog in Thermococcus kodakarensis was shown to associate with GINS and accelerate its nuclease activity and was, therefore, designated GAN (GINS-associated nuclease); however, to date, no archaeal RecJ·MCM·GINS complex has been isolated. The thermophilic archaeon Thermoplasma acidophilum has two RecJ-like proteins, designated TaRecJ1 and TaRecJ2. TaRecJ1 exhibited DNA-specific 5'-3' exonuclease activity, whereas TaRecJ2 had 3'-5' exonuclease activity and preferred RNA over DNA. TaRecJ2, but not TaRecJ1, formed a stable complex with TaGINS in a 2:1 molar ratio. Furthermore, the TaRecJ2·TaGINS complex stimulated activity of TaMCM (T. acidophilum MCM) helicase in vitro, and the TaRecJ2·TaMCM·TaGINS complex was also observed in vivo However, TaRecJ2 did not interact with TaMCM directly and was not required for the helicase activation in vitro These findings suggest that the function of archaeal RecJ in DNA replication evolved divergently from Cdc45 despite conservation of the CMG-like complex formation between Archaea and Eukarya..
9. Yoda, T., Tanabe, M., Tsuji, T.,Yoda, T., Ishino, S., Shirai, T., Ishino, Y., Takeyama, H., and Nishida, H. , Exonuclease processivity of archaeal replicative DNA polymerase in association with PCNA is expedited by mismatches in DNA. , Sci. Rep. , doi: 10.1038/srep44582., 7, 44582, 2017.03, Family B DNA polymerases comprise polymerase and 3' ->5' exonuclease domains, and detect a mismatch in a newly synthesized strand to remove it in cooperation with Proliferating cell nuclear antigen (PCNA), which encircles the DNA to provide a molecular platform for efficient protein-protein and protein-DNA interactions during DNA replication and repair. Once the repair is completed, the enzyme must stop the exonucleolytic process and switch to the polymerase mode. However, the cue to stop the degradation is unclear. We constructed several PCNA mutants and found that the exonuclease reaction was enhanced in the mutants lacking the conserved basic patch, located on the inside surface of PCNA. These mutants may mimic the Pol/PCNA complex processing the mismatched DNA, in which PCNA cannot interact rigidly with the irregularly distributed phosphate groups outside the dsDNA. Indeed, the exonuclease reaction with the wild type PCNA was facilitated by mismatched DNA substrates. PCNA may suppress the exonuclease reaction after the removal of the mismatched nucleotide. PCNA seems to act as a "brake" that stops the exonuclease mode of the DNA polymerase after the removal of a mismatched nucleotide from the substrate DNA, for the prompt switch to the DNA polymerase mode..
10. Yoshizumi Ishino, Sonoko Ishino, A functional endonuclease Q exists in the bacterial domain: identification and characterization of endonuclease Q from Bacillus pumilus., Biosci. Biotechnol. Biochem. 81, 931-937. doi: 10.1080/09168451.2016.1277946., doi: 10.1080/09168451.2016.1277946., 81, 931-937., 2017.01, DNA base deamination occurs spontaneously
under physiological conditions and is promoted by
high temperature. Therefore, hyperthermophiles are
expected to have efficient repair systems of the
deaminated bases in their genomes. Endonuclease Q
(EndoQ) was originally identified from the hyperthermophlic
archaeon, Pyrococcus furiosus, as a
hypoxanthine-specific endonuclease recently. Further
biochemical analyses revealed that EndoQ also recognizes
uracil, xanthine, and the AP site in DNA,
and is probably involved in a specific repair process
for damaged bases. Initial phylogenetic analysis
showed that an EndoQ homolog is found only in the
Thermococcales and some of the methanogens in
Archaea, and is not present in most members of the
domains Bacteria and Eukarya. A better understanding
of the distribution of the EndoQ-mediated
repair system is, therefore, of evolutionary interest.
We showed here that an EndoQ-like polypeptide
from Bacillus pumilus, belonging to the bacterial
domain, is functional and has similar properties
with the archaeal EndoQs..
11. Yoshizumi Ishino, Sonoko Ishino, Structure of the EndoMS-DNA complex as mismatch-restriction endonuclease. Structure 24, 1960-1971. doi: 10.1016/j.str.2016.09.005., Structure, http://dx.doi.org/10.1016/j.str.2016.09.005, 24, 1960-1971, 2016.11, Archaeal NucS nuclease was thought to degrade
the single-stranded region of branched DNA, which
contains flapped and splayed DNA. However,
recent findings indicated that EndoMS, the orthologous
enzyme of NucS, specifically cleaves doublestranded
DNA (dsDNA) containing mismatched
bases. In this study, we determined the structure of
the EndoMS-DNA complex. The complex structure
of the EndoMS dimer with dsDNA unexpectedly revealed
that the mismatched bases were flipped out
into binding sites, and the overall architecture most
resembled that of restriction enzymes. The structure
of the apo form was similar to the reported structure
of Pyrococcus abyssi NucS, indicating that movement
of the C-terminal domain from the resting state
was required for activity. In addition, a model of
the EndoMS-PCNA-DNA complex was preliminarily
verified with electron microscopy. The structures
strongly support the idea that EndoMS acts in a
mismatch repair pathway..
12. Yoshizumi Ishino, Sonoko Ishino, Takeshi YAMAGAMI, Atomic structure of an archaeal GAN suggests its dual roles as an exonuclease in DNA repair and a CMG component in DNA replication. , Nucleic Acids Res., doi: 10.1093/nar/gkw789 , 44, 19, 9509-9517, 2017.05, In eukaryotic DNA replication initiation, hexameric
MCM (mini-chromosome maintenance) unwinds the
template double-stranded DNA to form the replication
fork. MCM is activated by two proteins, Cdc45
and GINS, which constitute the ‘CMG’ unwindosome
complex together with the MCM core. The archaeal
DNA replication system is quite similar to that of
eukaryotes, but only limited knowledge about the
DNA unwinding mechanism is available, froma structural
point of view. Here, we describe the crystal
structure of an archaeal GAN (GINS-associated nuclease)
from Thermococcus kodakaraensis, the homolog
of eukaryotic Cdc45, in both the free form and
the complex with the C-terminal domain of the cognate
Gins51 subunit (Gins51C). This first archaeal
GAN structure exhibits a unique, ‘hybrid’ structure
between the bacterial RecJ and the eukaryotic Cdc45.
GAN possesses the conserved DHH and DHH1 domains
responsible for the exonuclease activity, and
an inserted CID (CMG interacting domain)-like domain
structurally comparable to that in Cdc45, suggesting
its dual roles as an exonuclease in DNA repair
and a CMG component in DNA replication. A
structural comparison of the GAN–Gins51C complex
with the GINS tetramer suggests that GINS uses the
mobile Gins51C as a hook to bind GAN for CMG formation.
INTRODUCTION
DNA replication is essential for all living organisms, and
the basic mechanism is conserved across the three domains
of life, Bacteria, Archaea, and Eukarya. DNA replication
must occur accurately in a highly coordinated manner regulated
by numerous proteins (1). At the initiation of DNA
replication, the parental double-stranded DNA (dsDNA)
is unwound to generate two single-stranded DNAs (ssDNAs),
which form a replication fork. In eukaryotes, the
hetero-hexameric MCM (mini-chromosome maintenance)
comprisingMCM2–7acts as the helicase core to unwind the
templateDNA (2,3). Although isolatedMCMgenerally exhibits
weak helicase activity, two protein factors, Cdc45 and
GINS, have been identified as MCM activators, and they
form the CMG complex holoenzyme together with MCM
(4,5). CMG is constructed on the DNA by the sequential
loading of MCM (as the double-hexameric ring at the formation
of Pre-RC; pre-replicating complex), Cdc45 (at the
formation of Pre-IC; pre-initiation complex) and GINS (at
the Pre-LC; pre-loading complex after pre-IC), rather than
the loading of the pre-assembled complex (3,6,7).CMGformation
is controlled by two protein kinases, CDK (cyclindependent
kinase) and DDK (dbf4-dependent kinase), and
is.
13. Yoshizumi Ishino, Sonoko Ishino, Archaeal DNA polymerase-B as a DNA template guardian: links between polymerases and base/alternative excision repair enzymes in handling the deaminated bases uracil and hypoxanthine., Archaea, http://dx.doi.org/10.1155/2016/1510938, 2016, Article ID 1510938, 8 pages, 2016.08, In Archaea repair of uracil and hypoxanthine, which arise by deamination of cytosine and adenine, respectively, is initiated by three
enzymes: Uracil-DNA-glycosylase (UDG, which recognises uracil); Endonuclease V (EndoV, which recognises hypoxanthine); and
Endonuclease Q (EndoQ), (which recognises both uracil and hypoxanthine). Two archaeal DNA polymerases, Pol-B and Pol-D,
are inhibited by deaminated bases in template strands, a feature unique to this domain.Thus the three repair enzymes and the two
polymerases show overlapping specificity for uracil and hypoxanthine. Here it is demonstrated that binding of Pol-D to primertemplates
containing deaminated bases inhibits the activity of UDG, EndoV, and EndoQ. Similarly Pol-B almost completely turns
off EndoQ, extending earlier work that demonstrated that Pol-B reduces catalysis by UDG and EndoV. Pol-B was observed to be
a more potent inhibitor of the enzymes compared to Pol-D. Although Pol-D is directly inhibited by template strand uracil, the
presence of Pol-B further suppresses any residual activity of Pol-D, to near-zero levels. The results are compatible with Pol-D acting
as the replicative polymerase and Pol-B functioning primarily as a guardian preventing deaminated base-induced DNAmutations..
14. Yoshizumi Ishino, Sonoko Ishino, DJ-1 family Maillard deglycases prevent acrylamide formation. Biochem Biophys Res Commun., Biochem Biophys Res Commun., http://dx.doi.org/10.1016/j.bbrc.2016.08.077, 478, 1111-1116, 2016.08, The presence of acrylamide in food is a worldwide concern because it is carcinogenic, reprotoxic and
neurotoxic. Acrylamide is generated in the Maillard reaction via condensation of reducing sugars and
glyoxals arising from their decomposition, with asparagine, the amino acid forming the backbone of the
acrylamide molecule. We reported recently the discovery of the Maillard deglycases (DJ-1/Park7 and its
prokaryotic homologs) which degrade Maillard adducts formed between glyoxals and lysine or arginine
amino groups, and prevent glycation damage in proteins. Here, we show that these deglycases prevent
acrylamide formation, likely by degrading asparagine/glyoxal Maillard adducts. We also report the discovery
of a deglycase from the hyperthermophilic archaea Pyrococcus furiosus, which prevents acrylamide
formation at 100 C. Thus, Maillard deglycases constitute a unique enzymatic method to prevent
acrylamide formation in food without depleting the components (asparagine and sugars) responsible for
its formation..
15. Yoshizumi Ishino, Sonoko Ishino, Takeshi YAMAGAMI, PCNA is involved in the EndoQ-mediated DNA repair process in Thermococcales. , Scientific Report, DOI: 10.1038/srep25532, 6, 25532, 2016.05, To maintain genome integrity for transfer to their offspring, and to maintain order in cellular processes,
all living organisms have DNA repair systems. Besides the well-conserved DNA repair machineries,
organisms thriving in extreme environments are expected to have developed efficient repair
systems. We recently discovered a novel endonuclease, which cleaves the 5′ side of deoxyinosine,
from the hyperthermophilic archaeon, Pyrococcus furiosus. The novel endonuclease, designated as
Endonulcease Q (EndoQ), recognizes uracil, abasic site and xanthine, as well as hypoxanthine, and cuts
the phosphodiester bond at their 5′ sides. To understand the functional process involving EndoQ, we
searched for interacting partners of EndoQ and identified Proliferating Cell Nuclear Angigen (PCNA).
The EndoQ activity was clearly enhanced by addition of PCNA in vitro. The physical interaction between
the two proteins through a PIP-motif of EndoQ and the toroidal structure of PCNA are critical for the
stimulation of the endonuclease activity. These findings provide us a clue to elucidate a unique DNA
repair system in Archaea..
16. Yoshizumi Ishino, Sonoko Ishino, Takeshi YAMAGAMI, Identification of a mismatch-specific endonuclease in hyperthermophilic Archaea. , Nucleic Acids Res. 44, 2977-2986., 10.1093/nar/gkw153, 44, 7, 2977-2986., 2016.03, The common mismatch repair system processed by
MutS and MutL and their homologs was identified
in Bacteria and Eukarya. However, no evidence of a
functional MutS/L homolog has been reported for archaeal
organisms, and it is not known whether the
mismatch repair system is conserved in Archaea.
Here, we describe an endonuclease that cleaves
double-stranded DNA containing a mismatched base
pair, from the hyperthermophilic archaeon Pyrococcus
furiosus. The corresponding gene revealed
that the activity originates from PF0012, and we
named this enzyme Endonuclease MS (EndoMS)
as the mismatch-specific Endonuclease. The sequence
similarity suggested that EndoMS is the ortholog
of NucS isolated from Pyrococcus abyssi,
published previously. Biochemical characterizations
of the EndoMS homolog from Thermococcus kodakarensis
clearly showed that EndoMS specifically
cleaves both strands of double-stranded DNA into 5′-
protruding forms, with the mismatched base pair in
the central position. EndoMS cleaves G/T, G/G, T/T,
T/C andA/Gmismatches,with amore preference for
G/T, G/G and T/T, but has very little or no effect on
C/C, A/C andA/Amismatches. The discovery of this
endonuclease suggests the existence of a novel mismatch
repair process, initiated by the double-strand
break generated by the EndoMS endonuclease, in Archaea
and some Bacteria..
17. Yoshizumi Ishino, Sonoko Ishino, Takeshi YAMAGAMI, A longer finger-subdomain of family A DNA polymerases found by metagenomic analysis strengthens DNA binding and primer extension abilities, doi: 10.1016/j.gene.2015.10.030., 576, (2 Pt 1), 690-695, 2016.02, The family A DNA polymerases from thermophilic bacteria are useful for PCR. The DNA polymerase from Thermus aquaticus (Taq polymerase) was the original enzyme used when practical PCR was developed, and it has remained the standard enzyme for PCR to date. .
18. Yoshizumi Ishino, Sonoko Ishino, Structural basis for substrate recognition and processive cleavage mechanisms of the trimeric exonuclease PhoExo I. , doi: 10.1093/nar/gkv654., 43, 14, 7122-7136, 2015.08, Nucleases play important roles in nucleic acid processes, such as replication, repair and recombination. Recently, we identified a novel single-strand specific 3'-5' exonuclease, PfuExo I, from the hyperthermophilic archaeon Pyrococcus furiosus, which may.
19. Yoshizumi Ishino, Issay Narumi, DNA repair in hyperthermophilic and hyperradioresistant microorganisms., doi: 10.1016/j.mib.2015.05.010., 25, 103-112., 2015.06, The genome of a living cell is continuously under attack by exogenous and endogenous genotoxins. Especially, life at high temperature inflicts additional stress on genomic DNA, and very high rates of potentially mutagenic DNA lesions, including deaminatio.
20. Yoshizumi Ishino, Sonoko Iahino, Takeshi YAMAGAMI, A novel endonuclease that may be responsible for damaged DNA base repair in Pyrococcus furious , doi: 10.1093/nar/gkv121, 43, 5, 2853-2863, 2015.02, DNA is constantly damaged by endogenous and environmental influences. Deaminated adenine (hypoxanthine) tends to pair with cytosine and leads to the A:T→G:C transition mutation during DNA replication. Endonuclease V (EndoV) hydrolyzes the second phosphodiester bond 3 from deoxyinosine in the DNA strand, and was considered to be responsible for hypoxanthine excision repair. However, the downstream pathway after EndoV cleavage remained unclear. The activity to cleave the phosphodiester bond 5 from deoxyinosine was detected in a Pyrococcus furiosus cell extract. The protein encoded by PF1551, obtained from the mass spectrometry analysis of the purified fraction, exhibited the corresponding cleavage activity. A putative homolog from Thermococcus kodakarensis (TK0887) showed the same activity. Further biochemical analyses revealed that the purified PF1551 and TK0887 proteins recognize uracil, xanthine and the AP site, in addition to hypoxanthine. We named this endonuclease Endonuclease Q (EndoQ), as it may be involved in damaged base repair in the Thermococcals of Archaea..
21. Yoshizumi Ishino, Sonoko Iahino, Activation of the MCM helicase from the thermophilic archaeon, Thermoplasma acidophilum by interactions with GINS and Cdc6-2., Springer, DOI 10.1007/s00792-014-0673-6, 18, 915-924, 2014.07, In DNA replication studies, the mechanism for regulation of the various steps from initiation to elongation is a crucial subject to understand cell cycle control. The eukaryotic minichromosome maintenance (MCM) protein complex is recruited to the replication origin by Cdc6 and Cdt1 to form the pre-replication complex, and participates in forming the CMG complex formation with Cdc45 and GINS to work as the active helicase. Intriguingly, Thermoplasma acidophilum, as well as many other archaea, has only one Gins protein homolog, contrary to the heterotetramer of the eukaryotic GINS made of four different proteins. The Gins51 protein reportedly forms a homotetramer (TaGINS) and physically interacts with TaMCM. In addition, TaCdc6-2, one of the two Cdc6/Orc1 homologs in T. acidophilum reportedly stimulates the ATPase and helicase activities of TaMCM in vitro. Here, we found a reaction condition, in which TaGINS stimulated the ATPase and helicase activities of TaMCM in a concentration dependent manner. Furthermore, the stimulation of the TaMCM helicase activity by TaGINS was enhanced by the addition of TaCdc6-2. A gel retardation assay revealed that TaMCM, TaGINS, and TaCdc6-2 form a complex on
ssDNA. However, glutaraldehyde-crosslinking was necessary to detect the shifted band, indicating that the ternary complex of TaMCM–TaGINS–TaCdc6-2 is not stable in vitro. Immunoprecipitation experiment supported a weak interaction of these three proteins in vivo. Activation of the replicative helicase by a mechanism including a Cdc6-like
protein suggests the divergent evolution after the division into Archaea and Eukarya..
22. Yoshizumi Ishino, Sonoko Iahino, Takeshi YAMAGAMI, Multiple interactions of the intrinsically disordered region between the helicase and the nuclease domains of the archaeal Hef protein. , ASBMB, DOI 10.1074/jbc.M114.554998, 289, 31, 21627-21639, 2014.06, Hef is an archaeal protein that probably functions mainly in stalled replication fork repair. The presence of an unstructured region was predicted between the two distinct domains of the Hef protein.Weanalyzed the interdomain region of Thermococcus kodakarensis Hef and demonstrated its disordered structure by CD, NMR, and high speed atomic force microscopy (AFM). To investigate the functions of this intrinsically disordered region (IDR), we screened for proteins interacting with the IDR of Hef by a yeast two-hybrid method, and 10 candidate proteins were obtained. We found that PCNA1 and a RecJ-like protein specifically bind to the IDR in vitro. These results suggested that
the Hef protein interacts with several different proteins that work together in the pathways downstream from stalled replication fork repair by converting the IDR structure depending on the partner protein.
.
23. Kazuo Tori, Sonoko Iahino, Shinichi Kiyonari, Saki Tahara, Yoshizumi Ishino, A novel single-strand specific 3'-5' exonuclease found in the hyperthermophilic archaeon,, Pyrococcus furiosus. , PLoS ONE, 10.1371/journal.pone.0058497, 8, 3, e58497-1-e58497-9, 2013.03, Nucleases play important roles in all DNA transactions, including replication, repair, and recombination. Many different nucleases from bacterial and eukaryotic organisms have been identified and functionally characterized. However, our knowledge about the nucleases from Archaea, the third domain of life, is still limited. We searched for 39–59 exonuclease activity in the hyperthermophilic archaeon, Pyrococcus furiosus, and identified a protein with the target activity. The purified protein, encoded by PF2046, is composed of 229 amino acids with a molecular weight of 25,596, and displayed singlestrand specific 39–59 exonuclease activity. The protein, designated as PfuExo I, forms a stable trimeric complex in solution and excises the DNA at every two nucleotides from the 39 to 59 direction. The amino acid sequence of this protein is conserved only in Thermococci, one of the hyperthermophilic classes in the Euryarchaeota subdomain in Archaea. The newly discovered exonuclease lacks similarity to any other proteins with known function, including hitherto reported 39–59 exonucleases. This novel nuclease may be involved in a DNA repair pathway conserved in the living organisms as a specific member for some hyperthermophilic archaea..
24. Yumani Kuba, Sonoko Iahino, Takeshi YAMAGAMI, Masahiro Tokuhara, Tamotsu Kanai, Ryosuke Fujikane, Hiromi Daiyasu, Haruyuki Atomi, Yoshizumi Ishino, Comparative analyses of the two PCNAs from the hyperthermophilic archaeon, Thermococcus kodakarensis , Genes to Cells , 10.1111/gtc.12007, 17, 11, 923-937, 2012.11, The DNA sliding clamp is a multifunctional protein involved in cellular DNA transactions. In Archaea and Eukaryota, proliferating cell nuclear antigen (PCNA) is the sliding clamp. The ring-shaped PCNA encircles double-stranded DNA within its central hole and tethers other proteins on DNA. The majority of Crenarchaeota, a subdomain of Archaea, have multiple PCNA homologues, and they are capable of forming heterotrimeric rings for their functions. In contrast, most organisms in Euryarchaeota, the other major subdomain, have a single PCNA forming a homotrimeric ring structure. Among the Euryarchaeota whose genome is sequenced,
Thermococcus kodakarensis is the only species with two genes encoding PCNA homologues on its genome. We cloned the two genes from the T. kodakarensis genome, and the gene products, PCNA1 and PCNA2, were characterized. PCNA1 stimulated the DNA synthesis reactions of the two DNA polymerases, PolB and PolD, from T. kodakarensis in vitro. PCNA2, however, only had an effect on PolB. We were able to disrupt the gene for PCNA2, whereas gene disruption for PCNA1 was not possible, suggesting that PCNA1 is essential for DNA replication. The sensitivities of the Dpcna2 mutant strain to ultraviolet irradiation (UV), methyl methanesulfonate
(MMS) and mitomycin C (MMC) were indistinguishable from those of the wild-type strain..
25. ISHINO Yoshizumi and ISHINO Sonoko, Rapid progress of DNA replication studies in Archaea, the third domain of life., SCIENCE CHINA  Life Sciences, doi: 10.1007/s11427-012-4324-9, 55, 1-18, 2012.05, Archaea, the third domain of life, are interesting organisms to study from the aspects of molecular and evolutionary biology. Archaeal cells have a unicellular ultrastructure without a nucleus, resembling bacterial cells, but the proteins involved in genetic information processing pathways, including DNA replication, transcription, and translation, share strong similarities with those of Eukaryota. Therefore, archaea provide useful model systems to understand the more complex mechanisms of genetic information processing in eukaryotic cells. Moreover, the hyperthermophilic archaea provide very stable proteins, which are especially useful for the isolation of replisomal multicomplexes, to analyze their structures and functions. This review focuses on the history, current status, and future directions of archaeal DNA replication studies..
26. Ishino, S., Kawamura, A., and Ishino, Y., Application of PCNA to processive PCR by reducing the stability of its ring structure. , J. Jap. Soc. Extremophiles. , 11, 2, 印刷中, 2012.05.
27. Ishino, S., Fujino, S., Tomita, H., Ogino, H., Takao, K., Daiyasu, H., Kanai, T., Atomi, H., and Ishino, Y., Biochemical and genetical analyses of the three mcm genes from the hyperthermophilic archaeon, Thermococcus kodakarensis. , Genes to Cells, 10.1111/j.1365-2443.2011.01562.x., 16, 12, 1176-1189, 2011.11, In eukaryotes, the replicative DNA helicase 'core' is the minichromosome maintenance (Mcm) complex (MCM), forming a heterohexameric complex consisting of six subunits (Mcm2-7). Recent studies showed that the CMG (Cdc45-MCM-GINS) complex is the actual helicase body in the replication fork progression complex. In Archaea, Thermococcus kodakarensis harbors three genes encoding the Mcm homologs on its genome, contrary to most archaea, which have only one homolog. It is thus, of high interest, whether and how these three Mcms share their functions in DNA metabolism in this hyperthermophile. Here, we report the biochemical properties of two of these proteins, TkoMcm1 and TkoMcm3. In addition, their physical and functional interactions with GINS, possibly an essential factor for the initiation and elongation process of DNA replication, are presented through in vitro ATPase and helicase assays, and an in vivo immunoprecipitation assay. Gene disruption and product quantification analyses suggested that TkoMcm3 is essential for cell growth and plays a key role as the main DNA helicase in DNA replication, whereas TkoMcm1 also shares some function in the cells..
28. Oyama T, Ishino S, Fujino S, Ogino H, Shirai T, Mayanagui K, Saito M, Nagasawa N, Ishino Y, Morikawa K., Architectures of archaeal GINS complexes, essential DNA replication initiation factors, BMC Biol., doi:10.1186/1741-7007-9-28, 9, 28, 2011.04.
29. Fujikane, R., Ishino, S., Ishino, Y., Forterre, P., Genetic analysis of DNA repair in the hyperthermophilic archaeon, Thermococcus kodakaraensis. , Genes Genet. Syst. , 10.1266/ggs.85.243, 85, , 243-257, 2010.08, Extensive biochemical and structural analyses have been performed on the putative DNA repair proteins of hyperthermophilic archaea, in contrast to the few genetic analyses of the genes encoding these proteins.  Accordingly, little is known about the repair pathways used by archaeal cells at high temperature.  Here, we attempted to disrupt the genes encoding the potential repair proteins in the genome of the hyperthermophilic archaeon Thermococcus kodakaraensis.  We succeeded in isolating null mutants of the hjc, hef, hjm, xpb, and xpd genes, but not the radA, mre11, rad50, herA, nurA, and xpg/fen1 genes.  Phenotypic analyses of the gene-disrupted strains showed that the xpb and xpd null mutants are only slightly sensitive to UV, methylmethane sulfonate and mitomycin C, as compared with the wild-type strain.  The hjm null mutant showed sensitivity specifically to mitomycin C.  On the other hand, the null mutants of the hjc gene lacked increasing sensitivity to any type of DNA damage.  The Hef protein is particularly important for maintaining genome homeostasis, by functioning in the repair of a wide variety of DNA damage in T. kodakaraensis cells.  Deletion of the entire hef gene or of the segments encoding either its nuclease or helicase domain produced similar phenotypes.  The high sensitivity of the Δhef mutants to mitomycin C suggests that Hef performs a critical function in the repair process of DNA interstrand cross-links.  These damage-sensitivity profiles suggest that the archaeal DNA repair system has processes depending on repair-related proteins different from those of eukaryotic and bacterial DNA repair systems using homologous repair proteins analyzed here..
30. Mayanagi, K., Kiyonari, S., Nishida, H., Saito, M., Kohda, D., Ishino, Y., Shirai, T., Morikawa, K. , The architecture of the DNA polymerase-PCNA-DNA ternary complex. , Proc. Natl. Acad. Sci. USA. , 10.1073/pnas.1010933108 , 107, , 5, 1845-1849 , 2011.02, DNA replication in archaea and eukaryotes is executed by family B DNA polymerases, which exhibit full activity when complexed with the DNA clamp, proliferating cell nuclear antigen (PCNA). This replication enzyme consists of the polymerase and exonuclease moieties responsible for DNA synthesis and editing (proofreading), respectively. Due to the editing activity, this enzyme ensures the high fidelity of DNA replication. However, it remains unclear how the PCNA-complexed enzyme temporally switches between the polymerizing and editing modes. Here, we present the threedimensional structure of the Pyrococcus furiosus DNA polymerase B-PCNA-DNA ternary complex, which is the core component of the replisome, determined by single particle electron microscopy of negatively stained samples. This structural view, representing the complex in the editing mode, revealed the whole domain configuration of the trimeric PCNA ring and the DNA polymerase, including protein-protein and protein-DNA contacts. Notably, besides the authentic DNA polymerase-PCNA interaction through a PCNA-interacting protein box (PIP-box), a novel contact was found between DNA polymerase and the PCNA subunit adjacent to that with the PIP contact. This contact appears to be responsible for the configuration of the complex
specific for the editing mode. The DNA was located almost at the center of PCNA, and exhibited a substantial and particular tilt angle against the PCNA ring plane. The obtained molecular architecture of the complex, including the new contact found in this work, provides clearer insights into the switching mechanism between the two distinct
modes, thus highlighting the functional significance of PCNA in the replication process..
31. Akita, M., Adachi, A, Takemura, K., Yamagami, T., Matsunaga, F., and Ishino, Y., Cdc6/Orc1 from Pyrococcus furiosus may act as the origin recognition protein and Mcm helicase recruiter. , Gens to Cells, 10.1111/j.1365-2443.2010.01402.x, 15, 5, 537-552, 2010.05.
32. 石野園子、石野良純, 耐熱性クランプ分子:環状構造の安定性と機能の関係, 生化学, 81, 12, 1056-1063., 2009.12.
33. Nishida, H., Mayanagi, K., Kiyonari, S., Sato, Y., Oyama, T., Ishino, Y., and Morikawa, K., Structural determinant for switching between the polymerase and exonuclease modes in the PCNA-replicative DNA polymerase complex., Proc. Natl. Acad. Sci. USA. , 106, 49, 20693-20698, 2009.12.
34. Kiyonari, S., Tahara, S., Shirai, T., Iwai, S., Ishino, S., and Ishino, Y., Biochemical properties and BER complex formation of AP endonuclease from Pyrococcus furiosus, Nucleic Acids Res., 37, 19, 6439-6453, 2009.09.
35. 石野良純, 特集にあたって, 蛋白質核酸酵素, 54, 101-107, 2009.08.
36. 石野園子、石野良純, アーキアのDNAトランスアクション〜その共通性と多様性〜 , 蛋白質核酸酵素, 54, 141-147, 2009.02.
37. Matsukawa H, Yamagami T, Kawarabayasi Y, Miyashita Y, Takahashi M, Ishino Y., A useful strategy to construct DNA polymerases with different properties by using genetic resources from environmental DNA, Genes Genet. Syst., 84, 3-13, 2009.02.
38. Oyama T, Oka H, Mayanagi K, Shirai T, Matoba K, Fujikane R, Ishino Y, Morikawa K, Atomic structures and functional implications of the archaeal RecQ-like helicase Hjm., BMC Struct. Biol., 9, 2 (1-12), 2009.03.
39. Mayanagi. K., Kiyonari, S., Saito, M., Shirai, T., Ishino, Y., and Morikawa, K., Mechanism of replication machinery assembly as revealed by the DNA ligase-PCNA-DNA complex architechture., Proc. Natl. Acad. Sci. USA., 106, 4657-4652 , 2009.03.
40. 石野良純, DNA複製フォーク進行停止とその修復機構, RADIOISOTOPES, 557, 467-470 (2008), 2008.10.
41. S. Kiyonari, S. Tahara, M. Uchimura, T. Shirai, S. Ishino, and Yoshizumi Ishino,, Studies on the base excision repair (BER) complex in Pyrococcus furiosus., Biochem. Soc. Transact., in press, 2008.12.
42. S . Kiyonari, M. Uchimura, T. Shirai, and Yoshizumi Ishino,, Physical and Functional Interactions between Uracil-DNA glycosylase and proliferating cell nuclear antigen from the euryarchaeon Pyrococcus furiosus., J. Biol. Chem., 283, No35, 24185-24193, 2008.08.
43. Yoshimochi, T., Fujikane, R., Kawanami, M., Matsunaga, F., and Ishino, Y.*, The GINS complex from Pyrococcus furiosus stimulates the MCM helicase activity., Journal of Biological Chemistry, 283, No3, 1601-1609, 2008.01.
44. 清成信一、石野良純, DNAリガーゼの構造と機能〜アーキアからの発見〜, 化学と生物, 45,705-711(2007), 2007.10.
45. Tori, K., Kimizu, M., Ishino, S., and Ishino. Y. , Both DNA polymerase BI and D from the hyperthermophilic archaeon, Pyrococcus furiosus bind to PCNA at the C-terminal PIP box motifs. , Journal of Bacteriology, 189, 5652-5657, 2007.05.
46. Kiyonari, S., Kamigochi, T., and Ishino, Y. , A single amino acid substitution in the DNA-binding domain of Aeropyrum pernix DNA ligase impairs its interaction with proliferating cell nuclear antigen, Extremophiles, 11, 675−684, 2007.05.
47. Matsunaga, F., Glatigny, A., Mucchielli-Giorgi, M. H., Agier, N., Delacroix, H., Marisa, M., Durosay, P., Ishino, Y., Aggerbeck, L., and Forterre, P. , Genomewide and Biochemical Analyses of DNA-binding activity of Cdc6/Orc1 and Mcm proteins in Pyrococcus sp. , Nucleic Acids Res. , 35, 3214-3222., 2007.04.
48. Imamura, K., Fukunaga, K., Kawarabayasi, Y., and Ishino, Y., Specific interactions of three PCNAs with replication-related proteins in Aeropyrum pernix., Molecular Microbiology , 64, 308-318, 2007.01.
49. Kiyonari, S., Takayama, K., Nishida, H., Ishino, Y., Identification of a novel binding motif in Pyrococcus furiosus DNA ligase for the functional interaction with proliferating cell nuclear antigen., J. Biol. Chem., 281 巻 28023-28032ページ, 2006.07.
50. Miyata, T., Suzuki, H., Oyama, T., Mayanagi, K., Ishino, Y., and Morikawa, K., Open clamp structure in the clamp-loading complex visualized by electron microscopic image analysis., Proc. Natl. Acad. Sci. USA, 10.1073/pnas.0506447102, 102, 39, 13795-13800, 102, 13795-13800, 2005.09.
51. Fujikane, R., Komori, K., Shinagawa, H., and Ishino, Y., Identification of a novel helicase activity unwinding branched DNAs from the hyperthermophilic archaeon, Pyrococcus furiosus., J. Biol. Chem., 10.1074/jbc.M413417200, 280, 13, 12351-12358, 280, 12351-12358., 2005.04.
主要総説, 論評, 解説, 書評, 報告書等
1. 石野良純, 遺伝子工学技術はDNA関連酵素に支えられている~制限酵素、PCRそしてゲノム編集へ~, 生物工学会誌, 2017.07, 20世紀後半にはじまった分子生物学の急速な発展は、試験管内で人工的にDNAを切り貼りしたものを生きた細胞に導入する遺伝子組換え実験ができるようになったことに大きく依存している。この技術を使って実験を成功させるためには少し熟練を必要としたが、この技術誕生の15年後、PCR の登場によって試験管内遺伝子操作が画期的に容易になった。PCRは分子生物学の各種実験手法を一変させ、煩雑な操作が大きく簡略化されたことを、筆者の世代の分子生物学者は実感している。そして、PCRの登場から15年後、新たな遺伝子操作実験手法として、CRISPR-Cas9を利用した簡便な「ゲノム編集」技術が開発され、それから数年を経た現在、ゲム編集実験は急速にその広がりを見せている。これらの遺伝子工学実験技術を支えているのは、DNAに作用する酵素であり、生物が自分の生命活動を維持するために有しているDNA関連酵素をうまく利用することによって、人工的にDNA鎖を合成したり、切断したり、連結したり、二本鎖を一本鎖に解くことが可能になった。本稿では、重要な遺伝子工学技術開発に繋がったDNA関連酵素に焦点を当てて、その歴史を辿ってみたい。.
2. Ishino, Y., Krupovic, M., Forterre, P. , History of CRISPR-Cas from Encounter with a Mysterious Repeated Sequence to Genome Editing Technology. , Journal of Bacteriology, doi: 10.1128/JB.00580-17., 2018.03, Clustered regularly interspaced short palindromic repeat (CRISPR)-Cas systems are well-known acquired immunity systems that are widespread in archaea and bacteria. The RNA-guided nucleases from CRISPR-Cas systems are currently regarded as the most reliable tools for genome editing and engineering. The first hint of their existence came in 1987, when an unusual repetitive DNA sequence, which subsequently was defined as a CRISPR, was discovered in the Escherichia coli genome during an analysis of genes involved in phosphate metabolism. Similar sequence patterns were then reported in a range of other bacteria as well as in halophilic archaea, suggesting an important role for such evolutionarily conserved clusters of repeated sequences. A critical step toward functional characterization of the CRISPR-Cas systems was the recognition of a link between CRISPRs and the associated Cas proteins, which were initially hypothesized to be involved in DNA repair in hyperthermophilic archaea. Comparative genomics, structural biology, and advanced biochemistry could then work hand in hand, not only culminating in the explosion of genome editing tools based on CRISPR-Cas9 and other class II CRISPR-Cas systems but also providing insights into the origin and evolution of this system from mobile genetic elements denoted casposons. To celebrate the 30th anniversary of the discovery of CRISPR, this minireview briefly discusses the fascinating history of CRISPR-Cas systems, from the original observation of an enigmatic sequence in E. coli to genome editing in humans.
KEYWORDS:

.
3. Yoshizumi Ishino, Sonoko Iahino, DNA polymerases as useful reagents for biotechnology - the history of developmental research in the field. , 2014.08, DNA polymerase is a ubiquitous enzyme that synthesizes complementary DNA strands according to the template DNA in the living cells. Multiple enzymes have been identified from each organism, and the shared functions of these enzymes have been investigated. In addition to their fundamental role in maintaining genome integrity during replication and repair, DNA polymerases are widely used for DNA manipulation in vitro, including DNA cloning, sequencing, labeling, mutagenesis, and other experiments. The fundamental ability of DNA polymerases to synthesize a deoxyribonucleotide chain is conserved. However, the more specific properties, including processivity, fidelity (synthesis accuracy), and substrate nucleotide selectivity, differ among the enzymes. The distinctive properties of each DNA polymerase may lead to the potential development of unique reagents, and therefore searching for novel DNA polymerase has been one of the major focuses in this research field. In addition, protein engineering techniques to create mutant or artificial DNA polymerases have been successfully developing powerful DNA polymerases, suitable for specific purposes among the many kinds of DNA manipulations. Thermostable DNA polymerases are especially important for PCR-related techniques in molecular biology. In this review, we summarize the history of the research on developing thermostable DNA polymerases as reagents for genetic manipulation and discuss the future of this research field..
4. Yoshizumi Ishino, Sonoko Iahino, The archaeal DNA replication machinery: past, present and future. , Genes and Genetic Systems, 2013.12, Living organisms are divided into three domains: Bacteria, Archaea, and Eukarya. Whereas Bacteria and Archaea are both prokaryotes, proteins involved in information processes; replication, transcription, and translation, are more similar in Archaea and Eukarya. Here the history of the research on archaeal DNA replication is summarized and the future of the field is discussed..
5. Ishino, Y. and Ishino, S., Rapid progress of DNA replication studies in Archaea, the third domain of life. , Sci. China, Ser-C Life Sci. , 2012.05, Archaea, the third domain of life, is a very interesting living organism to study from the aspects of molecular and evolutional biology. Archaeal cells have a unicellular ultrastructure without a nucleus, resembling bacterial cells, but the, proteins involved in the genetic information processing pathways, including DNA replication, transcription, and translation, share strong similarities with those of Eukaryota. Therefore, the archaeal processes provide useful model systems to understand the more complex mechanisms of genetic information processing in the eukaryotic cells. Moreover, the hyperthermophilic archaea provide very stable proteins, which are especially useful for the isolation of replisomal multicomplexes, to analyze their structures and functions. This review focuses on the history, current status, and future directions of archaeal DNA replication studies..
6. Ishino, S and Ishino, Y., Application of environmental DNA resources to create useful DNA polymerases with different properties., Microorganisms in Sustainable Agriculture and Biotechnology. (Springer Science+Business Media), 2012.01, DNA polymerases use deoxynucleotide triphosphates to synthesize new DNA strands according to the template DNA during DNA replication and repair, and are essential to maintain genome integrity in DNA metabolism. In addition, these enzymes are widely used for genetic engineering techniques, including dideoxy-sequencing, PCR, DNA labeling, mutagenesis, and other in vitro gene manipulations. Thermostable DNA polymerases are especially useful for PCR and cycle-sequencing. We describe in this chapter a powerful strategy to use environmental DNA as a genetic resource to create useful DNA polymerases. The region corresponding to the active center of the DNA polymerizing reaction in the structural gene of well known DNA polymerases, such as Pfu DNA polymerase and Taq DNA polymerase, can be substituted with gene fragments amplified by PCR from DNAs within soil samples from various world-wide locations. The constructed chimeric pol genes can be expressed in E. coli, and the produced chimeric enzymes, possessing DNA polymerase activities with different properties, can be evaluated in terms of their processivity, fidelity, and efficiency of primer usage, to select valuable DNA polymerases for genetic engineering techniques..
7. 石野園子、石野良純 , 耐熱性クランプ分子:環状構造の安定性と機能の関係, 生化学, 2009.12.
8. 石野良純, アーキア:第3の不思議な生物 特集にあたって , 蛋白質核酸酵素 54, 101-107, 2009.02.
9. 石野園子、石野良純, アーキアのDNAトランスアクション 〜その共通性と多様性〜, 蛋白質核酸酵素 54, 141-147, 2009.02.
10. 石野良純、山上 健、松川博昭、鬼塚尚子、鍋 健吾、興梠聖哉, メタゲノムを利用した新規DNA合成酵素の創製, 環境バイオテクノロジー学会誌, Vol 7, No.2, 87-92, 2007.07.
主要学会発表等
1. Sonoko Ishino, Yumani Kuba, Takeshi Yamagami, Masahiro Tokuhara, Tamotsu Kanai, Haruyuki Atomi, and Yoshizumi Ishino, NA replication proteins from Thermococcus kodakarensis., Gordon Research Conference on Archaea: Ecology, Metabolism & Molecular Biology, 2011.08.
2. Yoshizumi Ishino, DNA replication in Thermococcales -from initiation to elongation-, Gordon Research Conference on Archaea: Ecology, Metabolism & Molecular Biology, 2011.08.
3. 石野良純, アーキアにおけるDNA複製フォーク進行複合体の構造と機能, 第84回日本生化学会大会シンポジウム3S4a 「アーキア研究の最前線」, 2011.09.
4. 亀丸美里、山上 健、石野園子、石野良純, Thermococcus kodakarensisのDNAポリメラーゼDに含まれる天然変性領域に注目した構造・機能解析, 第12回極限環境生物学会年会, 2011.11.
5. Sonoko Ishino, Takeshi Yamagami, Makoto Kitamura, Yoshizumi Ishino, Two nucleases from Thermococcus kodakarensis, probably involved in replication fork progression., 第34回日本分子生物学会年会, 2011.12.
6. 石野良純, DNA複製フォーク進行停止の修復関連タンパク質に存在する天然変性領域について, 日本農芸化学会2012年度大会 シンポジウム 4SY23 「天然変性領域を介した相互作用に基づくタンパク質応用化学の新展開」, 2012.03.
7. 石野良純 , DNA 複製フォーク進行停止の修復に関わる天然変性蛋白質, 第10回日本蛋白質科学会 , 2010.06.
8. 石野良純,  DNA修復タンパク質 Hef 〜その発見から現在まで〜, 日本Archaea研究会第23回講演会 , 2010.07.
9. 石野良純, 超好熱性アーキアの複製フォーク進行と停止修復に関わる超分子複合体の構造と機能, ワークショップ「染色体複製とその制御における超高次複合体のダイナミクス」  第33回日本分子生物学会年会第83回日本生化学会大会合同大会 2010.12.7-10. 神戸ポートアイランド,神戸 , 2010.12.
10. Yoshizumi Ishino, Structural and functional analyses of a DNA repair protein Hef containing a ID region between the helicase and endonuclease domains. , The 1st International Symposium on Intrinsically disordered proteins, 2011.01.
11. 石野良純, DNA複製フォーク進行に関与するタンパク質の天然変性領域の役割解明を目指して, 「天然変性タンパク質の分子認識機構と機能発現」第1回公開シンポジウム, 2010.01.
12. 向井彩子、石野園子、佐藤 大、木野邦器、石野良純, 耐熱性ATP再生系のDNA組換え反応への応用, 第10回極限環境微生物学会年会, 2010.05.
13. 石野良純, Stability of the ring structure and its effect on the processive DNA strand synthesis構造解析から見えてきたアーキアPCNAのリング形成特性, 第82回日本生化学会大会, 2009.10.
14. Yoshizumi Ishino, Stability of the ring structure of PCNA and its effect on the in vitro DNA strand synthesis., Thermophiles 2009, 2009.08.
15. Sonoko Ishino, Eriko Nasu, Hiroaki Matsukawa, Maiko Uchimura, Shinichi Kiyonari, Kaori Imamura, Yoshizumi Ishino, Detection of the functional PCNA ring of Aeropyrum pernix based on analyzing interface between subunits and isolating clamp-loading complex., Thermophiles 2009, 2009.08.
16. Shinichi Kiyonari, Saki Tahara, Tsuyoshi Shirai, Shigenori Iwai, Sonoko Ishino, and Yoshizumi Ishino , Studies on interactions of PCNA-PCNA binding proteins in Archaea 〜Detection of the base excision repair (BER) complex in Pyrococcus furiosus〜 , Gordon Research Conference  Archaea: Ecology, Metabolism & Molecular Biology, 2009.07.
17. 石野良純, PCR 関連分子の機能改変〜キメラ DNA ポリメラーゼとクランプ分子工学〜, 日本農芸化学会2009年度大会, 2009.03.
18. 石野良純、清成信一、西田洋一、真柳浩太、白井 剛、森川耿右,  構造生物学から見たアーキア DNA 鎖伸長機構 , 日本生化学会九州支部例会 , 2008.05.
19. 石野園子、河村 啓、石野良純, (2008) 超好熱性アーキアPyrococcus furiosus由来 PCNAの機能解析からPCNA工学へ, 第31回日本分子生物学会年会 第81回日本生化学会大会 合同大会, 2008.12.
20. 石野良純, アーキア分子生物学の醍醐味 〜DNAポリメラーゼから複製、そして組換え修復機構研究へ 

日本アーキア研究会第21回講演会, 日本アーキア研究会第21回講演会, 2008.07.
21. Ishino Y. , Contribution of the proteins from hyperthermophilic archaea to structural and functional analyses of DNA replication/repair apparatus., Extremophiles 2008, 2008.09.
特許出願・取得
特許出願件数  10件
特許登録件数  3件
学会活動
所属学会名
日本アーキア研究会
American Society for Microbiology
極限環境微生物学会(International Society for Extremophiles)
日本農芸化学会
蛋白質科学会
日本遺伝学会
日本薬学会
日本生化学会
日本分子生物学会
学協会役員等への就任
2010.03~2010.09, Extremophiles 2010, Scientific Advisary board member.
2009.01~2009.08, Thermophiles 2009, INTERNATIONAL ADVISORY COMMITTEE.
2009.03~2011.03, 日本農芸化学会, 評議員.
2006.07, 日本アーキア研究会, 幹事.
2005.01, 日本生化学会, 「生化学」企画委員会委員.
2003.01, 極限環境微生物学会, 編集委員会副委員長.
2006.01, 極限環境微生物学会, 幹事.
学会大会・会議・シンポジウム等における役割
2012.03.22~2012.03.25, 農芸化学会 2012 年度大会, シンポジウム 4SY23 「天然変性領域を介した相互作用に基づくタンパク質応用化学の新展開」 企画、オーガナイザー(司会、座長を含む).
2011.09.21~2011.09.24, 第84回日本生化学会大会, シンポジウム3S4a 「アーキア研究の最前線」 企画、オーガナイザー (司会、座長を含む) .
2011.03.25~2011.03.28, 農芸化学会 2011 年度大会, シンポジウム 「核酸を標的とする微生物酵素の化学〜多様な分子機構の解明から応用への展開」提案、オーガナイザー (司会、座長を含む).
2009.10.21~2009.10.24, 第82回日本生化学会大会, シンポジウム 4S14a 「高次タンパク質複合体構造の変遷によるDNA複製の進行制御」 企画 オーガナイザー 、座長、司会も含む  .
2009.08.16~2009.08.20, Thermophiles 2009, インターナショナル サイエンティフィック アドバイザリーボード (演題査読、座長を含む).
2008.09.07~2008.09.11, Extremophiles 2008, インターナショナル サイエンティフィック アドバイザリーボード.
2008.12.07~2008.12.10, 第31回日本分子生物学会年会・第81回日本生化学会大会 合同大会, シンポジウム 4S17 「アーキアゲノム生物学が与える生命科学研究へのインパクト」 企画 オーガナイザー.
2008.05.21~2008.05.22, 日本生化学会九州支部例会 シンポジウム「構造解析を基にしてDNA複製の分子機構を解く」, シンポジウム企画 オーガナイザー.
2007.11.27~2007.11.28, 第8回極限環境微生物学会年会, オーガナイザー.
2006.08.02~2006.08.03, 日本 Archaea研究会 第 19 回後援会 , オーガナイザー.
2004.09, 日本遺伝学会第76回大会, 座長(Chairmanship).
2003.09, 日本遺伝学会第75回大会, 座長(Chairmanship).
2014.09.07~2014.09.11, Extremophiles 2015, Invited Speaker (招待講演者).
2014.05.19~2014.05.22, Molecular Biology of Archaea 4, Invited Speaker (招待講演者).
2014.11.24~2014.11.26, 第37回日本分子生物学会年会ワークショップ 「生命の3大ドメインの分子生物学から考察する遺伝情報制御系の原型」 「Prototype of gene-regulatory machineries based on the molecular biology in the three domains of life」, ワークショップ オーガナイザー.
2005.07, The XIIIth International Congress of Bacteriology and Applied Microbiology, オーガナイザー.
2004.12, 第27回分子生物学会ワークショップ「DNA複製フォーク進行阻害の回避・回復の分子機構 」, オーガナイザー.
2004.09, 第76回日本遺伝学会シンポジウム「ゲノムの安定性維持機構ー翻訳後修飾の関わり」, オーガナイザー.
2002.10, 第75回日本生化学会シンポジウム「古細菌の生化学」, オーガナイザー.
2001.12, 第24回分子生物学ワークショップ「古細菌の分子生物学」, オーガナイザー.
学会誌・雑誌・著書の編集への参加状況
2010.01~2020.12, Archaea, 国際, 編集委員.
2005.04, 生化学会/生化学, 国内, 編集委員.
2003.04, 極限環境微生物学会誌, 国内, 編集委員.
学術論文等の審査
年度 外国語雑誌査読論文数 日本語雑誌査読論文数 国際会議録査読論文数 国内会議録査読論文数 合計
2012年度 10      13 
2011年度 10  12 
2010年度 20    25 
2009年度 20  10    33 
2008年度    
2007年度 12      15 
2006年度 10      18 
2005年度        
2004年度 110  115 
2003年度
2002年度
その他の研究活動
海外渡航状況, 海外での教育研究歴
Lucca (Gordon Research Conference), Italy, 2013.07~2013.08.
University of Regensburg (Thermophiles 2013), Germany, 2013.09~2013.09.
Institute pasteur (Molecular Biology of Archaea 4), France, 2014.05~2014.05.
Saint Petersburg (Extremophiles 2014), Russia, 2014.09~2014.09.
Northwest A6F University , China, 2014.09~2014.09.
University of Illinois, Urbana Champaign, UnitedStatesofAmerica, 2015.09~2015.09.
Yale University, UnitedStatesofAmerica, 2015.09~2015.09.
University of Maryland, UnitedStatesofAmerica, 2015.09~2015.09.
University of Santiago de Chile (Thermophiles 2015), Chile, 2015.08~2015.09.
KAUST, Saudi Arabia, 2014.11~2014.12.
Institute Pasteur, France, 2015.02~2015.02.
Institute pasteur, France, 2015.10~2016.03.
University of Illinois, UnitedStatesofAmerica, 2003.08~2003.08.
University of Maryland, UnitedStatesofAmerica, 2003.08~2003.08.
New England Biolabs, UnitedStatesofAmerica, 2002.08~2002.08.
New England Biolabs, UnitedStatesofAmerica, 2000.08~2000.08.
University of Paris sud ( パリ南大学), France, UnitedStatesofAmerica, UnitedStatesofAmerica, 2000.08~2000.09.
University of Maryland, UnitedStatesofAmerica, 1998.01~1998.02.
University of illinois, UnitedStatesofAmerica, 1998.01~1998.02.
New England Biolabs, UnitedStatesofAmerica, 1998.01~1998.02.
外国人研究者等の受入れ状況
2014.10~2014.10, Christian-Albrechts-Universität zu Kiel , Germany.
2014.03~2014.04, 2週間以上1ヶ月未満, University of Bergen, Norway, Norway, 日本学術振興会.
2013.08~2013.09, 2週間未満, University of Vienna, Austria, 日本学術振興会.
2013.06~2013.07, 1ヶ月以上, National Institute of Standards and Technology (NIST) and the Institute for Bioscience and Biotechnology Research (IBBR)、 University of Maryland, UnitedStatesofAmerica, 日本学術振興会.
2011.06~2011.07, 1ヶ月以上, Ohio State University, UnitedKingdom, 日本学術振興会.
2010.03~2010.03, New England Biolabs Inc., UnitedStatesofAmerica, 学内資金.
2009.03~2009.03, 2週間未満, New England Biolabs Inc., UnitedStatesofAmerica, 学内資金.
2008.03~2008.03, 2週間未満, New England Biolabs Inc., UnitedStatesofAmerica, 学内資金.
2006.08, 1ヶ月以上, フランス パリ南大学, France, 日本学術振興会.
2005.10, 2週間以上1ヶ月未満, フランス パリ南大学, France, 外国政府・外国研究機関・国際機関.
受賞
九州大学農学研究院賞(第6回), 農学研究院, 2018.03.
平成30年度 農芸化学会賞, 日本農芸化学会, 2018.03.
日本医療研究開発大賞, 健康・医療戦略推進本部, 2017.12.
ポスター賞, 極限環境微生物学会, 2003.12.
研究資金
科学研究費補助金の採択状況(文部科学省、日本学術振興会)
2010年度~2014年度, 新学術領域研究, 代表, 核内ネットワークを制御する天然変性タンパク質の機能発現(計画代表).
2011年度~2013年度, 基盤研究(B), 代表, 超好熱生物の遺伝情報はどのように守られ、伝達されるか?.
2009年度~2014年度, 新学術領域研究, 代表, 核内ネットワークを制御する天然変性タンパク質の機能発現.
2006年度~2010年度, 特別研究, 分担, 特定領域「染色体サイクルの制御ネットワーク」

複製フォーク複合体の構築、維持、変換の研究.
科学研究費補助金の採択状況(文部科学省、日本学術振興会以外)
2008年度~2010年度, 科学技術振興機構バイオインフォーマティクス推進事業, 分担, 実践による超分子ネットワークモデリングシステムの開発.
2005年度~2008年度, , 分担, 実践による超分子複合体モデリングシステムの開発.
競争的資金(受託研究を含む)の採択状況
2011年度~2016年度, , 代表, 科学技術振興機構 (JST)
CREST 研究 「海洋生物多様性および生態系の保全・再生に資する基盤技術の創出」
「Digital DNA chip による生物多様性評価と環境予測法の開発」 (代表 五條堀 孝)の機関代表.
2011年度~2015年度, 農林水産技術会議プロジェクト研究, 代表, 農林水産技術会議委託プロジェクト研究「海洋微生物解析による沿岸漁業被害の予測、抑制技術の開発」.
2009年度~2010年度, 科学技術振興機構 バイオインフォマティクス推進センター事業, 分担, 実践による超分子ネットワークモデリングシステムの開発.
2006年度~2008年度, 科学技術振興機構 バイオインフォマティクス推進センター事業, 分担, 実践による超分子モデリングシステムの開発.
共同研究、受託研究(競争的資金を除く)の受入状況
2005.04~2006.03, 代表, 新規なFEN用補助因子.
寄附金の受入状況
2007年度, 財団法人 篷庵社, 研究助成金/複製フォークを正しく進行させるためのDNA 複製と協調した修復機構.
2006年度, 財団法人 篷庵社, 研究助成金/複製フォークを正しく進行させるためのDNA 複製と協調した修復機構.
2005年度, 財団法人 篷庵社, 研究助成金/複製フォークを正しく進行させるためのDNA 複製と協調した修復機構.
2004年度, 財団法人 篷庵社, 研究助成金/複製フォークを正しく進行させるためのDNA 複製と協調した修復機.
学内資金・基金等への採択状況
2010年度~2011年度, 九州大学教育研究プログラム・研究拠点形成プロジェクト, 分担, ネパールの食品および環境中の遺伝資源の高度有効利用

.
2010年度~2011年度, 九州大学教育研究プログラム・研究拠点形成プロジェクト, 分担, ネパールの食品および環境中の遺伝資源の高度有効利用

.
2009年度~2010年度, 九州大学教育研究プログラム・研究拠点形成プロジェクト , 分担, 超高次複合体解析に基づくゲノム動態研究プロジェクト.
2006年度~2007年度, 九州大学教育研究プログラム・研究拠点形成プロジェクト, 分担, DNA複製研究の次世代育成プログラム.

九大関連コンテンツ

pure2017年10月2日から、「九州大学研究者情報」を補完するデータベースとして、Elsevier社の「Pure」による研究業績の公開を開始しました。
 
 
九州大学知的財産本部「九州大学Seeds集」