Research Projects

• Grant-in-Aid for Scientific Research (C), Apr. 2022 - Mar. 2026, 22K05532, Establishment of the technological basis for color enhancement of red wine by yeast, 吉崎 隆之, Japan Society for the Promotion of Science, Grants-in-Aid for Scientific Research Grant-in-Aid for Scientific Research (C), Fukuyama University, 4030000, 3100000, 930000, kaken

  対象リソースIRI
• Grant-in-Aid for Scientific Research (C), 2010 - 2012, 22570020, Adaptive significance of dwarf males in deep-sea barnacles, YUSA Yoichi; IWAGUCHI Shinichi, Japan Society for the Promotion of Science, Grants-in-Aid for Scientific Research Grant-in-Aid for Scientific Research (C), Nara Women's University, 4550000, 3500000, 1050000, The adaptive significance of dwarf males is addressed in deep-sea barnacles. First, microsatellite markers were developed in Scalpellum stearnsii, and the reproductive success of their dwarf males was studied. Small males attained higher reproductive success than larger ones. Second, life history traits were studied and some pieces of information were obtained in this species and in a related species, S. scalpellum, in situ and/or in the aquarium. Third, using mathematical models and theoretical synthesis by reviewing relevant studies, it was suggested that dwarf males evolved under low-density conditions., url

  対象リソースIRI
• Grant-in-Aid for Challenging Exploratory Research, 2010 - 2012, 22659086, Development of early diagnostic method for Deep mycoses by monitoring volatile organic compounds, IWAGUCHI Shin-ichi; TAKEUCHI Takae; SUZUKI Takahito; YOKOYAMA Koji, Japan Society for the Promotion of Science, Grants-in-Aid for Scientific Research Grant-in-Aid for Challenging Exploratory Research, Nara Women's University, 3060000, 2700000, 360000, We evaluated the availability of VOCs (volatile organic compounds) as the biomarker during the pulmonary invasive aspergillosis in murine infection model. We established the murine infection model and developed the apparatus for trapping of VOCs in murine breath. The detection of VOCs was performed by the SPME (solid phase microextraction)/ GC-MS (gas chromatography- mass spectrometry) method. Although we found several VOCs (2-ethyl-1-decanol、 2,3,6-trimethyl-octane、Isothiocyanatocyclohexane、1, 4-Methanoazulene (Longifolene)) in mouse chamber, there is no significant difference in TIC chromatograph between healthy and infected mouse. Thus we must reconsider the method for fungal infection., url

  対象リソースIRI
• Grant-in-Aid for Scientific Research (B), 2008 - 2010, 20300288, Analytical studies on microbial volatile organic compounds (MVOCs) and their physiological role in the microbial communities participating in the degradation of cultural properties., SUZUKI Takahito; TAKEUCHI Takae; IWAGUCHI Shinーichi, Japan Society for the Promotion of Science, Grants-in-Aid for Scientific Research Grant-in-Aid for Scientific Research (B), Nara Women's University, 19370000, 14900000, 4470000, (1) Some microorganisms and their communities play a role of deterioration on cultural heritage materials. Detection of their emergence at an earlier stage allows for reducing risks of the deterioration. Emission of MVOCs seems to be an effective candidate for an indicator of the occurrence of their growth. The MVOCs were collected by solid phase micro extraction (SPME) and analyzed by gas-chromatography/mass spectrometry (GC/MS) from the representative microorganisms relating the deterioration, including five fungal genera ( Aspergillus, Penicillium, Acremonium, Fusarium and Aureobasidium) and bacterial Streptomyces. We constructed the Finger Print Database (FPD) of MVOCs determined by GC/MS. We succeeded in developing an innovative apparatus detecting MVOCs based on the principle of ion mobility spectrometry. The FPD of MVOCs was also constructed by IMS.
  (2) A species of springtail (Collembola) was isolated from soil and its behavior on the flavor of several fungal species, described above, was analyzed using a video-camera. The springtail was found to feed on Penicillium. They did not show a significant feeding attraction or avoidance behavior against those fungi, either. However, obvious attractive one was found to the fungus Cladosporium.
  (3) The co-culturing system was applied to allow free gas or volatile exchange between the colonies of two fungal species. The effects of Fusarium solani volatiles were found on growth and development of the other fungi Aspergillus fumigatus and Penicillium paneum. Colonies of F. solani were incubated in the same growth chamber as colonies of either A. fumigatus or P. paneum. Analzing MVOCs by GC/MS, 2-pentadecanone, emitted from F. solani, was suggested as an accelerator of conidiophore development in both A. fumigatus and P. paneum. However, benzaldehyde from F. solani, may be an allelochemical inhibitor of growth against both A. fumigatus and P. paneum.
  (4) As a cooperative research with Prof. Nishiyama at Nara University, we obtained fungal isolates from the Takaida burial cave located at Kashiwara-shi, Osaka, Japan. Using these isolates, the occurrence of growth was investigated by simulation analysis on the process of their growth on the surface of tuff stones in the controlled environment. Fungal growth and the formation of conidia were found on these experiments., url

  対象リソースIRI
• Grant-in-Aid for Scientific Research (C), 2007 - 2009, 19570019, Factors affecting sex-ratio variation and sex-determining mechanisms in the apple snail Pomacea canaliculata, YUSA Yoichi; IWAGUCHI Shin-ichi, Japan Society for the Promotion of Science, Grants-in-Aid for Scientific Research Grant-in-Aid for Scientific Research (C), Nara Women's University, 3770000, 2900000, 870000, The freshwater snail Pomacea canaliculata shows a large variation in offspring sex ratio. We investigated whether this variation is caused by a small number of sex-determining genes, with two series of experiments. First, we reared F2 offspring from several crosses involving full-sib sisters with the same male, and determined sex ratios of F2. The sex ratios of F2 converged into a few values, indicating that the sex-determining genotypes of the F1 full-sib sisters were also a few. This suggests that the sex in this snail is determined by a few genes rather than polygenes. Secondly, we developed genetic markers to study linkage with the putative sex-determining genes. We found >40 microsatellite loci, and developed four useful markers. These markers were repeatable in PCR, polymorphic and followed Mendelian segregation. We conducted fragment analyses in two populations (Kumamoto and Nara) with these markers. One of them showed significantly different frequencies between males and females in both populations ; it may be in the same linkage group as a sex-determining locus., url

  対象リソースIRI
• 萌芽研究, 2005 - 2006, 17657023, 不完全酵母カンジダで発見された接合型遺伝子は形態形成にどう関わるか。, 鈴木 孝仁; 岩口 伸一, 日本学術振興会, 科学研究費助成事業 萌芽研究, 奈良女子大学, 3000000, 3000000, Candida albicansは有性生殖世代を欠いているため不完全菌酵母に分類されており、通常二倍体の核相を有している。しかし、臨床分離株NUM51では、核相が二倍体の細胞と四倍体の細胞が混在し、細胞周期のG2遅滞を経て二倍体から四倍体に核相が増加する。四倍体の細胞では、S.cerevisiaeの減数第二分裂に相当する多極性の核内分裂を経て、二倍体に核相を減少させていることが連続超薄切片を用いた電子顕微鏡による紡錘体極の観察から明らかになった。C.albicansに関するゲノムプロジェクトが進展した結果、接合型遺伝子MTLが5番染色体上に座乗していることが報告された。NUM51株でも、この接合型遺伝子は異型接合(MTLa/MTLalpha)の状態で存在していることが判明した。そして、MTLaが座乗する5番染色体を喪失させたMTLalphaのみのヘミ接合体株や、MTLalphaを遺伝子破壊した株では、核相の変換がほとんど起こらず、合成培地での菌糸形成率も低下していた。さらに、接合型遺伝子の異型接合株では、核相が増加した細胞の出現と相まって抗真菌剤フルコナゾールに対する耐性が上昇することが明らかとなった。ところが、接合型遺伝子やそれらが座乗する染色体の一方を失った株では、核相が一定であることと相まってフルコナゾール耐性度に変化はみられなかった。これらの事実から、核相の変換にはMTL遺伝子座が異型接合の状態で存在することが必要であること、MTL遺伝子座が座乗する5番染色体には菌糸形成や抗真菌剤耐性に関わる遺伝子群も座乗し、この染色体性の変化(モノソミーとなったままか、あるいはその後に複製が起きて二染色体性になるか)や接合性の変化を通じて当該遺伝子量の変化がもたらされ、それらの結果として、形態形成の反応や抗真菌剤への耐性応答の違いという表現型が現れることが判明した。, kaken

  対象リソースIRI
• Grant-in-Aid for Scientific Research (C), 2001 - 2002, 13640664, The role of vitamin B1 as a regulator of yeast dimorphism (yeast-hypha conversion), SUZUKI Takahito; IWAGUCHI Shin-ichi, Japan Society for the Promotion of Science, Grants-in-Aid for Scientific Research Grant-in-Aid for Scientific Research (C), Nara Women's University, 3600000, 3600000, The dimorphic transition from yeast to pseudohyphae in the petroleum-assimilating yeast Candida tropicalis occurs following the addition of ethanol to glucose semi-defined medium. Subtractive gene cloning was performed on the cDNA from the yeast-bowing control culture and on that from the ethanol-supplemented one (the ethanol culture). A homologue of Schizosaccharomyces pombe nmtl^+ or Saccharomyces cerevisiae THI5 was isolated from the cDNA fraction as a preferentially expressed gene for the ethanol culture. This homologue was tentatively called CtTHI5, since exogenous thiamine repressed its expression in C.tropicalis growth media. The ethanol culture showed biphasic pattern of growth phases and the expression of CtTHI5 occurred at the first growth phase. The supplementation of thiamine to the ethanol culture at the first phase was followed by repression of CtTHI5 expression and also delay of pseudohyphal growth and the occurrence of pseudomycelia in a stubby form with branched filaments. A CtTHI5 disruptant of this organism did not show thiamine auxotrophy and produced pseudohyphal filaments even in the control culture. The supplement of oxythiamine, an analog of thiamine, to the control culture was followed by the appearance of pseudohyphal filaments. Supplementation of valproic acid to the ethanol culture inhibited pseudohyphal growth, however, additional supplementation of thiamine partially cancelled the inhibition. These facts indicate that the participation of thiamine during the process of pseudohyphal growth in this organism., kaken

  対象リソースIRI
• Grant-in-Aid for Scientific Research (C), 1998 - 2000, 10640650, Characterization of genes regulating dimorphic transition between yeast and pseudohyphal form., SUZUKI Takahito; KAMIHARA Teijiro; IWAGUCHI Shin-ichi, Japan Society for the Promotion of Science, Grants-in-Aid for Scientific Research Grant-in-Aid for Scientific Research (C), NARA WOMEN'S UNIVERSITY, 3600000, 3600000, Ethanol caused mycelial growth in the imperfect yeast Candida tropicalis Pk233. Cultivation with ethanol gave biphasic growth curves. During the first growth phase in the ethanol culture, there was an accumulation of swollen spherical yeast cells and after entering the second phase, pseudohyphal cells appeared, projecting from the swollen yeast cells. Subtractive cloning of genes was performed on the cDNAs of the ethanol culture, from which the cDNAs of the control cultur had been subtracted by hybridization. Isolated included Ctnmt1^+ homologous to S.pombe nmt1^+ oncoding the enzyme for thiamine biosynthesis and CtPP1 homologous to C.albicans CPP1 encoding tyrosine phosphatase.. Correlation was suggested of the process of ethanol-induced pseudomycelial formation with thiamine metabolism, since the elongation of the pseudohyphal growth was suppressed when exogenous thiamine was supplemented at the late stage of the first growth phase. Disruption of CtPP1 brought about the phenotype defective in pseudomycelial formation, suggesting the participation of MAP kinase cascade in the formation., kaken

  対象リソースIRI
• 奨励研究(A), 1999 - 1999, 10770117, 病原性酵母カンジダの倍数性変換と染色体再配列を制御する遺伝子の解析, 岩口 伸一, 日本学術振興会, 科学研究費助成事業 奨励研究(A), 奈良女子大学, 1900000, 1900000, 日和見感染菌Candida albicansは通常二倍体だが、いくつか菌株では多倍数化した細胞の出現とそれに続く倍数性の減少すなわち倍数性変換が認められ、同時に染色体再編成が生じている。したがって倍数性変換がゲノムの変化を誘導する可能性を示唆している。倍数性変換を示す株の表現型はSps^-(suppresser of ploidy shift)で表される。それに対し、核相が一定である株の表現型はSps^+で表される。細胞融合実験からSps変異は劣性であることが示されているので、Sps^+株のゲノムDNAライブラリーをSps^-株に形質転換し、フロキシンB培地上で白い集落を形成すること、集落形態が滑面となること、増殖速度が速くなること、核の形態が一核となることという選択条件を設定しSPS遺伝子の単離を試みた。前年度に遺伝子破壊株であるSTN21U-1株に形質転換実験を行い、上記の選択方法に基づいてSps^+を示す13個の形質転換体を得た。本年度はその中からNo.115株をSPS遺伝子をもつ候補として選び出した。この株は細胞集団中にSTN21U-1株で観察される細胞よりも小さな細胞を生じる傾向があった。このNo.115株からプラスミドDNAを回収し、インサート部分をサブクローニングし、塩基配列を決定後に相同性検索を行ったところ、RAD5(REV2)遺伝子と高い相同性が見られた。Rad5タンパクはDNA修復タンパクのひとつであり、DNAへリカーゼであると考えられているが、詳しい機能については明らかにされていない。Rad5変異株では組み換えの頻度があがることが報告されているので、Sps^-株で染色体再編成が高頻度に起こるのは、RAD5遺伝子の変異に由来しているかもしれない。また、Sps^-株で高次倍数化するのは、RAD5の未知の機能が損なわれたためかもしれない。以上のことからRAD5がSPS遺伝子群の有力な候補の1つである可能性が考えられた。現在、RAD5遺伝子の機能解析を試みている。, kaken

  対象リソースIRI
• Grant-in-Aid for Scientific Research (C), 1996 - 1997, 08640846, Genetic regulation of dimorphic transition between yeast and hyphal form growth in the imperfect yeast, Candida tropicalis., SUZUKI Takahito; KAMIHARA Teijiro; IWAGUCHI Shin-ichi, Japan Society for the Promotion of Science, Grants-in-Aid for Scientific Research Grant-in-Aid for Scientific Research (C), NARA WOMEN'S UNIVERSITY, 2200000, 2200000, Ethanol caused mycelial growth in the imperfect yeast Candida tropicalis Pk233. Cultivation with ethanol gave biphasic growth curves. During the first growth phase in the ethanol culture, there was an accumulation of swollen spherical yeast cells, instead of the oblong ones observed in the control culture, followed by the appearance of spherical daughter cells in chains. During the second growth phase, hyphal cells appeared, projecting from the swollen yeast cells. Subtractive cloning of genes was performed on the cDNAs of the ethanol culture, from which the cDNAs of the control cultur had been subtracted by hybridization. Isolated genes specific for the process of hyphal formation included nmt1 homolog and URP2 homolog. Correlation was suggested of the process of ethanol-induced hyphal formation with thiamine metabolism, since the mycelial growth was abolished by addition of thiamine., kaken

  対象リソースIRI
• 奨励研究(A), 1996 - 1996, 08770191, 病原性酵母カンジダ・アルビカンスの染色体組換えを制御するタンパク質の検索, 岩口 伸一, 日本学術振興会, 科学研究費助成事業 奨励研究(A), 奈良女子大学, 900000, 900000, 日和見感染菌である酵母C.albicansのゲノム全体に存在している反復配列RPS1は、染色体再配列の生じている部位に局在している。このような反復配列がゲノム上に存在している場合、組換えを制御するタンパク質が細胞中に存在しなければ、ランダムな組換えが生じ、ゲノムの不安定性を引き起こすと考えられる。本研究ではC.albicansの反復配列RPS1に結合する調節タンパク質をゲル・シフト法により探索した。ゲル・シフト法では、ポリアクリルアミド電気泳動において、DNA断片とタンパク質の複合体の移動度は遊離のDNA断片に比べて遅くなるという原理を利用する。RPS1配列から30〜100bpのDNA断片を調整、ラベルし、C.albicansからのタンパク質抽出液と反応させた。その結果、30bpのCOM29に対して強く結合するタンパク質と、RNAとして発現している領域のDNA断片に結合するタンパク質を検出できた。COM29配列に結合するタンパク質は、大過剰の未標識のプローブを加えた競合実験においてもシグナルが観察される事から配列特異性が非常に高く、かつ結合力がかなり大きい事が分かった。さらにサウス・ウエスタン法によりCOM29配列に結合するタンパク質の大きさを調べたところ、49、38、26.8kdの3つのポリペプチドが検出された。COM29配列はRPS1ユニット中に5個存在しているため、このタンパク質がRPS1内部、さらにはRPSクラスター全体をパッケージングすることにより、組換えの鋳型として機能させなくしている可能性が考えられる。現在、COM29配列をCNBr-sepharose 4Bに結合させたDNAアフィニティーカラムを作製し、結合タンパク質の単離を試みている。, kaken

  対象リソースIRI
• 奨励研究(A), 1995 - 1995, 07857019, 病原性酵母カンジダの染色体再配列に関わるRPS1配列とその転写反応, 岩口 伸一, 日本学術振興会, 科学研究費助成事業 奨励研究(A), 奈良女子大学, 900000, 900000, 本研究ではRPS1がRNAを介した組み換えに関与しているかを調べるために、まずC.albicansにおいて発現型RPS1の単離を試みた。発現型のRPS1相同配列は、1006、WO-1株ともに同じ大きさで、約1.3kbであった。発現量はアクチン遺伝子のそれとの比較から、2株とも同程度発現していると考えられる。本研究では新たな試みとして、マグネット・ビーズ上にcDNAライブラリーを構築を行った。これにより安定にライブラリーが保存することができ、さらに、マグネット上のcDNAを鋳型にしてPCR、あるいは、サブトラクションやDNA結合タンパク質などの検索が可能になる。RPS1配列の大きさは、2.1kbなので配列の全てがmRNAとして転写されているわけではない。そこで、RPS1配列のどの部分が発現されているかをPCR法により検討した。マグネット上のcDNAを鋳型として、RPS1由来のプライマーと(dT)_<18>プライマーによりPCRを行った。その結果、プライマー24279、24280、22531、13267とXhoI-(dT)18との組み合わせでPCR産物が得られた。このことは、発現型RPS1は少なくともnon-REP配列とCOM29配列を含むことを示している。REP配列については、その存在はこの実験からは定かではないが、COM29配列がオリジナルのRPS1配列では常にREP配列に接しているのでおそらく発現型RPS1においても存在しているものと推定された。また、COM29由来のプライマーである13267によるPCR産物の大きさは、最も大きいもので約1.2kbのものが認められるので5´末端側にCOM29由来の配列が存在すると予想される。それぞれのPCR産物のクローニングが現在行われている。, kaken

  対象リソースIRI
• Grant-in-Aid for General Scientific Research (C), 1994 - 1995, 06640865, MOLECULAR MECHANISM OF PLOIDY-SHIFT AND VARIATION OF COLONY MORPHOLOGY IN THE IMPERFECT YEASTS, SUZUKI Takahito; IWAGUCHI Shinichi, Japan Society for the Promotion of Science, Grants-in-Aid for Scientific Research Grant-in-Aid for General Scientific Research (C), NARA WOMEN'S UNIVERSITY, 2100000, 2100000, Some strains of Candida albicans poroduce a proportion of the cell population with a higher ploidy than the rest in normal diploid state. This phenotype was named as Pld^-by the author. Ploidy-shift was observed between two different ploidy states. Pld-STN21 (arg4, Pld^-) was isolated from a weakly growing sector of a half-sectored colony, after UV-irradiation of STN14, an arg4 derivative of C.albicans CBS5736 and from the other normally growing sector, STN22 (arg4, Pld^+) was isolated. STN22, as well as its progenitor, CBS5736, showed a homogeneus colony size when plated out, but STN21 showed a characteristic heterogeneity of colony size on YPD plates at an early stage of incubation : two types of colonies, small ones and large ones, were observed.
  The chromosomes of these strains were separated out by PFGE.Some culture of STN21 showed the single band of chromosome 7, suggesting due to the occurrence of chromosome non-disjunction. The other culture of STN21 gave an extrachromosomal band, the size of which was larger than that of chromosome 7, and the loss of one of the two bands of chromosome 7. Southern blot hybridization experiments showed the existence of a SfiI fragment 7F,a central portion of chromosome 7, on the extrachromosomal band but the absence of a terminal SfiI fragment 7C.Protoplast fusion experiments showed, at least, two complementation groups of PLD,the function of which may stabilize both ploidy state and chromosome fidelity., kaken

  対象リソースIRI
• 微生物が放出する揮発性有機化合物（MVOCs）を利用した微生物汚染、微生物感染の早期発見方法の開発, 0, 0, 0, 「揮発性有機化合物を指標とした真菌感染症の早期診断法の開発」日本学術振興会科学研究費補助金基盤研究（挑戦的萌芽研究）を実施中, Competitive research funding
• 微生物を利用した発酵食品開発, 0, 0, 0, 酵母、カビを利用した発酵食品の研究開発、商品化協力, Competitive research funding
• 野生酵母を利用した発酵食品開発, 0, 0, 0, Competitive research funding
• 不完全真菌の二形性変換, 0, 0, 0, Competitive research funding
• 不完全真菌の倍数性変換機構, 0, 0, 0, Competitive research funding
• 不完全真菌における染色体再配列のメカニズム, 0, 0, 0, Competitive research funding
• Dimorphism in fungi, 0, 0, 0, Competitive research funding
• Ploidy shift in Fungi, 0, 0, 0, Competitive research funding
• Chromosome rearrangement in Fungi, 0, 0, 0, Competitive research funding
• 科学研究費助成事業 基盤研究(C), 2022 - 2026, 22K05532, Coinvestigator, 酵母による赤ワインの色調増強に関わる技術基盤の確立