Gan P.,RIKEN |
Ikeda K.,Kyoto University |
Irieda H.,Kyoto University |
Narusaka M.,Research Institute for Biological science |
And 5 more authors.
New Phytologist | Year: 2013
Hemibiotrophic fungal plant pathogens represent a group of agronomically significant disease-causing agents that grow first on living tissue and then cause host death in later, necrotrophic growth. Among these, Colletotrichum spp. are devastating pathogens of many crops. Identifying expanded classes of genes in the genomes of phytopathogenic Colletotrichum, especially those associated with specific stages of hemibiotrophy, can provide insights on how these pathogens infect a large number of hosts. The genomes of Colletotrichum orbiculare, which infects cucurbits and Nicotiana benthamiana, and C. gloeosporioides, which infects a wide range of crops, were sequenced and analyzed, focusing on features with potential roles in pathogenicity. Regulation of C. orbiculare gene expression was investigated during infection of N. benthamiana using a custom microarray. Genes expanded in both genomes compared to other fungi included sequences encoding small, secreted proteins (SSPs), secondary metabolite synthesis genes, proteases and carbohydrate-degrading enzymes. Many SSP and secondary metabolite synthesis genes were upregulated during initial stages of host colonization, whereas the necrotrophic stage of growth is characterized by upregulation of sequences encoding degradative enzymes. Hemibiotrophy in C. orbiculare is characterized by distinct stage-specific gene expression profiles of expanded classes of potential pathogenicity genes. © 2012 The Authors. New Phytologist © 2012 New Phytologist Trust.
Abe H.,RIKEN |
Narusaka Y.,Research Institute for Biological science |
Sasaki I.,RIKEN |
Hatakeyama K.,National Institute of Vegetable and Tea Science |
And 5 more authors.
DNA Research | Year: 2011
Arabidopsis belongs to the Brassicaceae family and plays an important role as a model plant for which researchers have developed fine-tuned genome resources. Genome sequencing projects have been initiated for other members of the Brassicaceae family. Among these projects, research on Chinese cabbage (Brassica rapa subsp. pekinensis) started early because of strong interest in this species. Here, we report the development of a library of Chinese cabbage full-length cDNA clones, the RIKEN BRC B. rapa full-length cDNA (BBRAF) resource, to accelerate research on Brassica species. We sequenced 10 000 BBRAF clones and confirmed 5476 independent clones. Most of these cDNAs showed high homology to Arabidopsis genes, but we also obtained more than 200 cDNA clones that lacked any sequence homology to Arabidopsis genes. We also successfully identified several possible candidate marker genes for plant defence responses from our analysis of the expression of the Brassica counterparts of Arabidopsis marker genes in response to salicylic acid and jasmonic acid. We compared gene expression of these markers in several Chinese cabbage cultivars. Our BBRAF cDNA resource will be publicly available from the RIKEN Bioresource Center and will help researchers to transfer Arabidopsis-related knowledge to Brassica crops. © 2011 The Author.
Kawai F.,Kyoto Institute of Technology |
Kitajima S.,Kyoto Institute of Technology |
Oda K.,Research Institute for Biological science |
Higasa T.,Kyoto Institute of Technology |
And 3 more authors.
Archives of Microbiology | Year: 2013
Scanning electron microscopy (SEM) shows remarkable morphological surface changes in Sphingopyxis sp. 113P3 cells grown in polyvinyl alcohol (PVA) but not in Luria-Bertani medium (LB) (Hu et al. in Arch Microbiol 188: 235-241, 2007). However, transmission electron microscopy showed no surface changes in PVA-grown cells and revealed the presence of polymer bodies in the periplasm of PVA-grown cells, which were not observed in LB-grown cells. The presence of polymer bodies was supported by low-vacuum SEM observation of PVA- and LB-grown cells of strain 113P3, and the presence of similar polymer bodies was also found when Sphingopyxis macrogoltabida 103 and S. terrae were grown in polyethylene glycol (PEG). The extraction of PVA and PEG from the periplasmic fraction of cells using a modified Anraku and Heppel method and their analysis by MALDI-TOF mass spectrometry strongly suggested that the polymer bodies are composed of PVA and PEG, respectively, in Sphingopyxis sp. 113P3 (PVA degrader) and Sphingopyxis macrogoltabida 103 or S. terrae (PEG degraders). PEG-grown S. macrogoltabida 103 and S. terrae showed higher transport of 14C-PEG 4000 than LB-grown cells. Recombinant PegB (TonB-dependent receptor-like protein consisting of a barrel structure) interacted with PEG 200, 4000 and 20000, suggesting that the barrel protein in the outer membrane contributes to the transport of PEG into the periplasm. © 2012 Springer-Verlag Berlin Heidelberg.
Nahar K.,Okayama University |
Matsumoto I.,Okayama University |
Taguchi F.,Okayama University |
Inagaki Y.,Okayama University |
And 6 more authors.
Molecular Plant Pathology | Year: 2014
Summary: Ralstonia solanacearum is a Gram-negative soil-borne bacterium that causes bacterial wilt disease in more than 200 plant species, including economically important Solanaceae species. In R.solanacearum, the hypersensitive response and pathogenicity (Hrp) type III secretion system is required for both the ability to induce the hypersensitive response (HR) in nonhost plants and pathogenicity in host plants. Recently, 72 effector genes, called rip (Ralstonia protein injected into plant cells), have been identified in R.solanacearumRS1000. RS1002, a spontaneous nalixidic acid-resistant derivative of RS1000, induced strong HR in the nonhost wild eggplant Solanum torvum in an Hrp-dependent manner. An Agrobacterium-mediated transient expression system revealed that Rip36, a putative Zn-dependent protease effector of R.solanacearum, induced HR in S.torvum. A mutation in the putative Zn-binding motif (E149A) completely abolished the ability to induce HR. In agreement with this result, the RS1002-derived Δrip36 and rip36E149A mutants lost the ability to induce HR in S.torvum. An E149A mutation had no effect on the translocation of Rip36 into plant cells. These results indicate that Rip36 is an avirulent factor that induces HR in S.torvum and that a putative Zn-dependent protease motif is essential for this activity. © 2013 BSPP AND JOHN WILEY & SONS LTD.
Hanano S.,Research Institute for Biological science |
Goto K.,Research Institute for Biological science
Plant Cell | Year: 2011
TERMINAL FLOWER1 (TFL1) is a key regulator of flowering time and the development of the inflorescence meristem in Arabidopsis thaliana. TFL1 and FLOWERING LOCUS T (FT) have highly conserved amino acid sequences but opposite functions. For example, FT promotes flowering and TFL1 represses it; FT-overexpressing plants and TFL1 loss-of-function mutants have a similar phenotype production of terminal flowers in the shoot apex. FT is believed to function in a transcriptional activator complex by interacting with FD. Here, we demonstrate that TFL1 is involved in the transcriptional repression of genes that are activated by FT. We analyzed transgenic plants overexpressing TFL1 fused to a transcriptional repressor domain (TFL1-SRDX) or an activator domain (TFL1-VP16). Plants carrying 35S:TFL1-SRDX showed delayed flowering similar to 35S:TFL1 plants, and plants carrying 35S:TFL1-VP16 showed an early flowering phenotype and produced terminal flowers. Furthermore, the tfl1 and 35S:TFL1-VP16 plant phenotypes were strongly suppressed by the fd mutation, and TFL1 interacted with FD in the cell nucleus, as shown by bimolecular fluorescence complementation experiments. We conclude that TFL1 negatively modulates the FD-dependent transcription of target genes to fine-tune flowering time and the development of the inflorescence meristem. © 2011 American Society of Plant Biologists. All rights reserved.
Shiota H.,Okayama University |
Kanzaki H.,Okayama University |
Hatanaka T.,Research Institute for Biological science |
Nitoda T.,Okayama University
Carbohydrate Research | Year: 2013
TMG-chitotriomycin (1) produced by the actinomycete Streptomyces annulatus NBRC13369 was examined as a probe for the prediction of substrate specificity of β-N-acetylhexosaminidases (HexNAcases). According to the results of inhibition assays, 14 GH20 HexNAcases from various organisms were divided into 1-sensitive and 1-insensitive enzymes. Three representatives of each group were investigated for their substrate specificity. The 1-sensitive HexNAcases hydrolyzed N-acetylchitooligosaccharides but not N-glycan-type oligosaccharides, whereas the 1-insensitive enzymes hydrolyzed N-glycan-type oligosaccharides but not N-acetylchitooligosaccharides, indicating that TMG-chitotriomycin can be used as a molecular probe to distinguish between chitin-degrading HexNAcases and glycoconjugate-processing HexNAcases. © 2013 Elsevier Ltd. All rights reserved.
Mukaihara T.,Research Institute for Biological science |
Tamura N.,Research Institute for Biological science |
Tamura N.,Okayama Prefectural General Agriculture Center |
Iwabuchi M.,Research Institute for Biological science
Molecular Plant-Microbe Interactions | Year: 2010
The gram-negative plant-pathogenic bacterium Ralstonia solanacearum utilizes the hypersensitive response and pathogenicity (Hrp) type III secretion system (T3SS) to cause disease in plants. To determine the entire repertoire of effector proteins possessed by R. solanacearum RS1000, we constructed a transposon carrying a calmodulin-dependent adenylate cyclase reporter that can be used to specifically detect rip (Ralstonia protein injected into plant cells) genes by monitoring the cAMP level in plant leaves inoculated with insertion mutants. From the new functional screen using this transposon, we identified 38 new Rip proteins translocated into plant cells via the Hrp T3SS. In addition, most of the 34 known effectors of RS1000 could be detected by the screen, except for three effectors that appear to be small in size or only weakly expressed. Finally, we identified 72 Rips in RS1000, which include 68 effector proteins classified into over 50 families and four extracellular components of the Hrp T3SS. Interestingly, one-third of the effectors are specific to R. solanacearum. Many effector proteins contain various repeated amino acid sequences or known enzyme motifs. We also show that most of the R. solanacearum effector proteins, but not Hrp extracellular components, require an Hrp-associated protein, HpaB, for their effective translocation into plant cells. © 2010 The American Phytopathological Society.
A qualitative analysis of the regulation of cyclic electron flow around photosystem I from the post-illumination chlorophyll fluorescence transient in Arabidopsis: A new platform for the in vivo investigation of the chloroplast redox state
Gotoh E.,Kyushu University |
Matsumoto M.,Research Institute for Biological science |
Ogawa K.,Research Institute for Biological science |
Ogawa K.,Japan Science and Technology Agency |
And 2 more authors.
Photosynthesis Research | Year: 2010
A transient in chlorophyll fluorescence after cessation of actinic light illumination, which has been ascribed to electron donation from stromal reductants to plastoquinone (PQ) by the NAD(P)H-dehydrogenase (NDH) complex, was investigated in Arabidopsis thaliana. The transient was absent in air in a mutant lacking the NDH complex (ndhM). However, in ndhM, the transient was detected in CO2-free air containing 2% O2. To investigate the reason, ndhM was crossed with a pgr5 mutant impaired in ferredoxin (Fd)-dependent electron donation from NADPH to PQ, which is known to be redundant for NDH-dependent PQ reduction in the cyclic electron flow around photosystem I (PSI). In ndhM pgr5, the transient was absent even in CO2-free air with 2% O2, demonstrating that the post-illumination transient can also be induced by the Fd- (or PGR5)-dependent PQ reduction. On the other hand, the transient increase in chlorophyll fluorescence was found to be enhanced in normal air in a mutant impaired in plastid fructose-1,6-bisphosphate aldolase (FBA) activity. The mutant, termed fba3-1, offers unique opportunities to examine the relative contribution of the two paths, i.e., the NDH- and Fd- (or PGR5)-dependent paths, on the PSI cyclic electron flow. Crossing fba3-1 with either ndhM or pgr5 and assessing the transient suggested that the main route for the PSI cyclic electron flow shifts from the NDH-dependent path to the Fd-dependent path in response to sink limitation of linear electron flow. © Springer Science+Business Media B.V. 2010.
Mworia E.G.,Okayama University |
Yoshikawa T.,Okayama University |
Yokotani N.,Research Institute for Biological science |
Fukuda T.,Kagawa Agricultural Experiment Stn. 6117 1 Fuchu Cho |
And 4 more authors.
Postharvest Biology and Technology | Year: 2010
Ethylene biosynthesis in kiwifruit, Actinidia chinensis 'Sanuki Gold' was characterized using propylene, an ethylene analog, and 1-methylcyclopropene (1-MCP), an inhibitor of ethylene perception. In fruit harvested between a young stage (66 days after pollination) (DAP) and an early commercial harvesting stage (143 DAP), 2 days of exposure to propylene were sufficient to initiate ethylene biosynthesis while in fruit harvested at commercial harvesting stage (154 DAP), 4 days of propylene treatment were required. This observation suggests that response of ethylene biosynthesis to propylene treatment in kiwifruit declined with fruit maturity. Propylene treatment resulted in up-regulated expression of AC-ACO1, AC-ACO2, AC-SAM1 and AC-SAM2, prior to the induction of AC-ACS1 and ethylene production, confirming that AC-ACS1 is the rate limiting step in ethylene biosynthesis in kiwifruit. Treatment of fruit with more than 5 μL L-1 of 1-MCP after the induction of ethylene production subsequently suppressed ethylene production and expression of ethylene biosynthesis genes. Treatment of fruit with 1-MCP at harvest followed with propylene treatment delayed the induction of ethylene production and AC-ACS1 expression for 5 days. These observations suggest that in ripening kiwifruit, ethylene biosynthesis is regulated by positive feedback mechanism and that 1-MCP treatment at harvest effectively delays ethylene production by 5 days. © 2009 Elsevier B.V. All rights reserved.
PubMed | Research Institute for Biological science
Type: Journal Article | Journal: mBio | Year: 2016
The plant pathogen Ralstonia solanacearum uses a large repertoire of type III effector proteins to succeed in infection. To clarify the function of effector proteins in host eukaryote cells, we expressed effectors in yeast cells and identified seven effector proteins that interfere with yeast growth. One of the effector proteins, RipAY, was found to share homology with the ChaC family proteins that function as -glutamyl cyclotransferases, which degrade glutathione (GSH), a tripeptide that plays important roles in the plant immune system. RipAY significantly inhibited yeast growth and simultaneously induced rapid GSH depletion when expressed in yeast cells. The in vitro GSH degradation activity of RipAY is specifically activated by eukaryotic factors in the yeast and plant extracts. Biochemical purification of the yeast protein identified that RipAY is activated by thioredoxin TRX2. On the other hand, RipAY was not activated by bacterial thioredoxins. Interestingly, RipAY was activated by plant h-type thioredoxins that exist in large amounts in the plant cytosol, but not by chloroplastic m-, f-, x-, y- and z-type thioredoxins, in a thiol-independent manner. The transient expression of RipAY decreased the GSH level in plant cells and affected the flg22-triggered production of reactive oxygen species (ROS) and expression of pathogen-associated molecular pattern (PAMP)-triggered immunity (PTI) marker genes in Nicotiana benthamiana leaves. These results indicate that RipAY is activated by host cytosolic thioredoxins and degrades GSH specifically in plant cells to suppress plant immunity.Ralstonia solanacearum is the causal agent of bacterial wilt disease of plants. This pathogen injects virulence effector proteins into host cells to suppress disease resistance responses of plants. In this article, we report a biochemical activity of R. solanacearum effector protein RipAY. RipAY can degrade GSH, a tripeptide that plays important roles in the plant immune system, with its -glutamyl cyclotransferase activity. The high GSH degradation activity of RipAY is considered to be a good weapon for this bacterium to suppress plant immunity. However, GSH also plays important roles in bacterial tolerance to various stresses and growth. Interestingly, RipAY has an excellent safety mechanism to prevent unwanted firing of its enzyme activity in bacterial cells because RipAY is specifically activated by host eukaryotic thioredoxins. This study also reveals a novel host plant protein acting as a molecular switch for effector activation.