Annual Report 2006

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P lant Physiology Department The research activities of our department are mainly focused on the elucidation of molecular mechanisms of important physiological processes in plants. The major topics in fiscal 2005 are as follows. Photosynthesis and carbon metabolism Identification of critical residues for substrate recognition of CtpA, an integral protease for assembly of oxygen-evolving complex of photosystem II. Carboxyl-terminal processing protease A (CtpA) is an indispensable protease for assembling of the photosynthetic oxygen evolving machinery, which performs proteolytic processing of the precursor form of the D1 protein, a pivotal subunit of photosystem II. The CtpA is well known to have very narrow substrate specificity, which is one of the common properties of proteases involved in regulation of cellular processes. In order to approach molecular basis of the high specificity, we examined each residue of the spinach CtpA by several concepts that tendency of conservation in the CtpA family, distance from the catalytic center and accessibility of substrates. I400, V416 and Y419 were extracted as candidate residues involved in substrate recognition of the protease (Fig. 1A). We introduced a series of site-directed mutations about these residues of the spinach gene and transformed the mutagenized genes into deleted sp. PCC 6803 cells, which helps rapid evaluation of effects of each mutation onto the CtpA activity. These analyses revealed that V416 and Y419 are not so important for the CtpA action, because these positions could be accepted various substitutions without deactivation of the CtpA. In contrast, I400 is likely to be critical for CtpA activity. It is noteworthy that I400T, a relatively mild substitution, significantly abolishes the activity and then cells with the mutant gene could not grow on Fig. 1 (A) Three dimensional structure of the CtpA and positions of our target residues Catalytic center residues, S372 and K397 were colored with red and blue, respectively. CtpA has two grooves close to the catalytic center. I400, V416 and Y419 (colored with orange) emerged on surface of the narrow groove. Fig. 1 (B) Photoautotrophic growth test of the several mutants cells carrying the I400T mutant gene could not grow on photoautotrophic medium (- glucose). In contrast, V416T cells did not abolish the CtpA activity and then the mutant as well as WT cells grew well on the same medium.
photoautotrophic medium (Fig. 1B). It clearly suggested that hydrophobic environment supplied from this aliphatic residue is required for maintenance of the CtpA activity. It is most probable that the I400 is a key residue for substrate recognition of the CtpA and contributes the high specificity of the protease. Genetic analysis of C3-C4 intermediate photosynthesis C3-C4 intermediate plants have lower activities of photorespiration than C3 plants. We investigated structural and photosynthetic characteristics in leaves of reciprocal hybrids differing in genome constitution produced between (C3-C4; MaMa) and (C3; CC). We found that the hybrids had intermediate features between the parents and expressed more strongly the C3-C4 intermediate characteristics with an increase in the constitution ratio of the Ma:C genome. The same pattern of expression of the C3-C4 intermediacy was found between the hybrids that the parents were reciprocally exchanged. We conclude that the C3-C4 intermediate characteristics are inherited in the hybrids depending on the constitution ratio of the parent genomes and there is no evidence of maternal inheritance in these characteristics. Isolation of genes that are differentially expressed during C3 and C4 leaf development C4 leaves have a specialized structural feature, Kranz anatomy, that differs from C3 leaves. is a unique grass including both Kranz (C4) and non-Kranz (C3) forms within this one species. We used subtractive hybridization to isolate genes that are differentially expressed during leaf development of the two forms. Northern blot analyses of the subtracted clones revealed that six and 13 genes were C3- and C4-abundant, respectively, most of which showed high similarities to genes involved in mechanisms of regulation, signaling and transport, carbon metabolism, and protection from abiotic or biotic stress. We are now attempting to characterize these genes more in detail to elucidate possible involvements in C4 leaf development. Identification and physiological analyses of a locus for rice yield potential We analyzed quantitative trait loci (QTLs) for the ratio of filled grains, a yield component, in backcrossed inbred lines of Nipponbare and Kasalath. Only one QTL (), with a positive Kasalath allele, was detected across environments. In a near-isogenic line (NILrg5) with a Kasalath chromosome segment containing carbohydrate storage capacity before heading or sink activity was significantly higher than in Nipponbare (control). The ratio of filled grains and yield per plant were significantly higher in NILrg5 than in Nipponbare, by 5% ( < 0.01) and 15% ( < 0.05), respectively. These results suggest that improves carbohydrate storage capacityandkeepssinkactivityhigherinthe reproductive stage, and consequently increases yield potential. Light response Distinct functions of different phytochrome species in rice We have isolated phytochrome B () and mutants from rice ( ) and have produced all combinations of double mutants. Seedlings of and mutants exhibited a partial loss of sensitivity to continuous red light (Rc) but still showed significant deetiolation responses. The responses to Rc were completely canceled in double mutants. These results indicate that phyA and phyB act in a highly redundant manner to control deetiolation under Rc. Under continuous far-red light (FRc), mutants showed partially impaired deetiolation, and double mutants showed no significant residual phytochrome responses, indicating that not only phyA but also phyC is involved in the photoperception of FRc in rice. Interestingly, the double mutant displayed clear R/FR reversibility in the pulse irradiation experiments, indicating that both
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2006
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Teruo Ishige, President The Nationa
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List of Publication Original paper
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Topics of Research in This Year Pos
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studies. To construct a fully satur
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Fig. 2 Geographical distribution of
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(ABR0015, ABR0257, ABR0417, ABR0495
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Mulberry latex defense mulberry tre
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Fig. 2 Growth inhibitory effects of
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Fig. 3 Change of the levels of luci
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Next, we examined the effect of tra
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G enome Research Department Introdu
Page 25 and 26: The Rice Annotation Project Databas
Page 27 and 28: industrial and agricultural concern
Page 29 and 30: One of the main problems involving
Page 31 and 32: Fig. 3 Mapping of QTL for rachis in
Page 33 and 34: two genes, XOO2263 and XOO4134 cont
Page 35 and 36: G enebank Genebank Project The NIAS
Page 37 and 38: using dormant winter buds. Also, we
Page 39 and 40: Insect and Animal Sciences Division
Page 41 and 42: Expression patterns of two novel ge
Page 43 and 44: Fig. 5 PGC and EBC fused cells PGCs
Page 45 and 46: cells. The epidermal cells maintain
Page 47 and 48: Animal and cell culture models for
Page 49 and 50: terminus, respectively. A signal se
Page 51 and 52: Fig. 2 Trehalose content of body pa
Page 53 and 54: Fig. 6 Birth weight of 11 adult SNT
Page 55 and 56: The role of glucose as a metabolic
Page 57 and 58: analysis using microsatellite marke
Page 59 and 60: Fig. 2 Recorded Electromyograms (le
Page 61 and 62: Table 1 Menu of Commercial silkworm
Page 63 and 64: Fig. 3 Expression of GFP gene in th
Page 65 and 66: also surveyed in the genetic stocks
Page 67 and 68: esearch were located in or near gen
Page 69 and 70: elatives results in severe reductio
Page 71 and 72: and circadian clocks may control th
Page 73 and 74: B iochemistry Department Structural
Page 75: Proteomic analysis Based on the pro
Page 79 and 80: Fig. 4 Blast resistance of OsWRKY45
Page 81 and 82: phosphatase activity by SIPK depend
Page 83 and 84: the endosperm in co-operation with
Page 85 and 86: isolated genes in transgenic Anal
Page 87 and 88: Fig. 1 Gene structures and mutation
Page 89 and 90: cytochimera among the mulberries (
Page 91 and 92: 15 G. Hariraj, Kinoshita Haruo, T.
Page 93 and 94: 49 Kameda Tsunenori, Teramoto Hidet
Page 95 and 96: 85 Matsuyama Shuichi, Ohkura Satosh
Page 97 and 98: 120 Ozawa Kenjiro, Kawahigashi Hiro
Page 99 and 100: 155 Taguchi-Shiobara Fumio, Yamamot
Page 101 and 102: 192 Ueno Osamu, Agarie Sakae, (2005
Page 103 and 104: 8 Murakami Ritsuko, Koyama Akio, Sh
Page 105 and 106: 20 Todoroki Junichi, Kaneko Hiroyuk
Page 107 and 108: Executive Members and Research Staf
Page 109 and 110: Developmental Biology Department Di
Page 111 and 112: Research Leader Hayasaka Shoji Rese
Page 113 and 114: Mutation Genetics Laboratory Head N
Page 115 and 116: thousands of yen TOTAL BUDGET OPERA
Page 117: Annual Report 2006 (Apr. 2005Mar. 2
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Annual Report 2006

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