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1 COMPLEX ADAPTIVE TRAITS Newsletter Vol. 2 No

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8 Ishida JK 1,2, Yoshida S 1, Wafula E 3, depamphilis CW 3, Namba S 2, Shirasu K 1. ( 1 Plant Science Center. RIKEN, Japan 2 Graduate School of Agricultural and Life Science - The University of Tokyo, Japan 3 Department of Biology, The Pennsylvania State University, USA) Parasitic plant Phtheirospermum japonicum invades host to take water and nutrients via haustorium. To find the genes essential for haustorium development the transcriptome of P. japonicum were sequenced. The assembly resulted in contigs. The 60 bp probes from assembled sequences were immobilized in microarray slides and hybridized with labeled crna extracted at different time points during the treatment with DMBQ (2, 6-dimethoxy-pbenzoquinone), a chemical able to induce the haustorium in vitro. Differential gene expression was confirmed using qrt-pcr. The expression levels of transcription factors WRKY33 and the nuclear protein Pirin were increased during the first three hours of DMBQ treatment, suggesting those proteins may play important roles during the early haustoria development. In addition DMBQinduced differential regulation of genes encoding oxygen processing enzymes such as peroxidases, glutathione transferases and quinone redutases, allow us to infer the pivotal role of reaction oxygen species during the haustoria development. In future, reverse genetic approaches will be taken to further analyze the function of these genes. Sucrases are digestive enzymes that hydrolyze α-glucosyl (α-glucosidase, EC ) or β- fructosyl residue (β-fructofuranosidase, β-ffase, EC ) of the substrate. α-glucosidase but not β-ffase activity is inhibited by the sugar-mimic alkaloids. We identified the β-ffase cdnas not only from Bombyx mori, but also from Rondotia menciana, Glyphodes pyloalis, Antheraea pernyi, Samia cynthia ricini, and Trilocha varians. qrt-pcr indicated that β-ffase genes (Suc1) were much more highly expressed in the midgut of the mulberry-feeders (Bm, Rm, Gp) than in that of the non-mulberry feeders (Ap, Scr, Tv), indicating that Suc1 potentially contributes to the adaptation of the insects to mulberry. To investigate the transcriptional regulation of the β-ffase genes, we determined the sequences of the 5' upstream regions of GpSuc1, TvSuc1 and BmSuc1 and found that putative silk gland factor 1 (SGF1) binding sites in the promoter regions were conserved among three species, suggesting some roles of SGF1. One the other hand, we found that the inhibitory effect of DNJ and D-AB1 on soluble midgut α- glucosidase activities of non-mulberry-feeders was stronger than that in the mulberry-feeders. It suggests that not only Suc1 but also the α-glucosidases genes have evolved to adapt to the mulberry alkaloids. 6

japonicum invades host to take water and nutrients via haustorium. To find the genes essential for haustorium development the transcriptome of P. japonicum were sequenced.

9 Oleg Gusev, Kyosuke Mukae, Richard Cornette, Takashi Okuda and Takahiro Kikawada (Anhydrobiosis Research Group, National Institute of Agrobiological Sciences, Ohwashi 1-2, Tsukuba, , JAPAN) Larvae of the sleeping chironomid Polypedilim vanderplanki are able to survive decades of complete dehydration. In addition, in the dry form the larvae show high cross-resistance to various abiotic stresses, including vacuum and ionizing radiation. We have recently found that the anhydrobiotic larvae experience massive DNA damage and excessive level of oxidative stress upon every cycle of dehydration-rehydration. The sleeping chironomid is characterized by significant expansion in homologous and paralogous copies of defensive genes, such as major antioxidants of thioredoxin-based cycle, heat shock proteins and protein-reparation methyltransferases. We have observed clear differences in expression patterns of the paralogs in relation to anhydrobiosis. In addition, we hypothesize that ability to stand prolonged period of severe nuclear DNA fragmentation is linked to adaptive changes in apoptosis control network, as many of typical apoptosis initiators are missing in the sleeping chironomid genome

In addition, in the dry form the larvae show high cross-resistance to various abiotic stresses, including vacuum and ionizing radiation.

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18 COMPLEX ADAPTIVE TRAITS Newsletter Vol. 2 No. 4 COMPLEX ADAPTIVE TRAITS Newsletter URL

日本消化器外科学会雑誌第29巻第9号

日本消化器外科学会雑誌第29巻第9号 Table 1 Oligonucleotide primers used for RT-PCR and internal probes used for Southern blot hybridization Cytokine Primer Sequence (5'-3') 5' 3' Internal probe s' 3' Internal probe 5' 3' Internal probe