著者

Shimizu Tetsu Takaya Naoki Nakamura Akira

出版者

American Society for Biochemistry and Molecular Biology

雑誌

The journal of biological chemistry (ISSN:00219258)

巻号頁・発行日

vol.287, no.48, pp.40448-40456, 2012-11

被引用文献数

36 6

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An l-glucose-utilizing bacterium, Paracoccus sp. 43P, was isolated from soil by enrichment cultivation in a minimal medium containing l-glucose as the sole carbon source. In cell-free extracts from this bacterium, NAD+-dependent l-glucose dehydrogenase was detected as having sole activity toward l-glucose. This enzyme, LgdA, was purified, and the lgdA gene was found to be located in a cluster of putative inositol catabolic genes. LgdA showed similar dehydrogenase activity toward scyllo- and myo-inositols. l-Gluconate dehydrogenase activity was also detected in cell-free extracts, which represents the reaction product of LgdA activity toward l-glucose. Enzyme purification and gene cloning revealed that the corresponding gene resides in a nine-gene cluster, the lgn cluster, which may participate in aldonate incorporation and assimilation. Kinetic and reaction product analysis of each gene product in the cluster indicated that they sequentially metabolize l-gluconate to glycolytic intermediates, d-glyceraldehyde-3-phosphate, and pyruvate through reactions of C-5 epimerization by dehydrogenase/reductase, dehydration, phosphorylation, and aldolase reaction, using a pathway similar to l-galactonate catabolism in Escherichia coli. Gene disruption studies indicated that the identified genes are responsible for l-glucose catabolism.

著者

Kamimura Manabu Saito Hitoshi Niwa Ryusuke Niimi Teruyuki Toyoda Kinuko Ueno Chihiro Kanamori Yasushi Shimura Sachiko Kiuchi Makoto

出版者

American Society for Biochemistry and Molecular Biology

雑誌

The journal of biological chemistry (ISSN:00219258)

巻号頁・発行日

vol.287, no.20, pp.16488-16498, 2012-05

被引用文献数

22

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Steroid hormones ecdysteroids regulate varieties of developmental processes in insects. Although the ecdysteroid titer can be increased experimentally with ease, its artificial reduction, although desirable, is very difficult to achieve. Here we characterized the ecdysteroid-inactivating enzyme ecdysteroid-22-oxidase (E22O) from the entomopathogenic fungus Nomuraea rileyi and used it to develop methods for reducing ecdysteroid titer and thereby controlling insect development. Km and Kcat values of the purified E22O for oxidizing ecdysone were 4.4 μm and 8.4/s, respectively, indicating that E22O can inactivate ecdysone more efficiently than other ecdysteroid inactivating enzymes characterized so far. The cloned E22O cDNA encoded a FAD-dependent oxidoreductase. Injection of recombinant E22O into the silkworm Bombyx mori interfered with larval molting and metamorphosis. In the hemolymph of E22O-injected pupae, the titer of hormonally active 20-hydroxyecdysone decreased and concomitantly large amounts of inactive 22-dehydroecdysteroids accumulated. E22O injection also prevented molting of various other insects. In the larvae of the crambid moth Haritalodes basipunctalis, E22O injection induced a diapause-like developmental arrest, which, as in normal diapause, was broken by chilling. Transient expression of the E22O gene by in vivo lipofection effectively decreased the 20-hydroxyecdysone titer and blocked molting in B. mori. Transgenic expression of E22O in Drosophila melanogaster caused embryonic morphological defects, phenotypes of which were very similar to those of the ecdysteroid synthesis deficient mutants. Thus, as the first available simple but versatile tool for reducing the internal ecdysteroid titer, E22O could find use in controlling a broad range of ecdysteroid-associated developmental and physiological phenomena.

著者

Nagamine Satoshi Tamba Michiko Ishimine Hisako Araki Kota Shiomi Kensuke Okada Takuya Ohto Tatsuyuki Kunita Satoshi Takahashi Satoru Wismans Ronnie G. P. Kuppevelt Toin H. van Masu Masayuki Keino-Masu Kazuko

出版者

American Society for Biochemistry and Molecular Biology

雑誌

The journal of biological chemistry (ISSN:00219258)

巻号頁・発行日

vol.287, no.12, pp.9579-9590, 2012-03

被引用文献数

70

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Heparan sulfate endosulfatases Sulf1 and Sulf2 hydrolyze 6-O-sulfate in heparan sulfate, thereby regulating cellular signaling. Previous studies have revealed that Sulfs act predominantly on UA2S-GlcNS6S disaccharides and weakly on UA-GlcNS6S disaccharides. However, the specificity of Sulfs and their role in sulfation patterning of heparan sulfate in vivo remained unknown. Here, we performed disaccharide analysis of heparan sulfate in Sulf1 and Sulf2 knock-out mice. Significant increases in ΔUA2S-GlcNS6S were observed in the brain, small intestine, lung, spleen, testis, and skeletal muscle of adult Sulf1−/− mice and in the brain, liver, kidney, spleen, and testis of adult Sulf2−/− mice. In addition, increases in ΔUA-GlcNS6S were seen in the Sulf1−/− lung and small intestine. In contrast, the disaccharide compositions of chondroitin sulfate were not primarily altered, indicating specificity of Sulfs for heparan sulfate. For Sulf1, but not for Sulf2, mRNA expression levels in eight organs of wild-type mice were highly correlated with increases in ΔUA2S-GlcNS6S in the corresponding organs of knock-out mice. Moreover, overall changes in heparan sulfate compositions were greater in Sulf1−/− mice than in Sulf2−/− mice despite lower levels of Sulf1 mRNA expression, suggesting predominant roles of Sulf1 in heparan sulfate desulfation and distinct regulation of Sulf activities in vivo. Sulf1 and Sulf2 mRNAs were differentially expressed in restricted types of cells in organs, and consequently, the sulfation patterns of heparan sulfate were locally and distinctly altered in Sulf1 and Sulf2 knock-out mice. These findings indicate that Sulf1 and Sulf2 differentially contribute to the generation of organ-specific sulfation patterns of heparan sulfate.

著者

Nakano Naoko Itoh Susumu Watanabe Yukihide Maeyama Kota Itoh Fumiko Kato Mitsuyasu

出版者

American Society for Biochemistry and Molecular Biology

雑誌

The journal of biological chemistry (ISSN:00219258)

巻号頁・発行日

vol.285, no.49, pp.38023-38033, 2010-12

被引用文献数

43

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The TGF-β and Wnt pathways are involved in cell fate and tumorigenicity. A recent report indicated that a TGF-β target gene, TMEPAI (transmembrane prostate androgen-induced RNA), is possibly also a downstream target of Wnt signaling. Although TMEPAI was believed to be involved in tumorigenicity because of its blockage of TGF-β signaling, how TGF-β and Wnt signals affect the activation of the TMEPAI gene is not well understood. Herein, we show that the TMEPAI promoter is regulated synergistically by TGF-β/Smad and Wnt/β-catenin/T cell factor (TCF) 7L2. The critical cis-element for dual signals, termed TGF-β-responsive TCF7L2-binding element (TTE), is located in intron 1 of the TMEPAI gene. TCF7L2, but not Smad proteins, bound to TTE, whereas the disruption of TTE by mutagenesis remarkably counteracted both TGF-β and TCF7L2 responses. The introduction of mutations in critical Smad-binding elements blocked the activation of the TMEPAI promoter by TCF7L2. Furthermore, our DNA-protein interaction experiments revealed the indirect binding of TCF7L2 to Smad-binding elements via Smad3 upon TGF-β stimulation as well as its TGF-β-dependent association with TTE. We demonstrate that the Wnt/β-catenin/TCF7L2 pathway is preferentially able to alter the transcriptional regulation of the TGF-β-target gene, TMEPAI.