Osamu Numata Ph.D. Doctoral Program in Structural Biosciences,
Graduate School of Life and Environmental Sciences,
University of Tsukuba
Office: F502 Agricultural and Biosciences Building
Lab： D408 Agricultural and Biosciences Building
Mailing address： 1-1-1 Tennohdai, Tsukuba, Ibaraki 305-8572, Japan
TEL 81(29)-853-6648/4530
FAX 029-853-6648
numata.osamu.gb@u.tsukuba.ac.jp

Research

To answer numbers of question, we have choosen Tetrahymena for our biologic tool. The Ciliated protozoan Tetrahymena is an unicellular organism that lives in fresh water such as ponds, rivers, and lakes. Tetrahyemna cell is surrounded by hundreds of cilia, and the beating of cilia produces the force to swim in water (the name Ciliates comes from the cilia). Tetrahymena have two different nucleus in single cell, the macronucleus and the micronucleus (This is the important character of Ciliates).

(1) Cytokinesis (Actin associated proteins)

Cytokinesis is the final event of the cell division cycle, and results in partition of a mother cell into two daughter cells. Tetrahymena actin forms so called, contractile ring or actin ring across the presumptive division plane to contract a mother cell. Before the formation of contractile ring, p85 and calmodulin localize to the basal body at presumptive division plane. Moreover, once the contractile ring is formed, profilin, fimbrin, and elongation factor-1 alpha become localized to the contractile ring. We hypothesize that Calmodulin/p85 play role in the initiation of contractile ring formation and profilin, fimbrin, EF-1 alpha, and myosin plays role in maintenance and contraction of the actin ring.

(2) Actin and Myosin

Actin and myosin play crutial role in cell motility , such as contractile ring contraction, organell transport, cytoplasmic flow, and etc. Since Tetrahymena cell is unicellular, we expect that many myosins play roles to maintain cells to be functional. To investigate this possibility, we have so far newly isolated 11 candidates of Tetrahymena myosins. On the other hand, we have also isolated several actin related genes including novel actin related genes.

(3) Cilia

The avoiding reaction of Tetrahyemna cell is incoroporated with the change of forward swimming to backward swimming. This process is strictly regulated by a Calcium flux. How this calcium signal is passed to swimming change is not knowm. We have biochemichally investigated Ca/Calmodulin associated protein from the ciliary fraction and found EF-1alpha as a Ca/CaM binding proetin. Electron micrograph showed both EF-1alpha and CaM localized in cilia.

(4) Oolong Tea

Oolong tea is considered to be good for health and effects on human body have been examined recently. After drinking oolong tea an increase has been observed in the amount of CO2 exhaled. This indicates that some substance in oolong tea may have the potential to activate oxygen respiration. To test this hypothesis we treated Tetrahymena and mouse sperm with oolong tea and investigated the effect of this on the mitochondrial membrane potential. The mitochondrial membrane potential was detected with rhodamine 123 and the intensity of fluorescence was measured by fluorophotometer. We found that the oolong tea highly activated the mitochondrial membrane potential in Tetrahymena and mouse sperm. Hydrophobic fraction of oolong tea contained high molecule polyphenol and had high activation. The hydrophobic fraction of oolong tea also elevated swimming velocity by up to 25~33% and 100%~ in Tetrahymena and mouse sperm, respectively, when compared with a control. The results suggest that the hydrophobic fraction of oolong tea increases ATP synthesis through activation of the cellular metabolism, such as TCA cycle or oxidative phosphorylation and then elevates swimming velocity.

(5) Tetrahymena calcium binding proetin (TCBP)

The Tetrahymena Ca2+-binding protein of 25 kDa (TCBP-25) and 23kDa (TCBP-23) are members of the calmodulin family containing four EF-hand Ca(2+)-binding loops. TCBPs localize to the whole cell cortex except for the oral apparatus and around the basal bodies. TCBP-25 localizes around both the migratory and the stationary gametic pronuclei at the pronuclear exchange stage during conjugation. Anti-sense gene suppression of TCBP-25, failed to exchange pronucleus, thus TCBP-25 plays imporant role in the nuclear exchange.

(6) Macronuclear division

There are two functionally and structually different nuclei in single Tetrahymena cell. They are the micronucleus (germinal) and the macronucleus (vegitative). Our focus is on the dividing mechanism of macronucelus. Macronuclear chromosome lack centrosome and during the macronuclear division, spindle MTs is absent, and chromosome condensation does not take place. Eventhough, MTs are assembled in macronucleus and disruption of macronucleus inhibits its division. Hence the macronuclear division is said to be amitotic, though it depends on MTs. Then how? We have so far clarified the localization of MTs which suggested that the macronuclear division may be a premitive mitosis rather than amitosis. To learn the molecular mechanism of the macronuclear division, we have searched for MTs associated protein form the macronucelus, and found, termed p138. p138 interact with MTs and most likely to interact with chromosome. p138 encoded a protein homologus to atranscription elongation factor FACT (facilitates chromatin transcription) 140 kDa subunit. We are now examining the in vivo function of p138

Recent publications

* : Corresponding author

  1. Yuki Aoki, Tetsuo Ozawa, Tohru Takemasa and Osamu Numata* (2017) Black Tea High-Molecular-Weight Polyphenol-Rich Fraction Promotes Hypertrophy during Functional Overload in Mice. Molecules., Vol. 22, 548
  2. Rikuri Morita, Masak Takaine, Osamu Numata and Kentaro Nakano*. (2017) Molecular dissection of the actin-binding ability of the fission yeast α-actinin, Ain1, in vitro and in vivo. J Biochem.
  3. Yasuharu Kushida, Masak Takaine, Kentaro Nakano, Toshiro Sugai, Krishna Kumar Vasudevan, Mayukh Guha, Yu-yang Jiang, Jacek Gaertig and Osamu Numata* (2016) Kinesin-14 Is Important for Chromosome Segregation during Mitosis and Meiosis in the Ciliate Tetrahymena thermophila. J Eukaryot Microbiol., Vol. 64, 293-307
  4. Tsuyoshi Yasuda, Masak Takaine, Osamu Numata and Kentaro Nakano* (2016) Anillin-related protein Mid1 regulates timely formation of the contractile ring in the fission yeast Schizosaccharomyces japonicus. Genes Cells, vol. 21, 594-607.
  5. Yasuharu Kushida, Masak Takaine, Kentaro Nakano, Toshiro Sugai and Osamu Numata*. (2015) Analysis of localization of γ-tubulin during conjugation of ciliate Tetrahymena thermophila using confocal lazar-scanning microscope. Zoological Sci., vol. 32, 25-32.
  6. Masak Takaine, Kazuki Imada, Osamu Numata, Taro Nakamura and Kentaro Nakano. (2014) The meiosis-specific nuclear passenger protein is required for proper assembly of forespore membrane in fission yeast. J. Cell Sci., vol. 127, 4429-4442.
  7. Masak Takaine*, Osamu Numata and Kentaro Nakano* (2013) Fission yeast IQGAP maintains F-actin-independent localization of myosin-II in the contractile ring. Genes Cells, vol. 19, 161-176.
  8. Tomonori Eguchi, Chisato Kumagai, Takashi Fujihara, Tohru Takemasa, Tetsuo Ozawa and Osamu Numata* (2013) Black Tea High-Molecular-Weight Polyphenol Stimulates Exercise Training-Induced Improvement of Endurance Capacity in Mouse via the Link between AMPK and GLUT4.PLoS ONE 8(7): e69480　doi:10.1371/journal.pone.0069480
  9. Yuhta Shimizu, Yasuharu Kushida, Shuhei Kiriyama, Kentaro Nakano* and Osamu Numata* (2013) Formation of division furrow and its ingression can progress under the inhibitory condition of actin polymerization in ciliate Tetrahymena pyriformis. Zoological Sci., vol. 30, 1044-1049.
  10. Nanami Shiozaki, Kentaro Nakano*, Yasuharu Kushida, Taro QP Noguchi, Taro QP Uyeda, Dorota Wloga, Drashti Dave, Krishna K. Vasudevan, Jacek Gaertig and Osamu Numata (2013) ADF/cofilin is not essential but critically important for actin activities during phagocytosis in Tetrahymena thermophila. Eukaryot. Cell, vol. 12, 1080-1086.
  11. Ayako Kikuchi, Kogiku Shiba, Tetsuo Ozawa Kentaro Nakano, Kazuo Inaba and Osamu Numata* (2012) Black tea high-molecular-weight polyphenol increases motility of sea urchin sperm by activating mitochondrial respiration. Biosci. Biotech. Biochem. vol. 76, 2321-2324.
  12. Sugita, M., Iwataki, Y., Nakano, K. and Numata, O. (2011) Unique sequences and predicted functions of myosins in Tetrahymena thermophila. Gene, 480, 10-20.
  13. Kushida, Y., Nakano, K. and Numata, O. (2011) Amitosis requires γ-tubulin-mediated microtubule assembly in Tetrahymena thermophila. Cytoskeleton, 68, 89-96.
  14. Saito-Nakano, Y., Nakahara T., Nakano K., Nozaki T. and Numata, O. (2010) Marked amplification and diversification of products of ras genes from rat brain, Rab GTPases, in the ciliates Tetrahymena thermophila and Paramecium tetraurelia. J Eukaryot Microbiol, 57, 389-399.
  15. Nakano, K., Kuwayama, H., Kawasaki, M., Numata, O. and Takaine M. (2010) GMF is an evolutionarily developed Adf/cofilin-super family protein involved in the Arp2/3 complex-mediated organization of the actin cytoskeleton. Cytoskeleton, 67, 373-382.
  16. Shiozaki, N., Nakano, K., Takaine, M., Abe, H. and Numata, O. (2009) Usual and unusual biochemical properties of ADF/cofilin-like protein Adf73p in ciliate Tetrahymena thermophila. Biochem Biophys Res Commun, 390, 54-9.
  17. Takaine, M., Numata, O. and Nakano, K. (2009) Fission yeast IQGAP arranges actin filaments into the cytokinetic contractile ring. EMBO J, 28, 3117-3131.
  18. Li, W., Zhang, S., Numata, O., Nozawa, Y. and Wang, S. (2009) TpMRK regulates cell division of Tetrahymena in response to oxidative stress. Cell Biochem Funct, 27, 364-369.
  19. Sugita, M., Nakano, K., Sato, M., Toyooka, K. and Numata, O. (2009) The roles of actin cytoskeleton and microtubules for membrane recycling of a food vacuole in Tetrahymena thermophila. Cell Motil Cytoskeleton, 66, 371-377.
  20. Nakagawa, T., Fujiu, K., Cole, E.S. and Numata, O. (2008) Involvement of a 25 kDa Tetrahymena Ca2+-binding Protein in Pronuclear Exchange. Cell Structure and Function, 33, 151-162.
  21. Morita, K., Bunai, F. and Numata, O. (2008) Roles of three domains of Tetrahymena eEF1A in bundling F-actin. Zoological Science, 25, 22-29.
  22. Fujihara, T., Nakagawa-Izumi, A., Ozawa, T. and Numata, O. (2007) High-Molecular-Weight Polyphenols from Oolong Tea and Black Tea: Purification, Some Properties, and Role in Increasing Mitochondrial Membrane Potential. Bioscience, Biotechnology, and Biochemistry, 71, 711-719.
  23. Ueno, H., Iwataki, Y. and Numata, O. (2006) Homologues of radial spoke head proteins interact with Ca2+/calmodulin in Tetrahymena cells. J. Biochem., 140, 525-533.
  24. Bunai, F., Ando, K., Ueno, H. and Numata, O. (2006) Tetrahymena eukaryotic translation elongation factor 1A (eEF1A) bundled filamentous actin through dimer formation. J. Biochem., 140, 393-399.
  25. Sakiyama, T., Ueno, H., Homma, H., Numata, O. and Kuwabara, T. (2006) Purification and characterization of a hemolysin-like protein Sll1951, a non-toxic member of the RTX protein family from cyanobacterium Synechocystis sp. PCC 6803. J. Bacteriology, 188, 3535-3542.
  26. Kuribara, S., Kato, M., Kato-Minoura, T. and Numata, O. (2006) Identification of a novel actin-related protein in Tetrahymena cilia. Cell Motility and the Cytoskeleton, 63, 437-446.
  27. Bunai, F., Ando, K., Ueno, H. and Numata, O. (2006) Tetrahymena eukaryotic translation elongation factor 1A (eEF1A) bundled filamentous actin through dimer formation. J. Biochem., 140, 393 - 399.
  28. Ueno, H., Iwataki, Y. and Numata, O. (2006) Homologues of radial spoke head proteins interact with Ca2+/calmodulin in Tetrahymena cilia. J. Biochem., 140, 525 - 533.
  29. Nakano, K., Bunai, F. and Numata, O. (2005) Stg1 is a novel SM22/transgelin-like actin-modulating protein in fission yeast. FEBS Letter, 579, 6311-6316.
  30. Fujiu, K. and Numata, O. (2004) Identification and molecular cloning of Tetrahymena 136kDa protein, a transcription elongation factor homologue that interacts with microtubules in vitro. Biochem. Biophys. Res. Commun. 315, 196-203.
  31. Ueno, H., Gonda, K., Takeda, T. and Numata, O. (2003) Identification of elongation factor-1α as a Ca2+/calmodulin-binding protein in Tetrahymena cilia. Cell Motility and the Cytoskeleton, 55, 51-60.
  32. Shirayama, S. and Numata, O. (2003) Tetrahymena fimbrin localized in the division furrow bundles actin filaments in a calcium-independent manner. J. Biochem. 134, 591-598.
  33. Ishida, H., Suzaki, T., Kuribayashi, C., Mauyama, E. and Numata, O. (2003) Distribution of actin-like proteins in the ciliate Spirostomum ambiguum. Jpn. J. Protozool. 36, 141-146.
  34. Kojima, H. and Numata, O. (2002) Enzymatic form and cytoskeletal form of bifunctional Tetrahymena 49kDa protein is regulated by phosphorylation. Zool. Sci., 19, 37-42.
  35. Chang, J-S., Seok, H., Kwon, T-K., Min, D. S., Ahn, B-H., Lee, Y. H., Suh, J-W., Kim, J-W., Iwashita, S., Omori, A., Ichinose, S., Numata, O., Seo, J-K., Oh, Y-S. and Suh, P-G. (2002) Interaction of elongation factor-1α and pleckstrin homology domain of phospholipase C-γ1 with activating its activity. J. Biol. Chem., 277, 19697-19702.
  36. Gonda, K. and Numata, O. (2002) p85 binds to G-actin in Ca2+/calmodulin-dependent manner, thus regulating the initiation of cytokinesis in Tetrahymena. Biochem. and Biophys. Res. Commun., 292, 1098- 1103.
  37. Takeda, T., Yoshihama, I. and Numata, O. (2001) Identification of Tetrahymena hsp60 as a 14-nm filament protein/citrate synthase-binding protein and its possible involvement in the oral apparatus formation. Genes to Cells, 6, 139-149.
  38. Numata, O., Kurasawa, Y., Gonda, K. and Watanabe. Y. (2000) Tetrahymena elongation factor-1α is localized with calmodulin in the division furrow. Journal of Biochemistry, 127, 51-56.
  39. Watanabe, A., Yonemura, I., Gonda, K. and Numata, O. (2000) Cloning and sequencing of the gene for a Tetrahymena fimbrin-like protein. Journal of Biochemistry, 127, 85-94.
  40. Fujiu, K. and Numata, O. (2000) Reorganization of microtubules in the amitotically dividing Macronucleus of Tetrahymena. Cell Motility and the Cytoskeleton, 46, 17-27.
  41. Gonda, K., Komatsu, M. and Numata, O. (2000) Calmodulin and Ca2+/calmodulin-binding proteins are involved in Tetrahymena thermophila phagocytosis. Cell Structure and Function, 25, 243-251.