研究者紹介システム

李 智博
リ トモヒロ
大学院農学研究科 資源生命科学専攻
准教授
農学関係
Last Updated :2023/12/12

研究者情報

所属

  • 【主配置】

    大学院農学研究科 資源生命科学専攻
  • 【配置】

    農学部 資源生命科学科

学位

  • 博士(農学), 神戸大学

授業科目

ジャンル

  • 科学・技術 / 生命科学

コメントテーマ

  • 発生生物学
  • 減数分裂
  • 卵母細胞
  • 精母細胞
  • マウス

研究活動

研究キーワード

  • meiosis
  • developmental biology
  • 減数分裂
  • 発生生物学

研究分野

  • ライフサイエンス / 動物生命科学

受賞

  • 2013年 日本繁殖生物学会, 第106回日本繁殖生物学会大会優秀発表賞(ポスター部門), マウス精母細胞における減数分裂型コヒーシンの詳細な局在位置

    美 栄, 松田 厚志, 平岡 泰, 李 智博

    国際学会・会議・シンポジウム等の賞

  • 2012年 日本繁殖生物学会, 日本繁殖生物学会賞奨励賞, 哺乳類減数分裂における染色体動態の分子機構

    李 智博

    国内学会・会議・シンポジウム等の賞

論文

  • Yuto Taniuchi, Kazutaka Hiraide, Rilige Su, Kazune Ijuin, XingQiang Wei, Takuro Horii, Izuho Hatada, Jibak Lee

    RAD2lL and REC8, meiosis-specific paralogs of the canonical cohesin subunit RAD21, are essential for proper formation of axial/lateral elements of the synaptonemal complex, synapsis of homologous chromosomes, and crossover recombination in mammalian meiosis. However, how many meiotic cohesins are present in germ cells has not been investigated because of the lack of an appropriate method of analysis. In the present study, to examine the intracellular amount of meiotic cohesins, we generated two strains of knock-in (KI) mice that expressed a 3×FLAG-tag at the C-terminus of RAD21L or REC8 protein using the CRISPR/Cas9 genome editing system. Both KI mice were fertile. Western blot analyses and immunocytochemical studies revealed that expression levels and localization patterns of both RAD21L-3×FLAG and REC8-3×FLAG in KI mice were similar to those in wild-type mice. After confirming that tagging of endogenous RAD21L and REC8 with 3×FLAG did not affect their expression profiles, we evaluated the levels of RAD21L-3×FLAG and REC8-3×FLAG in the testes of 2-week-old mice in which only RAD21L and REC8 but little RAD21 are expressed in the meiocytes. By comparing the band intensities of testicular RAD21L-3×FLAG and REC8-3×FLAG with 3×FLAG-tagged recombinant proteins of known concentrations in western blot analysis, we found that there were approximately 413,000 RAD21L and 453,000 REC8 molecules per spermatocyte in the early stages of prophase I. These findings provide new insights into the role played by cohesins in the process of meiotic chromosome organization in mammalian germ cells.

    2023年04月03日, The Journal of reproduction and development, 69 (2), 78 - 86, 英語, 国内誌

    [査読有り]

    研究論文(学術雑誌)

  • Riho Morikawa, Hirohisa Kyogoku, Jibak Lee, Takashi Miyano

    Oocytes communicate with the surrounding somatic cells during follicular development. We examined the effects of two oocyte-derived growth factors, growth differentiation factor 9 (GDF9) and bone morphogenetic protein 15 (BMP15), on the development of porcine oocyte-cumulus cell complexes (OCCs) in vitro. We collected OCCs from early antral follicles (1.2-1.5 mm) and prepared oocytectomized cumulus cell complexes (OXCs), which were then cultured in a growth medium supplemented with 0-100 ng/ml GDF9 and/or BMP15 for 7 days. In the medium without GDF9 or BMP15, OCCs developed during culture, and approximately 30% of them formed antrum-like structures. GDF9 promoted OCC development and structure formation in a dose-dependent manner. However, OXCs did not form antrum-like structures without growth factors. GDF9 promoted the development of OXCs, and 50 and 100 ng/ml GDF9 promoted the formation of the structures by 8% and 26%, respectively; however, BMP15 did not promote the formation of these structures. OXCs were then cultured with 100 ng/ml GDF9 and various concentrations of BMP15 to investigate their cooperative effects on the formation of antrum-like structures. BMP15 promoted the formation of antrum-like structures in a dose-dependent manner. In conclusion, GDF9 derived from oocytes is probably important for the formation of antrum-like structures in porcine OXCs, and BMP15 cooperates with GDF9 to form these structures.

    2022年05月02日, The Journal of reproduction and development, 68, 238 - 245, 英語, 国内誌

    [査読有り]

    研究論文(学術雑誌)

  • Riho Morikawa, Jibak Lee, Takashi Miyano

    During oocyte growth and follicle development, oocytes closely communicate with cumulus cells. We examined the effects of oocyte-derived growth factors, growth differentiation factor 9 (GDF9) and bone morphogenetic protein 15 (BMP15), on the growth and acquisition of meiotic competence of porcine oocytes collected from early antral follicles (1.2-1.5 mm). First, we confirmed that GDF9 and BMP15 mRNAs were expressed almost exclusively in the oocytes. Oocyte-cumulus cell complexes (OCCs) collected from early antral follicles were cultured in growth medium supplemented with 0-100 ng/ml of GDF9 or BMP15 for 5 days. GDF9 dose-dependently increased the OCC diameter, while BMP15 did not. GDF9 and BMP15 had no significant effects on oocyte growth (P > 0.05). When OCCs that had been cultured with 50 and 100 ng/ml BMP15 were subjected to a subsequent maturation culture, they expanded fully by gonadotropic stimulation and 49% and 61% of oocytes matured to metaphase II (MII), respectively. In contrast, GDF9 did not promote cumulus expansion, and < 10% of oocytes matured to MII. Based on the difference in cumulus expansion, we compared the expression of luteinizing hormone/choriogonadotropin receptor (LHCGR) and follicle stimulating hormone receptor (FSHR) mRNAs in cumulus cells. The level of LHCGR mRNA was increased in cumulus cells of the BMP15 group, although there were no significant differences in FSHR mRNA levels among the groups. These results suggest that GDF9 promotes the growth of OCCs and that BMP15 promotes LHCGR mRNA expression in cumulus cells during oocyte growth culture, which may contribute to cumulus expansion and oocyte maturation.

    2021年08月27日, The Journal of reproduction and development, 67 (4), 273 - 281, 英語, 国内誌

    [査読有り]

    研究論文(学術雑誌)

  • Mihoko Fushii, Rie Yamada, Jibak Lee, Takashi Miyano

    Transzonal projections (TZPs) that maintain bidirectional communication between oocytes and granulosa cells or cumulus cells are important structures for oocyte growth. However, whether TZPs develop between TZP-free oocytes and granulosa cells, and whether reestablished TZPs support oocyte growth, is unknown. We first examined changes in TZPs after denudation of bovine oocytes collected from early antral follicles (0.5-0.7 mm). Twenty-four hours after denudation, almost all the TZPs disappeared. We also examined the reestablishment of TZPs by coculturing TZP-free denuded oocytes (DOs) with mural granulosa cells (MGCs) collected from early antral follicles. In addition, to confirm if the reestablished TZPs were functional, the reconstructed complexes (DO+MGCs) were subjected to in vitro growth culture and found that the MGCs adhered to TZP-free DOs and TZPs were reestablished. During in vitro growth culture, DO+MGCs developed and formed antrum-like structures. After culture, the number of TZPs in DO+MGCs increased, and the oocytes grew fully and acquired meiotic competence. These results suggest that reestablished TZPs are able to support oocyte growth.

    2021年08月20日, The Journal of reproduction and development, 67, 300 - 306, 英語, 国内誌

    [査読有り]

    研究論文(学術雑誌)

  • Is age‐related increase of chromosome segregation errors in mammalian oocytes caused by cohesin deterioration?

    Lee J

    2020年01月, Reprod Med Biol, 19, 32 - 41

    [査読有り][招待有り]

    研究論文(学術雑誌)

  • Md Hasanur Alam, Jibak Lee, Takashi Miyano

    Bovine growing oocytes with a diameter of 105–115 μm from early antral follicles (1.2–1.8 mm) are able to resume meiosis, but lack the competence to mature to metaphase II. To confer full maturation competence onto the oocytes, culture systems which can support their growth and prevent their meiotic resumption during culture are needed. In this study, we cultured growing oocytes for 5 days to examine the effects of different phosphodiesterase (PDE) inhibitors on meiotic arrest and acquisition of full maturation competence of growing oocytes, and their gap junctional communication with cumulus cells. Growing oocyte-cumulus complexes (OCCs) were cultured with 3-isobutyl-1-methylxanthine (IBMX broad-spectrum PDE inhibitor), rolipram (PDE4D inhibitor), cilostamide and milrinone (PDE3A inhibitors). The mean diameters of oocytes increased similarly in all groups. IBMX, cilostamide and milrinone induced antrum formation by OCCs and maintained meiotic arrest of oocytes during culture, whereas rolipram neither promoted antrum formation nor maintained oocyte meiotic arrest. Gap junctional communication between oocytes and cumulus cells was maintained by IBMX and cilostamide, but not by rolipram as judged by the transfer of injected lucifer yellow dye from oocytes to cumulus cells. In subsequent in vitro maturation, oocytes grown with IBMX, cilostamide and milrinone showed full maturation competence. These results suggest that PDE3A inhibition maintains the meiotic arrest of bovine growing oocytes and sustains their gap junctional communication with cumulus cells for 5 days, thereby contributing to their acquisition of full maturation competence.

    Elsevier Inc., 2018年09月15日, Theriogenology, 118, 110 - 118, 英語

    [査読有り]

    研究論文(学術雑誌)

  • Alam MH, Lee J, Miyano T

    2018年, J Reprod Dev, 64 (5), 423 - 431, 英語

    [査読有り]

    研究論文(学術雑誌)

  • Mei Rong, Sachi Miyauchi, Jibak Lee

    Pairing, synapsis, and crossover recombination of homologous chromosomes (homologs) are prerequisite for the proper segregation of homologs during meiosis I. The meiosis-specific cohesin subunit, RAD21L, is essential for such meiotic chromosomal events, but it is unknown to what extent RAD21L by itself contributes to the process since various meiotic genes are also involved. To reveal the exclusive contribution of RAD21L to the specific regulation of homologs, we examined the effects of ectopic RAD21L expression on chromosome dynamics in somatic cells. We found that expression of GFP-fused RAD21L by plasmid transfection significantly shortened the distance between two FISH signals representing a pair of homologs for chromosome X or chromosome 11 in the nuclei compared to that in control (non-transfected) cells whereas expression of GFP-fused RAD21, a mitotic counterpart of RAD21L, showed no detectable effects. This indicates that RAD21L, when ectopically expressed in somatic cells, can promote homolog adjacency, which resembles the homolog pairing normally seen during meiosis. Furthermore, deletion of the N-terminal winged helix domain from RAD21L, prevented its association with another cohesin subunit, SMC3, and abolished the phenomenon of homolog adjacency upon ectopic expression. Our findings suggest that RAD21L-containing cohesin can promote homolog adjacency independently of other meiotic gene products, which might be central to the process of meiotic homolog paring.

    SOCIETY REPRODUCTION & DEVELOPMENT-SRD, 2017年06月, JOURNAL OF REPRODUCTION AND DEVELOPMENT, 63 (3), 227 - 234, 英語

    [査読有り]

    研究論文(学術雑誌)

  • Mei Rong, Atsushi Matsuda, Yasushi Hiraoka, Jibak Lee

    Cohesins containing a meiosis-specific alpha-kleisin subunit, RAD21L or REC8, play roles in diverse aspects of meiotic chromosome dynamics including formation of axial elements (AEs), assembly of the synaptonemal complex (SC), recombination of homologous chromosomes (homologs), and cohesion of sister chromatids. However, the exact functions of individual alpha-kleisins remain to be elucidated. Here, we examined the localization of RAD21L and REC8 within the SC by super-resolution microscopy, 3D-SIM. We found that both RAD21L and REC8 were localized at the connection sites between lateral elements (LEs) and transverse filaments (TFs) of pachynema with RAD21L locating interior to REC8 sites. RAD21L and REC8 were not symmetrical in terms of synaptic homologs, suggesting that the arrangement of different cohesins is not strictly fixed along all chromosome axes. Intriguingly, some RAD21L signals, but not REC8 signals, were observed between unsynapsed regions of AEs of zygonema as if they formed a bridge between homologs. Furthermore, the signals of recombination intermediates overlapped with those of RAD21L to a greater degree than with those of REC8. These results highlight the different properties of two meiotic alpha-kleisins, and strongly support the previous proposition that RAD21L is an atypical cohesin that establishes the association between homologs rather than sister chromatids.

    SOCIETY REPRODUCTION & DEVELOPMENT-SRD, 2016年12月, JOURNAL OF REPRODUCTION AND DEVELOPMENT, 62 (6), 623 - 630, 英語

    [査読有り]

    研究論文(学術雑誌)

  • Martin Houlard, Jonathan Godwin, Jean Metson, Jibak Lee, Tatsuya Hirano, Kim Nasmyth

    In addition to inter-chromatid cohesion, mitotic and meiotic chromatids must have three physical properties: compaction into 'threads' roughly co-linear with their DNA sequence, intra-chromatid cohesion determining their rigidity, and a mechanism to promote sister chromatid disentanglement. A fundamental issue in chromosome biology is whether a single molecular process accounts for all three features. There is universal agreement that a pair of Smc-kleisin complexes called condensin I and II facilitate sister chromatid disentanglement, but whether they also confer thread formation or longitudinal rigidity is either controversial or has never been directly addressed respectively. We show here that condensin II (beta-kleisin) has an essential role in all three processes during meiosis I in mouse oocytes and that its function overlaps with that of condensin I (gamma-kleisin), which is otherwise redundant. Pre-assembled meiotic bivalents unravel when condensin is inactivated by TEV cleavage, proving that it actually holds chromatin fibres together.

    NATURE PUBLISHING GROUP, 2015年06月, NATURE CELL BIOLOGY, 17 (6), 771 - +, 英語

    [査読有り]

    研究論文(学術雑誌)

  • Mihiro Shibutani, Jibak Lee, Takashi Miyano, Masashi Miyake

    The embryo culture technique has been improving, but the detailed demands for energy substrates such as glucose, fructose, pyruvate and lactate of preimplantation embryos are still unclear. In the present study, the demands of pig preimplantation embryos at each different developmental stage were investigated by use of parthenogenetic diploids as a model of pig preimplantation embryos. Pig parthenogenetic diploids showed different use of glucose and fructose before and after the 4-cell stage. Although glucose supported the development of pig embryos throughout the preimplantation stages and even maintained the expansion and hatching of blastocysts, it suppressed development to the blastocyst stage when glucose coexisted with pyruvate and lactate from 4 h after activation, but not after 48 h (early 4-cell stage). Since ketohexolcinase that metabolizes fructose was not expressed in 2-cell and 4-cell diploids, a medium that included only fructose as a major energy substrate did not support early cleavage of pig diploids beyond the 4-cell stage, and almost no diploids developed to the morula stage just as in a medium without carbohydrates. These results may explain the different suppressive effects on pig preimplantation development between glucose and fructose when pyruvate and lactate were present in a medium. In addition, 4-cell diploids that had been cultured in a medium with pyruvate and lactate developed to the expanded blastocyst stage without any carbohydrates as a major energy substrate. These results show that the demands for carbohydrates are different depending on the developmental stage in pig preimplantation embryos.

    SOCIETY REPRODUCTION & DEVELOPMENT-SRD, 2015年04月, JOURNAL OF REPRODUCTION AND DEVELOPMENT, 61 (2), 106 - 115, 英語

    [査読有り]

    研究論文(学術雑誌)

  • Jibak Lee

    Meiosis is a key step for sexual reproduction in which chromosome number is halved by two successive meiotic divisions after a single round of DNA replication. In the first meiotic division (meiosis I), homologous chromosomes pair, synapse, and recombine with their partners in prophase I. As a result, homologous chromosomes are physically connected until metaphase I and then segregated from each other at the onset of anaphase I. In the subsequent second meiotic division (meiosis II), sister chromatids are segregated. Chromosomal abnormality arising during meiosis is one of the major causes of birth defects and congenital disorders in mammals including human and domestic animals. Hence understanding of the mechanism underlying these unique chromosome behavior in meiosis is of great importance. This review focuses on the roles of cohesin and condensin, and their regulation in chromosome dynamics during mammalian meiosis.

    SOCIETY REPRODUCTION & DEVELOPMENT-SRD, 2013年10月, JOURNAL OF REPRODUCTION AND DEVELOPMENT, 59 (5), 431 - 436, 英語

    [査読有り][招待有り]

    研究論文(学術雑誌)

  • Shangdan Xu, Jibak Lee, Masashi Miyake

    Expression of mRNAs and proteins of ZO-1 and occludin was analyzed in pig oocytes and parthenogenetic diploid embryos during preimplantation development using real-time RT-PCR, western blotting and immunocytochemistry. All germinal vesicle (GV) and metaphase (M)II oocytes and preimplantation embryos expressed mRNAs and proteins of ZO-1 and occludin. mRNA levels of both ZO-1 and occludin decreased significantly from GV to MII, but increased at the 2-cell stage followed by temporal decrease during the early and late 4-cell stages. Then, both mRNAs increased after compaction. Relative concentration of zo1 alpha(-) was highest in 2-cell embryos, while zo1 alpha(+) was expressed from the morula stage. Occludin expression greatly increased after the morula stage and was highest in expanded blastocysts. Western blotting analysis showed constant expression of ZO-1 alpha(-) throughout preimplantation development and limited translation of ZO-1 alpha(+) from the blastocysts, and species-specific expression pattern of occludin. Immunocytochemistry analysis revealed homogeneous distribution of ZO-1 and occludin in the cytoplasm with moderately strong fluorescence in the vicinity of the contact region between blastomeres, around the nuclei in the 2-cell to late 4-cell embryos, and clear network localization along the cell-boundary region in embryos after the morula stage. Present results show that major TJ proteins, ZO-1 and occludin are expressed in oocytes and preimplantation embryos, and that ZO-1 alpha(+) is transcribed by zygotic gene activation and translated from early blastocysts with prominent increase of occludin at the blastocyst stage.

    CAMBRIDGE UNIV PRESS, 2012年05月, ZYGOTE, 20 (2), 147 - 158, 英語

    [査読有り]

    研究論文(学術雑誌)

  • Jibak Lee, Sugako Ogushi, Mitinori Saitou, Tatsuya Hirano

    In many eukaryotes, condensins I and II associate with chromosomes in an ordered fashion during mitosis and play nonoverlapping functions in their assembly and segregation. Here we report for the first time the spatiotemporal dynamics and functions of the two condensin complexes during meiotic divisions in mouse oocytes. At the germinal vesicle stage (prophase I), condensin I is present in the cytoplasm, whereas condensin II is localized within the nucleus. After germinal vesicle breakdown, condensin II starts to associate with chromosomes and becomes concentrated onto chromatid axes of bivalent chromosomes by metaphase I. REC8 "glues" chromosome arms along their lengths. In striking contrast to condensin II, condensin I localizes primarily around centromeric regions at metaphase I and starts to associate stably with chromosome arms only after anaphase I. Antibody injection experiments show that condensin functions are required for many aspects of meiotic chromosome dynamics, including chromosome individualization, resolution, and segregation. We propose that the two condensin complexes play distinctive roles in constructing bivalent chromosomes: condensin II might play a primary role in resolving sister chromatid axes, whereas condensin I might contribute to monopolar attachment of sister kinetochores, possibly by assembling a unique centromeric structure underneath.

    AMER SOC CELL BIOLOGY, 2011年09月, MOLECULAR BIOLOGY OF THE CELL, 22 (18), 3465 - 3477, 英語

    [査読有り]

    研究論文(学術雑誌)

  • Jibak Lee, Tatsuya Hirano

    Cohesins are multi-subunit protein complexes that regulate sister chromatid cohesion during mitosis and meiosis. Here we identified a novel kleisin subunit of cohesins, RAD21L, which is conserved among vertebrates. In mice, RAD21L is expressed exclusively in early meiosis: it apparently replaces RAD21 in premeiotic S phase, becomes detectable on the axial elements in leptotene, and stays on the axial/lateral elements until mid pachytene. RAD21L then disappears, and is replaced with RAD21. This behavior of RAD21L is unique and distinct from that of REC8, another meiosis-specific kleisin subunit. Remarkably, the disappearance of RAD21L at mid pachytene correlates with the completion of DNA double-strand break repair and the formation of crossovers as judged by colabeling with molecular markers, gamma-H2AX, MSH4, and MLH1. RAD21L associates with SMC3, STAG3, and either SMC1 alpha or SMC1 beta. Our results suggest that cohesin complexes containing RAD21L may be involved in synapsis initiation and crossover recombination between homologous chromosomes.

    ROCKEFELLER UNIV PRESS, 2011年01月, JOURNAL OF CELL BIOLOGY, 192 (2), 263 - 276, 英語

    [査読有り]

    研究論文(学術雑誌)

  • Shunsuke Tate, Kazumi Nakamura, Chihiro Suzuki, Taichi Noda, Jibak Lee, Hiroshi Harayama

    The aim of this study is to provide evidence of the existence of the adenylyl cyclase 10 (ADCY-10) ortholog proteins in boar spermatozoa. Experiments with RT-PCR techniques, nucleotide sequence analyses and Northern blot analyses revealed that boar testes exclusively express approximately 5.1-kbp RNA, the nucleotide sequence of which is highly similar to that of human A DCY10. Database analyses with CDART suggested that pig ADCY10 ortholog proteins conserve two catalytic domains of adenylyl cyclase. Western blot techniques and indirect immunofluorescence with a specific antiserum to pig recombinant ADCY10 ortholog proteins showed that 48-kDa and 70-kDa truncated forms of pig ADCY10 ortholog proteins are localized in the equatorial segments and connecting pieces of boar ejaculated spermatozoa. Finally, cell imaging techniques with fluo-3/AM indicated that incubation with sodium bicarbonate (an ADCY10 activator) can initiate the calcium influx in the boar sperm heads that is controlled via the cyclic AMP signaling cascades. These results are consistent with the suggestion that functional ADCY10 ortholog proteins exist in the heads of boar spermatozoa. This is the first direct evidence of the existence of ADCY10 proteins in the heads of mammalian spermatozoa.

    SOCIETY REPRODUCTION & DEVELOPMENT-SRD, 2010年04月, JOURNAL OF REPRODUCTION AND DEVELOPMENT, 56 (2), 271 - 278, 英語

    [査読有り]

    研究論文(学術雑誌)

  • Mohammad Moniruzzaman, Jibak Lee, Mai Zengyo, Takashi Miyano

    Mammalian ovaries are endowed with a large number of primordial follicles, each containing a nongrowing oocyte. Only a small population of primordial oocytes (oocytes in primordial follicles) is activated to enter the growth phase throughout a female's reproductive life. Little is known about the mechanism regulating the activation of primordial oocytes. Here, we found that the primordial oocytes from infant pigs (10- to 20-day-old) grew to full size at 2 months after xenografting to immunodeficient mice, whereas those from prepubertal pigs (6-month-old) survived without initiation of their growth even after 4 months; thereafter, they started to grow and reached full size after 6 months. These results suggest that the mechanism regulating the activation of primordial oocytes in prepubertal pigs is different from that in infant pigs. in this regard, the involvement of FOXO3, a forkhead transcription factor, was studied. in prepubertal pigs, FOXO3 was detected in almost all (94 +/- 2%) primordial oocyte nuclei, and in infant pigs, 42 +/- 7% primordial oocytes were FOXO3 positive. At 4 months after xenografting, the percentage of FOXO3-positive primordial oocytes from prepubertal pigs had decreased to the infant level. Further, siRNA was designed to knock down porcine FOXO3. FOXO3-knockdown primordial follicles from prepubertal pigs developed to the antral stage accompanied by oocyte growth at 2 months after xenografting. These results suggest that primordial oocytes are dormant in prepubertal pigs by a FOXO3-related mechanism to establish a nongrowing oocyte pool in the ovary, and that a transient knockdown of the FOXO3 activates the primordial oocytes to enter the growth phase. Reproduction (2010) 139 337-348

    BIOSCIENTIFICA LTD, 2010年02月, REPRODUCTION, 139 (2), 337 - 348, 英語

    [査読有り]

    研究論文(学術雑誌)

  • Jibak Lee, Tomoya S. Kitajima, Yuji Tanno, Kayo Yoshida, Takashi Morita, Takashi Miyano, Masashi Miyake, Yoshinori Watanabe

    Reductional chromosome segregation in germ cells, where sister chromatids are pulled to the same pole, accompanies the protection of cohesin at centromeres from separase cleavage. Here, we show that mammalian shugoshin Sgo2 is expressed in germ cells and is solely responsible for the centromeric localization of PP2A and the protection of cohesin Rec8 in oocytes, proving conservation of the mechanism from yeast to mammals. However, this role of Sgo2 contrasts with its mitotic role in protecting centromeric cohesin only from prophase dissociation, but never from anaphase cleavage. We demonstrate that, in somatic cells, shugoshin colocalizes with cohesin in prophase or prometaphase, but their localizations become separate when centromeres are pulled oppositely at metaphase. Remarkably, if tension is artificially removed from the centromeres at the metaphase-anaphase transition, cohesin at the centromeres can be protected from separase cleavage even in somatic cells, as in germ cells. These results argue for a unified view of centromeric protection by shugoshin in mitosis and meiosis.

    NATURE PUBLISHING GROUP, 2008年01月, NATURE CELL BIOLOGY, 10 (1), 42 - U29, 英語

    [査読有り]

    研究論文(学術雑誌)

  • C. Sakai, F. Konno, O. Nakano, T. Iwai, T. Yokota, J. Lee, C. Nishida-Umehara, A. Kuroiwa, Y. Matsuda, M. Yamashita

    An interspecific hybrid medaka (rice fish) between Oryzias latipes and O. hubbsi is embryonically lethal. To gain an insight into the cellular and molecular mechanisms that cause the abnormalities occurring in the hybrid medaka, we investigated the behavior of chromosomes and the expression patterns of proteins responsible for the chromosome behavior. The number of chromosomes in the hybrid embryos gradually decreased to nearly half, since abnormal cell division with lagging chromosomes at anaphase eliminated the chromosomes from the cells. The chromosome lagging occurred at the first cleavage and continued throughout embryogenesis even after the midblastula transition. Fluorescent in-situ hybridization analyses revealed that the chromosomes derived from O. hubbsi are preferentially eliminated in both O. latipes-hubbsi and O. hubbsi-latipes embryos. Whole-mount immunocytochemical analyses using antibodies against alpha-tubulin, gamma-tubulin, inner centromere protein, Cdc20, Mad2, phospho-histone H3 and cohesin subunits (SMC1 alpha, SMC3 and Rad21) showed that the expression patterns of these proteins in the hybrid embryos are similar to those in the wild-type embryos, except for phospho-histone H3. Phospho-histone H3 present on chromosomes at metaphase was lost from normally separated chromosomes at anaphase, whereas it still existed on lagging chromosomes at anaphase, indicating that the lagging chromosomes remain in the metaphase state even when the cell has proceeded to the anaphase state. On the basis of these findings, we discuss the cellular and molecular mechanisms of chromosome elimination in the hybrid medaka.

    SPRINGER, 2007年10月, CHROMOSOME RESEARCH, 15 (6), 697 - 709, 英語

    [査読有り]

    研究論文(学術雑誌)

  • Evidence for existence of cAMP-Epac signaling in the heads of mouse epididymal spermatozoa

    Reiko Amano, Jibak Lee, Namiko Goto, Hiroshi Harayama

    Cyclic adenosine 3',5'-monophosphate (cAMP) signaling regulates the expression of fertilizing ability in mammalian spermatozoa. Many articles indicate that this signaling is mediated mainly via protein kinase A. Recently, a guanine nucleotide exchange factor for small G protein Rap1 (an exchange protein directly activated by cAMP: Epac) was discovered as a new mediator of cAMP signaling in somatic cells. The aim of this study was to reveal the existence of cAMP-Epac signaling in mouse spermatozoa. Northern blot analysis and in situ hybridization suggested that Epac1 and Epac2 mRNAs were transcribed in the seminiferous epithelia of the testis. This shows that expression of Epac mRNAs is present in mouse testicular germ cells. Indirect immunofluorescence with specific polyclonal antibodies suggested possible co-localization of Epac1 and Rap1 proteins in the heads of epididymal spermatozoa. Moreover, treatment of epididymal spermatozoa with an Epac-specific cAMP analog, 8-pMeCPT-2'-O-Me-cAMP, induced activation of Rap1, as revealed with a commercial kit for pull-down assay. These results indicate the existence of cAMP-Epac signaling in the heads of mouse epididymal spermatozoa.

    SOCIETY REPRODUCTION & DEVELOPMENT-SRD, 2007年02月, JOURNAL OF REPRODUCTION AND DEVELOPMENT, 53 (1), 127 - 133, 英語

    [査読有り]

    研究論文(学術雑誌)

  • Takashi Miyano, Sugako Ogushi, Hong-Thuy Bui, Jibak Lee

    Fully grown mammalian oocytes arrested at prophase I resume meiosis after gonadotropic stimulation. Oocytes undergo a series of changes including chromosome condensation, nucleolus disassembly, germinal vesicle breakdown (GVBD), and spindle formation. The mouse is the best model for studying the molecular mechanisms underlying the maturation of mammalian oocytes. However, some of the maturational events are different from those in other mammalian species. To study these events, we use pig ovaries, which are available as a byproduct from local slaughterhouses. It has long been known that pig oocytes have a dependence on de novo protein synthesis for GVBD, whereas GVBD in mouse oocytes occurs independently of protein synthesis. The reason seems to be the lack of Cyclin B1 molecules in pig GV-oocytes, although the synthesis of other protein(s) may be required for the GVBD. In mouse oocytes, the spindle is formed through the action of cytoplasmic microtubule organizing centers (MTOCs), and the oocytes are able to form the spindle without chromosomes. However, pig oocytes don't have such distinguished cytoplasmic MTOCs and never form the spindle without chromosomes. In this species, the condensing chromosome plays the role of the organizer nucleating spindle microtubules. We should develop some other mammalian models that will help us understand the mechanisms underlying oocyte maturation. © 2007, JAPANESE SOCIETY OF OVA RESEARCH. All rights reserved.

    2007年, Journal of Mammalian Ova Research, 24 (3), 92 - 98, 英語

    [査読有り]

    研究論文(学術雑誌)

  • Loss of Rec8 from chromosome arm and centromere region is required for homologous chromosome separation and sister chromatid separation, respectively, in mammalian meiosis

    Jibak Lee, Konosuke Okada, Sugako Ogushi, Takashi Miyano, Masashi Miyake, Masakane Yamashita

    Chromosome separation in meiosis I is different from those in mitosis and meiosis II in that homologs separate from each other in the former while sisters do so in the latter. We show here that meiosis-specific cohesin subunit Rec8 in mouse oocytes shows essentially the same pattern of localization to those reported in yeasts and mammalian spermatocytes; Rec8 along chromosome arm ( armRec8) is lost at the metaphase I-to-anaphase I transition, although centromeric Rec8 (cenRec8) is maintained until the onset of anaphase II. Suppression of the loss of armRec8 by microinjection of anti-Rec8 antibody into the oocytes inhibits homolog separation but not the first polar body emission ( cytokinesis). Similarly, the injection of anti-Rec8 antibody into metaphase II oocytes prevents sister separation in anaphase II after oocyte activation. These data demonstrate that the loss of armRec8 and cenRec8 is required for separation of homologs and sisters, respectively, but both are not required for other late mitotic events such as spindle elongation and cytokinesis in mouse oocytes. Further, by using some inhibitors for spindle assembly, proteasome and Topoisomerase II and overexpression of Securin, we propose that loss of armRec8 ( homolog separation) and cytokinesis are suppressed until anaphase I by Securin whose destruction is regulated by spindle checkpoint-proteasome pathway, and that Topoisomerase II is required for homolog separation independently from such pathway.

    LANDES BIOSCIENCE, 2006年07月, CELL CYCLE, 5 (13), 1448 - 1455, 英語

    [査読有り]

    研究論文(学術雑誌)

  • T Iwai, J Lee, A Yoshii, T Yokota, K Mita, M Yamashita

    Until the onset of anaphase, sister chromatids are bound to each other by a multi-subunit protein complex called cohesin. Since chromosomes in meiosis behave differently from those in mitosis, the cohesion and separation of homologous chromosomes and sister chromatids in meiosis are thought to be regulated by meiosis-specific cohesin subunits. Actually, several meiosis-specific cohesin subunits, including Rec8, STAG3 and SMC1beta, are known to exist in mammals; however, there are no reports of meiosis-specific cohesin subunits in other vertebrates. To investigate the protein expression and localization of cohesin subunits during meiosis in non-mammalian species, we isolated cDNA clones encoding SMC1alpha, SMC1beta, SMC3 and Rad21 in the medaka and produced antibodies against recombinant proteins. Medaka SMC1beta was expressed solely in gonads, while SMC1alpha, SMC3 and Rad21 were also expressed in other organs and in cultured cells. SMC1beta forms a complex with SMC3 but not with Rad21, in contrast to SMC1alpha, which forms complexes with both SMC3 and Rad21. SMC1alpha and Rad21 were mainly expressed in mitotically dividing cells in the testis (somatic cells and spermatogonia), although their weak expression was detected in pre-leptotene spermatocytes. SMC1beta was expressed in spermatogonia and spermatocytes. SMC1beta was localized along the chromosomal arms as well as on the centromeres in meiotic prophase 1, and its existence on the chromosomes persisted up to metaphase H, a situation different from that reported in the mouse, in which SMC1beta is lost from the chromosome arms in late pachytene despite its universal presence in vertebrates. (C) 2004 Elsevier B.V. All rights reserved.

    ELSEVIER SCIENCE BV, 2004年09月, GENE EXPRESSION PATTERNS, 4 (5), 495 - 504, 英語

    [査読有り]

    研究論文(学術雑誌)

  • Takashi Miyano, Jibak Lee, Josef Fulka

    Wiley, 2003年09月, Reproductive Medicine and Biology, 2 (3), 91 - 99

    [査読有り]

    研究論文(学術雑誌)

  • J Lee, T Iwai, T Yokota, M Yamashita

    Sister chromatid cohesion is maintained from DNA replication to metaphase-to-anaphase transition by multisubunit protein complexes called cohesin, which include at least four proteins, SMC1alpha, SMC3, Rad21. and either SA1 or SA2, in mammalian somatic cells. We report here the first evidence of the involvement of Rec8 protein, a mammalian homolog of yeast Rec8p, in meiosis-specific chromosome behavior in mammals. In immunoblotting and immunohistochemical analysis using specific antibodies against mouse Rec8, we found that Rec8 was expressed in the testis but not in the kidney or liver; more precisely, it was expressed in spermatocytes and spermatids but not in spermatogonia or other somatic cells. We also found that Rec8 is present in both phosphorylated and dephosphorylated states in vivo. Immunoprecipitation analyses revealed that Rec8 associates with other cohesin proteins, SMC1beta (meiosis-specific protein) and SMC3 and with a component of synaptonemal complexes, SCP3, but not with SMC1alpha. In meiotic chromosome spreads, Rec8 was localized along the axial/lateral elements of the synaptonemal complexes in meiotic prophase from the leptotene to diplotene stages. At later stages, diakinesis and metaphase I, Rec8 was localized along the interstitial axes of chromosomes, including both centromere and arm regions of chromosomes. However, concomitantly with separation of homologous chromosomes in anaphase I, Rec8 was no longer detected along the arm regions, while it persisted on centromere regions up to metaphase II. In anaphase II, the centromeric signals were diminished. We propose from these results that mammalian Rec8 protein, in association with SMC3 and SMC1beta but not SMC1alpha, is involved in meiosis-specific chromosome behavior, and that homologous chromosome separation is triggered by selective loss of Rec8 from chromosome arms in meiosis I, while sister chromatid cohesion is maintained until metaphase II/anaphase II transition by centromeric Rec8 during mammalian meiosis.

    COMPANY OF BIOLOGISTS LTD, 2003年07月, JOURNAL OF CELL SCIENCE, 116 (13), 2781 - 2790, 英語

    [査読有り]

    研究論文(学術雑誌)

  • N Kanayama, T Miyano, J Lee

    Meiotic maturation of mammalian oocytes is under the control of cell cycle molecules Cdc2 kinase and MAP kinase (mitogen-activated protein kinase). In the present study, we investigated the relationship between the ability to activate Cdc2 kinase and MAP kinase and the acquisition of meiotic competence during pig oocyte growth. Growing and fully grown pig oocytes were collected from four groups of antral follicles of various diameters (A, 0.5-0.7 mm; B, 1.0-1.5 mm; C, 2.0-2.5 mm; D, 4.0-6.0 mm) and cultured in vitro. Fully grown oocytes from class D follicles, which have full competence to mature to metaphase II, had the ability to activate both Cdc2 kinase and MAP kinase. In contrast, growing oocytes from class A follicles, which have limited competence to resume meiosis, had no such ability. Cyclin B1 molecules did accumulate, however, with phosphorylated 35 and 36 kDa bands of p34(cdc2) appearing in the cultured oocytes. Of the growing oocytes from class B follicles, 60% resumed meiosis but arrested at metaphase I. Some of the oocytes in this class were capable of activating Cdc2 kinase, although they did not appear to have established a MAP kinase-activating pathway or the ability to activate MEK. These results suggest that limited meiotic competence in growing oocytes from class A follicles is due to their inability to activate Cdc2 kinase and their incomplete MEK-MAP-kinase pathway, although the oocytes are capable of accumulating cyclin B1 molecules. During the final growth phase, pig oocytes acquire the ability to activate Cdc2 kinase and then establish the MEK-MAP-kinase pathway for full meiotic competence.

    CAMBRIDGE UNIV PRESS, 2002年08月, ZYGOTE, 10 (3), 261 - 270, 英語

    [査読有り]

    研究論文(学術雑誌)

  • Analyses of mRNA expression patterns of cohesin Subunits rad21 and rec8 in mice: Germ cell-specific expression of rec8 mRNA in both male and female mice

    J Lee, T Yokota, M Yamashita

    A multisubunit protein complex called cohesin is required for the cohesion between sister chromatids in both mitosis and meiosis in yeast. We investigate here the mRNA expression patterns of mouse homologues of the yeast mitotic cohesin rad21 and the meiotic cohesin rec8 in various organs, with special attention to their expression in gonads. Northern blot and reverse transcription-polymerase chain reaction (RT-PCR) analyses revealed that, in contrast to the ubiquitous expression of rad21 mRNA in all of the organs examined, rec8 was expressed only in the gonads. We conducted in situ hybridization analysis to identify the cells that express rad21 and rec8 mRNAs in the gonads. In the testis, rad21 mRNA was expressed in somatic cells and spermatogonia but not in spermatocytes, and conversely, rec8 mRNA was expressed in spermatocytes but not in spermatogonia or somatic cells. Spermatids expressed rad21 and rec8 mRNAs simultaneously. In the ovary, rad21 rnRNA was detected in all of the ovarian cells including germ cells and somatic cells, whereas rec8 mRNA was detected only in oocytes. Unlike the widespread expression of rad21 gene, therefore, the gene expression of rec8 is strictly confined to spermatocytes and spermatids in male mouse and oocytes in female mouse. The restricted expression pattern of rec8 mRNA implies its essential role in meiosis in both sexes of mammals, as has been reported in yeast. We also discuss the cooperative functions of Rad21 and Rec8 on the basis of the finding that their mRNAs are coexpressed in oocytes and spermatids.

    ZOOLOGICAL SOC JAPAN, 2002年05月, ZOOLOGICAL SCIENCE, 19 (5), 539 - 544, 英語

    [査読有り]

    研究論文(学術雑誌)

  • Spindle formation and dynamics of γ-tubulin and NuMA distribution during meiosis in pig and mouse oocytes

    Lee J, Miyano T, Moor RM

    This work focuses on the assembly and transformation of the spindle during the progression through the meiotic cell cycle. For this purpose, immunofluorescent confocal microscopy was used in comparative studies to determine the spatial distribution of alpha- and gamma-tubulin and nuclear mitotic apparatus protein (NuMA) from late G2 to the end of M phase in both meiosis and mitosis. In pig endothelial tells, consistent with previous reports, gamma-tubulin was localized at the centrosomes in both interphase and M phase, and NuMA was localized in the interphase nucleus and at mitotic spindle poles. During meiotic progression in pig oocytes, gamma-tubulin and NuMA were initially detected in a uniform distribution across the nucleus. In early diakinesis and just before germinal vesicle breakdown, microtubules were first detected around the periphery of the germinal vesicle and cell cortex. At rate diakinesis, a mass of multi-arrayed microtubules was formed around chromosomes. In parallel, NuMA localization changed from an amorphous to a highly aggregated form in the vicinity of the chromosomes, but gamma-tubulin localization remained in an amorphous form surrounding the chromosomes. Then the NuMA foci moved away from the condensed chromosomes and aligned at both poles of a barrel-shaped metaphase I spindle white gamma-tubulin was localized along the spindle microtubules, suggesting that pig meiotic spindle poles are formed by the bundling of microtubules at the minus ends by NuMA. interestingly, in mouse oocytes, the meiotic spindle pole was composed of several gamma-tubulin foci rather than NuMA. Further, nocodazole, an inhibitor of microtubule polymerization, induced disappearance of the pole staining of NuMA in pig metaphase ii oocytes, whereas the mouse meiotic spindle pole has been reported to be resistant to the treatment. These results suggest that the nature of the meiotic spindle differs between species. The axis of the pig meiotic spindle rotated from a perpendicular to a parallel position relative to the cell surface during telophase I. Further, in contrast to the stable localization of NuMA and gamma-tubulin at the spindle poles in mitosis, NuMA and gamma-tubulin became relocalized to the spindle midzone during anaphase I and telophase I in pig oocytes. We postulate that in the centrosome-free meiotic spindle, NuMA aggregates the spindle microtubules at the midzone during anaphase and telophase and that the polarity of meiotic spindle microtubules might become inverted during spindle elongation.

    SOC STUDY REPRODUCTION, 2000年05月, Biol Reprod, 62 (5), 1184 - 1192, 英語

    [査読有り]

    研究論文(学術雑誌)

  • Specific regulation of CENP-E and kinetochores during meiosis I/meiosis II transition in pig oocytes

    J Lee, T Miyano, YF Dai, P Wooding, TJ Yen, RM Moor

    To understand the mechanisms which regulate meiosis-specific cell cycle and chromosome distribution in mammalian oocytes, the level and the localization of CENP-E and the kinetochore number and direction on a half bivalent were examined during pig oocyte maturation. CENP-E is a kinetochore motor protein whose intracellular level and localization are strictly regulated in the somatic cell cycle. The localizations of CENP-E on meiotic chromosomes from diakinesis stage to anaphase I and at the spindle midzone at telophase I were shown by immunofluorescent confocal microscopy to be similar to those in somatic cells of pig and other species. Further, ultrastructural analysis revealed the presence of CENP-E on fibrous corona and outer plate of kinetochores of the meiotic chromosomes. However, unlike mitosis, CENP-E staining was continuously detected either at the spindle midzone or on the kinetochores of segregated chromosomes during the first polar body emission. Consistent with this, immunoblot analysis revealed that CENP-E level remained high during meiosis l/meiosis II (MI/MII) transition and that some of CENP-E survived through the transition even in cycloheximide-treated oocytes in which cyclin B1 was completely degraded. Furthermore, examinations of CENP-E signals in confocal microscopy and kinetochores in electron microscopy in MI and MH oocytes provide the cytological evidence in mammalian oocytes which suggests that each sister chromatid in a pair has its own kinetochore which localizes side-by-side so that two sister chromatids on a half bivalent are oriented toward and connected to the same pole in M1. (C) 2000 Wiley-Liss, Inc.

    WILEY-LISS, 2000年05月, MOLECULAR REPRODUCTION AND DEVELOPMENT, 56 (1), 51 - 62, 英語

    [査読有り]

    研究論文(学術雑誌)

  • Localisation of phosphorylated MAP kinase during the transition from meiosis I to meiosis II in pig oocytes

    J Lee, T Miyano, RM Moor

    Mitogen-activated protein kinase (MAPK) has been reported to be involved in oocyte maturation in all animals so far examined. In the present study we investigate the expression and localisation of active phosphorylated MAPKs (p44(ERK1)/p42(ERK2)) during maturation of pig oocytes. In immunoblot analysis using anti-p44(ERK1) antibody which recognised both active and inactive forms of p44(ERK1) and p42(ERK2),we confirmed that MAPKs were phosphorylatred around the time of germinal vesicle breakdown (GVBD) and the active phosphorylated MRPKs (pMAKs) were maintained until metaphase II, as has been reported. On immunofluorescent confocal microscopy using anti-pMAPK antibody which recognised only phosphorylated forms of MAPKs, pMAPK was localised at the spindle poles in pig mitotic cells. On the other hand, in pig oocytes, no signal was detected during GV stage. After GVBD, the area around condensed chromosomes was preferentially stained at metaphase I although whole cytoplasm was faintly stained. nt early anaphase I, the polar regions of the meiotic spindle were prominently stained. However, during the progression of anaphase I and telophase I pMAPK was detected at the mid-zone of the elongated spindle, gradually becoming concentrated at the centre. Finally, at the time of emission of the first polar body, pMAPK was detected as a ring-like structure between the condensed chromosomes and the first polar body, and the staining was maintained even after the metaphase II spindle was formed. The inhibition of MAPK activity with the MAPK kinase inhibitor U0126 during the meiosis I/meiosis II transition suppressed chromosome separation, first polar body emission and formation of the metaphase II spindle. From these results, we propose that the spindle-associated pMAPKs play an important role in the events occurring during the meiosis I/meiosis II transition, such as chromosome separation, spindle elongation and cleavage furrow formation in pig oocytes.

    CAMBRIDGE UNIV PRESS, 2000年05月, ZYGOTE, 8 (2), 119 - 125, 英語

    [査読有り]

    研究論文(学術雑誌)

  • Degradation of pig cyclin B1 molecules precedes MAP kinase dephosphorylation during fertilisation of the oocytes

    T Miyano, YF Dai, J Lee, K Kano, RM Moor

    Pig oocytes at metaphase II were activated. by penetration of spermatozoa in cycloheximide-free and cycloheximide-containing fertilisation media. The precise nuclear stage, and the kinetics of degradation of cyclin B1 and dephosphorylation of MAP kinase were assessed after insemination. After maturation culture, 96% of oocytes reached metaphase II. At 6 h after insemination in cycloheximide-free medium, 68% of the oocytes were activated and had progressed to anaphase II or beyond. After 8 h, 89% of the oocytes were activated: a female pronucleus had formed and the heads of penetrating spermatozoa had enlarged and changed to male pronuclei. In the cycloheximide-containing medium, activation of oocytes started earlier than in cycloheximide-free medium. After 2 h, 43% of the oocytes were activated, and the percentage increased to 97% after 6 h. Pig cyclin B1 disappeared in the oocytes at 6 h after insemination in both cycloheximide-containing and cycloheximide-free media. Pig oocytes at metaphase II contained two types of MAP kinase - ERK 1 and ERK 2 - in their active phosphorylated forms. At 8 h after insemination ERE; 2 changed to the fast-migrating inactive form in the oocytes cultured in both cycloheximide-containing and cycloheximide-free media, although the shift-down was not complete. The change was delayed by 2 h after the degradation of cyclin B1 molecules. These results demonstrate that degradation of pig cyclin B1 molecules corresponds to the transition of the oocytes from metaphase II arrest to anaphase II/telophase II and was followed by MAP kinase dephosphorylation.

    CAMBRIDGE UNIV PRESS, 2000年05月, ZYGOTE, 8 (2), 153 - 158, 英語

    [査読有り]

    研究論文(学術雑誌)

  • Tyrosine phosphorylation of p34(cdc2) in metaphase II-arrested pig oocytes results in pronucleus formation without chromosome segregation

    J Lee, K Hata, T Miyano, M Yamashita, YF Dai, RM Moor

    At the G2-M boundary, maturation-promoting factor (MPF) activation is usually induced in one or both of two ways; tyrosine dephosphorylation of p34(cdc2) Or synthesis of cyclin B according to cell type and species. At the end of M-phase, however, MPF inactivation is normally triggered only by cyclin degradation. We investigated whether tyrosine phosphorylation of p34(cdc2) can inactivate MPF and what kinds of events are induced in pig metaphase II (MII)-arrested oocytes. First, cyclin B1 in MII-arrested oocytes is degraded upon fertilization. Second, when MII oocytes were treated with vanadate, an inhibitor of tyrosine phosphatases, they were released from MII arrest, but MPF was inactivated by further tyrosine phosphorylation of p34(cdc2) rather than cyclin B1 degradation. The vanadate-induced exit from M-phase is distinct from normal M-phase exit, which is accompanied by cyclin B1 degradation; the former lacks both sister chromatid separation and second polar body emission. Vanadate itself has no inhibitory effect on chromosome segregation since calcium ionophore induced chromosome segregation in the presence of vanadate. Furthermore, when MII oocytes were treated with olomoucine, an inhibitor of cyclin-dependent kinases, they exited from MII arrest in a manner similar to vanadate-treated MII oocytes. Finally, we propose that MPF inactivation by tyrosine phosphorylation of p34(cdc2) enables MII oocytes to form an interphase nucleus, but not to segregate sister chromatid due to the absence of the mechanisms required to trigger sister chromatid separation such as anaphase-promoting complex (APC)-mediated proteolysis, on the signaling pathway from intracellular Ca2+ increase to MPF inactivation, Mol. Reprod. Dev. 52:107-116, 1999. (C) 1999 Wiley-Liss, Inc.

    WILEY-LISS, 1999年01月, MOLECULAR REPRODUCTION AND DEVELOPMENT, 52 (1), 107 - 116, 英語

    [査読有り]

    研究論文(学術雑誌)

MISC

  • 卵胞発達に伴うウシ壁顆粒膜細胞の卵母細胞発育支持能力の変化

    伏井 実穂子, 京極 博久, 李 智博, 宮野 隆

    (公社)日本畜産学会, 2022年09月, 日本畜産学会大会講演要旨集, 130回, 112 - 112, 日本語

  • 卵胞発達に伴うウシ顆粒膜細胞のTranszonal Projection形成能の変化

    伏井 実穂子, 京極 博久, 李 智博, 宮野 隆

    (一社)日本生殖医学会, 2022年07月, 日本生殖医学会雑誌, 67 (3), 134 - 134, 日本語

  • 美 栄, 松田 厚志, 平岡 泰, 李 智博

    【目的】第一減数分裂前期には,三層の軸構造からなるシナプトネマ複合体が形成され,その上で相同染色体の対合や組換えが進行する。減数分裂型コヒーシンサブユニットであるRAD21LやREC8は,シナプトネマ複合体の軸構造 (axial element: AE) の形成に必須であることが示されている。しかし,どのようにこの両者が相同染色体の対合・組換えやシナプトネマ複合体の形成にかかわるか,その詳細なメカニズムや役割分担については不明である。そこで本研究では,これらのことを明らかにするために,第一減数分裂前期におけるRAD21LやREC8のシナプトネマ複合体上のより詳細な局在位置の解析を試みた。【方法】C57BL/6系統マウスの精巣を採取し,コラゲナーゼ処理などにより,細胞を分散させた。精巣由来細胞を2%パラホルムアルデヒドで固定後,RAD21LやREC8に対する抗体と,AEの構成分子であるSYCP3に対する抗体,あるいはDNA修復関連タンパク質であるRAD51やMSH4に対する抗体により,共免疫染色した。サンプルを高解像度の次世代光学顕微鏡システム (3D-SIM) で観察した。【結果】3D-SIMを用いて,対合状態の2本のAEをSYCP3のシグナルとして観察できた。RAD21LやREC8はAE上に非連続的に存在した。第一減数分裂前期のパキテン期において,2本のAE上のSYCP3間あるいは RAD21L間の距離の測定や局在を比較解析したところ,RAD21LはSYCP3と部分的に共局在するが,2本の AEのより内側に存在することが分かった。また,2本のAE間を架橋するような RAD21Lのシグナルも観察されたことから,RAD21Lは,AEの内側で相同染色体から伸びるクロマチンループを接着することにより,対合に関与すると考えられる。さらに,RAD21LやREC8,MSH4やRAD51 の二重免疫染色の解析から得られた結果について紹介し,組換えにおけるRAD21LやREC8の役割について検討する。

    日本繁殖生物学会, 2013年, 日本繁殖生物学会 講演要旨集, 106, P - 30-P-30, 日本語

  • 渋谷 海大, 李 智博, 三宅 正史

    【目的】着床前胚のグルコース(Glu)要求性は種によって異なる。マウス胚において培地へのGlu添加は,初期卵割期胚の発生を抑制するが,胚盤胞形成には必須である。そのためGlu不含培地では,胚盤胞を形成せずに退行する。一方ブタ胚はGluを含まないPZM-3培地において,受精卵,単為発生2倍体ともに,胚盤胞まで高率に発生する。マウス胚とブタ胚における,このようなGlu代謝の違いは,糖新生の有無に起因すると仮定した。つまり,ブタ着床前胚では糖新生が行われているために,培地へのGlu添加を必要としないと考えた。本研究では,着床前のブタ胚が糖新生能力を持つかどうかを明らかにするために,着床前の各発生段階にあるブタ単為発生2倍体において,糖新生の不可逆的反応に関わる酵素[ホスホエノールピルビン酸カルボキシキナーゼ (PEPCK), ピルビン酸カルボキシラーゼ (PC), フルクトース-1,6-ビスホスファターゼ (FBP), グルコース-6-ホスファターゼ (G6P)]の発現を調べた。【方法】直径4~6 mmの卵胞からGV卵を採取し,44~46時間成熟培養後,MII卵を活性化させ,サイトカラシンB処理によって単為発生2倍体を作出した。2倍体をPZM-3で培養し,活性化後24, 48, 72, 96, 120, 144時間に,それぞれ48~50個の正常胚を回収した。これらの2倍体胚,および50個のMII期卵母細胞からtotal RNAを抽出した。抽出したRNAを元にcDNAを合成し,RT-PCR法を用いて糖新生関連酵素(PEPCK, PC, FBP, G6P)のmRNA発現を調べた。【結果】MII期卵母細胞,ならびに2細胞から拡張胚盤胞期まですべての胚において,PEPCK, PC, FBP, G6PのmRNA発現が認められた。以上の結果から,哺乳動物のMII期卵母細胞,および胚盤胞までのすべての発生段階の活性化2倍体で糖新生を行えることが示唆された。

    日本繁殖生物学会, 2012年, 日本繁殖生物学会 講演要旨集, 105, 1010 - 1010

  • DETECTION OF SOLUBLE ADENYLYL CYCLASE (ADCY10) HOMOLOG PROTEINS IN BOAR SPERMATOZOA

    Kazumi Nakamura, Chihiro Suzuki, Shunsuke Tate, Jibak Lee, Hiroshi Harayama

    AMER SOC ANDROLOGY, INC, 2009年03月, JOURNAL OF ANDROLOGY, 37 - 37, 英語

    研究発表ペーパー・要旨(国際会議)

  • Localization of Claudin family proteins in pig embryos during pre-implantation development

    S. Xu, J. Lee, H. Harayama, M. Miyake

    CSIRO PUBLISHING, 2007年, REPRODUCTION FERTILITY AND DEVELOPMENT, 19 (1), 197 - 197, 英語

    研究発表ペーパー・要旨(国際会議)

  • The expression and localization of cohesin subunits in the gonads of medaka, Oryzias latipes

    Toshiharu Iwai, Jibak Lee, Atsushi Yoshii, Takehiro Yokota, Koichi Mita, Masakane Yamashita

    ZOOLOGICAL SOC JAPAN, 2004年12月, ZOOLOGICAL SCIENCE, 21 (12), 1288 - 1288, 英語

    [査読有り]

    研究発表ペーパー・要旨(国際会議)

  • Growth of oocytes in KIT-deficient Fas-knockout mice

    M. MONIRUZZAMAN, K. SAKAMAKI, Y. AKAZAWA, J. LEE, T. MIYANO

    2004年11月, Kobe University the 21st Century COE Program Symposium

  • Involvement of meiosis-specific cohesin rec8 in chromosome cohesion during pig gametogenesis.

    J Lee, M Muroga, S Ogushi, T Miyano, M Miyake

    SOC STUDY REPRODUCTION, 2004年, BIOLOGY OF REPRODUCTION, 188 - 188, 英語

    研究発表ペーパー・要旨(国際会議)

  • マウス減数分裂におけるRec8蛋白質の発現解析

    李智博, 岩井俊治, 横田雄洋, 山下正兼

    2003年, 日本畜産学会大会講演要旨, 101st

  • POSSIBLE INVOLVEMENT OF PROTEASES IN FERTILIZATION OF THE MEDAKA FISH(Developmental Biology)(Proceedings of the Seventy-Third Annual Meeting of the Zoological Society of Japan) :

    Iwai Toshiharu, Ogiwara Katsueki, Kotani Tomoya, Lee Jibak, Mita Koichi, Takahashi Takayuki, Yamashita Masakane

    Zoological Society of Japan, 2002年, Zoological science, 19 (12), 1442 - 1442, 英語

書籍等出版物

  • 繁殖生物学 (改定版) 日本繁殖生物学会 [編]

    李 智博;宮野 隆

    分担執筆, 第2章-1 生殖細胞, インターズー, 2020年03月, 日本語, ISBN: 9784866711102

  • “Oocytes -Maternal information and functions” Malgorzata Kloc (Ed)

    Jibak Lee

    分担執筆, Chapter 15: The regulation and function of cohesin and condensin in mammalian oocytes and spermatocytes. pp.355-372., Springer, Results and Problems in Cell Differentiation 63, 2017年08月, 英語

    学術書

  • 『染色体と細胞核のダイナミクス DNAを操る細胞の仕組み』平岡泰・原口徳子 編

    李 智博, 平野 達也

    分担執筆, 第7章「コヒーシンとコンデンシン」, 化学同人, 2013年, 日本語

    学術書

  • 「分子細胞生物学辞典(第2版)」篠崎一雄, 清水孝雄, 谷口克, 月田承一郎, 西村善文, 林崎良英, 御子柴克彦, 柳田充弘, 米田悦啓 編

    李 智博, 平野 達也

    分担執筆, 東京化学同人, 2008年, 日本語

    事典・辞書

  • “Reproductive Biotechnology: Reproductive Biotechnology and its Related Physiology” Miyamoto, and N. Manabe (eds.)

    Takashi Miyano, Jibak Lee

    分担執筆, Change in cyclin B1 and MAP kinase molecules during maturation and fertilization of pig oocytes. pp. 9-19, Hokuto Shobo, 2001年

  • “Gametes: Development and Function” A. Lauria, F. Gandolfi, G. Enne and L. Gianaroli (eds.)

    Dai, Y, Zhang, J, Lee, J, Moor, RM

    分担執筆, Molecules involved in the metaphase I to metaphase II transition in mammalian oocytes, pp.155-175, SeronoSymposia, 1998年

講演・口頭発表等

  • Antrum formation in bovine oocyte-cumulus cell complexes requires participation of oocytes via GDF9 and BMP15

    Alam MH, Lee J, Miyano T

    第111回日本繁殖生物学会大会, 2018年09月, 英語, 信州大学繊維学部, 国内会議

    口頭発表(一般)

  • How do different cohesins contribute to the connection between homologs in mammalian meiosis?

    李 智博

    蛋白研セミナー Genome stability and instability in mitotic and meiotic cells, 2018年04月, 英語, 大阪大学蛋白質研究所, 国際会議

    [招待有り]

    口頭発表(招待・特別)

  • GDF9 and BMP15 promote antrum formation by bovine cumulus cells in vitro

    Alam Md, Hasanur, Lee J, Miyano T

    日本畜産学会第124回大会, 2018年03月, 英語, 東京大学, 国内会議

    口頭発表(一般)

  • Meiotic cohesins during spermatogenesis

    李 智博

    4th World Congress of Reproductive Biology, 2017年09月, 英語, Ginowan City, Okinawa, Japan, 国際会議

    [招待有り]

    口頭発表(招待・特別)

  • Ectopic expression of RAD21L-containing cohesin brings homologous chromosomes closer in somatic cells

    Rong Mei, Miyauchi Sachi, 李 智博

    4th World Congress of Reproductive Biology, 2017年09月, 英語, Ginowan City, Okinawa, Japan, 国際会議

    ポスター発表

  • Effects of ectopic expression of meiotic cohesin subunit RAD21L in somatic cells

    Miyauchi Sachi, Rong Mei, 李 智博

    The 2nd meeting on SMC proteins, 2017年06月, 英語, Nanyo City, Yamagata, Japan, 国際会議

    ポスター発表

  • 減数分裂型コヒーシンサブユニットRAD21Lの体細胞への異所発現の影響

    宮内 紗千, 美 栄, 李 智博

    日本畜産学会第122回大会, 2017年03月, 日本語, 神戸, 国内会議

    口頭発表(一般)

  • The detailed localization of meiotic cohesin subunits, REC8 and RAD21L, in mouse spermatocytes

    美 栄, 松田 厚志, 平岡 泰, 李 智博

    World Congress of Reproductive Biology 2014, 2014年09月, 英語, 国際会議

    ポスター発表

  • Potential roles of condensins during mouse oocyte maturation and early embryo genesis

    李 智博

    Departmental Seminar, Department of Biochemstry, Oxford University, 2014年09月, 英語, Oxford, UK, 国際会議

    公開講演,セミナー,チュートリアル,講習,講義等

  • Essential roles of condensins in chromosome organization during mouse early embryogenesis

    李 智博, 清水 萌子, 西出 賢次, 平野 達也

    World Congress of Reproductive Biology 2014, 2014年09月, 英語, Edinburgh, 国際会議

    ポスター発表

  • ブタ胚盤胞におけるタイトジャンクション関連タンパク質claudin familyの発現性

    三宅 正史, 董 哲, 渋谷 海大, 李 智博, 原山 洋, 宮野 隆

    第107回日本繁殖生物学会大会, 2014年08月, 日本語, 日本繁殖生物学会, 帯広, 国内会議

    口頭発表(一般)

  • マウス初期胚におけるコンデンシンサブユニットSMC2の機能解析

    清水 萌子, 西出 賢次, 平野 達也, 李 智博

    日本畜産学会第114回大会, 2014年03月, 日本語, つくば, 国内会議

    口頭発表(一般)

  • マウス精母細胞におけるコヒーシンの解析

    李 智博, 美 栄

    第45回精子研究会・神戸大学重点研究チーム学術講演会, 2014年01月, 日本語, 国内会議

    [招待有り]

    シンポジウム・ワークショップパネル(指名)

  • マウス発育前卵母細胞におけるコヒーシンサブユニットRAD21LおよびRAD21の発現解析

    松浦 倫子, 李 智博

    第106回日本繁殖生物学会大会, 2013年, 日本語, 国内会議

    ポスター発表

  • マウス精母細胞における減数分裂型コヒーシンの詳細な局在位置

    美 栄, 松田 厚志, 平岡 泰, 李 智博

    第106回日本繁殖生物学会大会, 2013年, 日本語, 国内会議

    ポスター発表

  • マウス減数分裂における染色体動態の制御機構

    李 智博

    分子細胞生物学セミナー,北海道大学先端生命科学研究院, 2013年, 日本語, 国内会議

    公開講演,セミナー,チュートリアル,講習,講義等

  • 着床前発生過程のブタ単為発生2倍体における糖新生関連酵素の発現

    渋谷 海大, 李 智博, 三宅 正史

    第105回日本繁殖生物学会大会, 2012年, 日本語, 国内会議

    ポスター発表

  • 減数分裂前期における染色体上のコヒーシンの位置

    李 智博

    特定領域「生殖系列の世代サイクルとエピゲノムネットワーク」第5回公開シンポジウム, 2012年, 日本語, 国内会議

    [招待有り]

    シンポジウム・ワークショップパネル(指名)

  • マウス卵母細胞の二価染色体構築におけるコンデンシンIとIIの役割

    李 智博, 大串 素雅子, 斎藤 通紀, 平野 達也

    第29回染色体ワークショップ, 2012年, 日本語, 国内会議

    口頭発表(一般)

  • なぜコヒーシンとコンデンシンなのか?

    李 智博

    特定領域「生殖系列の世代サイクルとエピゲノムネットワーク」生殖サイクル若手勉強会2012, 2012年, 日本語, 国内会議

    口頭発表(一般)

  • Regulation of meiotic chromosome dynamics by cohesin, condensin, and shugoshin in mouse oocytes

    LEE Jibak

    EMBO Workshop “Cell Biology of Early Mouse Development”, 2012年, 英語, 国際会議

    [招待有り]

    シンポジウム・ワークショップパネル(指名)

  • 初期ブタ胚培養培地に対するグルコースおよびフルクトー添加の影響と利用性

    渋谷 海大, 李 智博, 三宅 正史

    第18回日本胚移植研究会, 2011年, 日本語, 国内会議

    口頭発表(一般)

  • 減数分裂特異的な新規コヒーシンサブユニットRAD21Lの動態

    李 智博, 平野 達也

    第104回日本繁殖生物学会大会, 2011年, 日本語, 国内会議

    口頭発表(一般)

  • 減数分裂期コヒーシンと相同染色体の対合・組換え

    李 智博

    特定領域「生殖系列の世代サイクルとエピゲノムネットワーク」第4回公開シンポジウム, 2011年, 日本語, 国内会議

    [招待有り]

    シンポジウム・ワークショップパネル(指名)

  • 減数分裂期コヒーシンとエピジェネティクスとの関係

    李 智博

    特定領域「生殖系列の世代サイクルとエピゲノムネットワーク」生殖サイクル若手勉強会2011, 2011年, 日本語, 国内会議

    口頭発表(一般)

  • ブタ単為発生2倍体の着床前発生におけるclaudin-1の役割

    許 尚丹, 李 智博, 三宅 正史

    日本畜産学会第114回大会, 2011年, 日本語, 国内会議

    ポスター発表

  • タイトジャンクションタンパク質,Claudin-1,-2のブタ着床前発生における発現動向

    許 尚丹, 李 智博, 三宅 正史

    第61回関西畜産学会大会, 2011年, 日本語, 国内会議

    ポスター発表

  • Regulation of chromosome dynamics by cohesins in mammalian meiosis

    LEE Jibak, HIRANO Tatsuya

    Czech–Japan Joint Symposium for Animal Reproduction, 2010年, 英語, 国際会議

    [招待有り]

    シンポジウム・ワークショップパネル(指名)

  • マウス卵子形成過程におけるデスレセプターFasの関与

    赤澤 由起子, Moniruzzaman M, 李 智博, 酒巻 和弘, 宮野 隆

    第25回日本受精着床学会学術講演会, 2007年08月, 日本語, 仙台, 国内会議

    口頭発表(一般)

  • ブタおよびマウス着床前胚のタイトジャンクション関連タンパク質の発現性

    許 尚丹, 木村 隼人, 松村 宏和, 李 智博, 原山 洋, 三宅 正史

    第99回日本繁殖生物学会大会, 2006年09月, 日本語, 日本繁殖生物学会, 名古屋, 国内会議

    ポスター発表

  • 着床前胚におけるタイトジャンクション構造タンパク質Claudin-1の核移行性

    許 尚丹, 木村 隼人, 松村 宏和, 李 智博, 原山 洋, 三宅 正史

    第13回日本胚移植研究会, 2006年08月, 日本語, 日本胚移植研究会, 広島, 国内会議

    口頭発表(一般)

  • 哺乳類第一減数分裂後期の進行にはAPCcdc20が必要である

    李 智博, 宮野 隆, 三宅 正史

    第28回日本分子生物学会年会, 2005年12月, 日本語, 日本分子生物学会, 福岡, 国内会議

    口頭発表(一般)

  • マウス卵母細胞の成熟過程におけるCdc20 とCdh1の発現解析

    李 智博, 宮野 隆, 三宅 正史

    日本畜産学会第105 回大会, 2005年09月, 日本語, 日本畜産学会, 札幌, 国内会議

    ポスター発表

  • マウス卵母細胞の第一減数分裂におけるSecurin の過剰発現は染色体腕部のRec8 の消失を阻害する

    李 智博, 宮野 隆, 三宅 正史

    日本畜産学会第104 回大会, 2005年03月, 日本語, 日本畜産学会, 東京, 国内会議

    口頭発表(一般)

  • ウシ体外受精卵母細胞の核膜形成ならびにヒストンH3の脱リン酸化状態による前核形成異常の検出

    伊佐治 麻実子, 岩田 尚孝, 原山 洋, 李 智博, 三宅 正史

    第11回日本胚移植研究会大会, 2004年08月, 日本語, 日本胚移植研究会, 松山, 国内会議

    口頭発表(一般)

  • ブタ卵母細胞減数分裂過程における染色体分離に及ぼすproteasome阻害の影響

    李 智博, 大串 素雅子, 宮野 隆, 三宅 正史

    日本畜産学会第103回大会, 2004年03月, 日本語, 日本畜産学会, 東京, 国内会議

    口頭発表(一般)

所属学協会

  • 日本分子生物学会

  • 日本繁殖生物学会

  • 日本畜産学会

共同研究・競争的資金等の研究課題

  • 減数分裂の染色体の振る舞いを規定する2種類のコヒーシンの特性の解明

    李 智博

    日本学術振興会, 科学研究費助成事業 基盤研究(B), 基盤研究(B), 神戸大学, 2023年04月01日 - 2027年03月31日

  • 体細胞におけるコヒーシンの異所性発現による減数分裂模倣系の構築

    李 智博

    日本学術振興会, 科学研究費助成事業 挑戦的研究(萌芽), 挑戦的研究(萌芽), 神戸大学, 2022年06月 - 2024年03月, 研究代表者

  • 李 智博

    科学研究費補助金/基盤研究(B), 2018年04月 - 2022年03月, 研究代表者

    競争的資金

  • 李 智博

    科学研究費補助金/基盤研究(B), 2014年04月 - 2018年03月, 研究代表者

    競争的資金

  • 李 智博

    日本学術振興会, 科学研究費補助金/若手研究(A), 若手研究(B), 独立行政法人理化学研究所, 2011年04月 - 2015年03月, 研究代表者

    申請者は、マウス卵母細胞の減数分裂において、コンデンシンIとコンデンシンIIの時空間的な動態を明らかにした。また、二価染色体が形成されるときに、2つのコンデンシンが染色体の凝縮や分離、姉妹動原体の一方向性の確立に関与することを示した。それと平行して、減数分裂特異的な新規のコヒーシンサブユニットRAD21Lを発見した。RAD21Lは、第一減数分裂前期にだけ発現し、シナプトネマ複合体上に局在することから、相同染色体の対合や組換えに関与することが示唆された。

    競争的資金

  • 李 智博

    科学研究費補助金/特定領域研究, 2011年04月 - 2013年03月, 研究代表者

    競争的資金

  • 哺乳類減数分裂におけるコンデンシンの役割

    李 智博

    日本学術振興会, 科学研究費助成事業 若手研究(B), 若手研究(B), 独立行政法人理化学研究所, 2008年 - 2010年

    申請者は、マウス卵母細胞の減数分裂において、コンデンシンIとコンデンシンIIの時空間的な動態を明らかにした。また、二価染色体が形成されるときに、2つのコンデンシンが染色体の凝縮や分離、姉妹動原体の一方向性の確立に関与することを示した。それと平行して、減数分裂特異的な新規のコヒーシンサブユニットRAD21Lを発見した。RAD21Lは、第一減数分裂前期にだけ発現し、シナプトネマ複合体上に局在することから、相同染色体の対合や組換えに関与することが示唆された。

  • 宮野 隆

    科学研究費補助金/基盤研究(B), 2005年 - 2008年

    競争的資金

  • 李 智博

    科学研究費補助金/若手研究(B), 2004年 - 2006年, 研究代表者

    競争的資金

  • メダカおよびブタ卵母細胞における第一減数分裂特有の染色体チェックポイント機構

    李 智博

    日本学術振興会, 科学研究費助成事業 特別研究員奨励費, 特別研究員奨励費, 北海道大学, 2000年 - 2002年

    真核生物における姉妹染色分体の結合には、コヒーシンと呼ばれる蛋白質複合体が必須である。コヒーシンは、酵母では少なくともScc1(Rad21)、Scc、Smc1、Smc3の4種のサブユニットからなる。また、酵母の減数分裂においては、Scc1サブユニットがRec8に置き換わることが知られている。本研究では、脊椎動物の減数分裂におけるコヒーシンの発現を調べた。マウスRec8に対する抗体を作製し、その発現パターンを調べた。Rec8は、精巣および卵巣の生殖細胞にだけ発現しており、それ以外の組織や細胞では発現は見られなかった。生体内では、Rec8はリン酸化型のものと脱リン酸化型のものが存在し、他のコヒーシンサブユニットである減数分裂型のSMC1β、SMC3とは複合体を形成するが、体細胞型のSMC1αとは複合体を形成しないことが明らかとなった。減数分裂過程での、Rec8の詳細な細胞内局在を精母細胞で調べたところ、Rec8は、第一減数分裂前期では、SCP3と呼ばれるシナプトネマ構造蛋白質と同様の局在を示した。また、Rec8は、第一減数分裂中期では、染色体の軸に沿って、その腕部分とセントロメア部分に局在した。相同染色体の分離が起こる第一減数分裂の後期には、Rec8は染色体の腕部分から解離したが、姉妹染色分体が結合している部分(セントロメア部分)には存続した。このセントロメア部分に残ったRec8は、最終的に姉妹染色分体の分離が起こる第二減数分裂後期に、消失した。このRec8の染色体上の局在と段階的な解離は、Rec8が第一減数分裂における相同染色体の結合と第二減数分裂における姉妹染色分体の結合に関与し、Rec8の染色体の腕部分からの解離とセントロメア部分からの解離がそれぞれ相同染色体の分離と姉妹染色分体の分離を引き起こすことを強く示唆するものである。現在、この結果をまとめた論文を投稿中である。また、マウスの卵母細胞における同様の研究と、メダカにおけるコヒーシンの研究の論文を投稿する予定である。