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ABSTRACT: It has been shown that constitutively active Wnt5a-Ror2 signaling in osteosarcoma cell lines plays crucial roles in induced expression of matrix metalloproteinase-13 (MMP-13), required for their invasiveness; however, it remains largely unclear about the molecular basis of MMP-13 gene induction by Wnt5a-Ror2 signaling. Here we show by reporter assay that the activator protein 1 (AP1) (binding site in the promoter region of MMP-13 gene is primarily responsible for its transcriptional activation by Wnt5a-Ror2 signaling in osteosarcoma cell lines SaOS-2 and U2OS. Chromatin immunoprecipitation assays revealed that c-Jun and ATF2 are crucial transcription factors recruited to the AP1-binding site in the MMP-13 gene promoter during Wnt5a-Ror2 signaling in SaOS-2 cells. Using siRNA-mediated suppression or specific inhibitors, we also show that Dishevelled2 (Dvl2) and c-Jun N-terminal kinase are required for MMP-13 gene induction presumably via phosphorylation of c-Jun and ATF2 during Wnt5a-Ror2 signaling in SaOS-2 cells. Interestingly, Dvl2 and Rac1, but not Dvl3, are required for MMP-13 expression in SaOS-2 cells, whereas Dvl3, but not Dvl2 and Rac1, is required for its expression in U2OS cells, indicating the presence of distinct intracellular signaling machineries leading to expression of the same gene, in this case MMP-13 gene in different osteosarcoma cell lines. Moreover, we provide evidence suggesting that Wnt5a-Ror2 signaling might also be required for expression of MMP-13 gene during the development of the cartilaginous tissue.
Journal of Biological Chemistry 11/2011; 287(2):1588-99. · 4.77 Impact Factor
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ABSTRACT: Correct spatio-temporal regulation of Wnt5a signaling is required for normal developmental morphogenesis, and defects in this pathway are linked to tumorigenesis. The precise role of Wnt5a signaling in cancer has, however, been a matter of controversy. Loss of Wnt5a signaling is related to development of lymphoid malignancies, whereas constitutively active Wnt5a signaling is involved in invasion or metastasis of several cancers. Interestingly, recent studies in Drosophila and mouse have revealed that disrupted cell polarity might contribute to invasion/metastasis of cancers. Wnt5a activates the planar cell polarity (PCP) pathway, partly through the receptor tyrosine kinase Ror2. Here, we review developments in our understanding of the molecular mechanisms underlying Wnt5a signaling, with an emphasis on the role of Ror2 in cancer. We also propose a model where the outcomes of normal and aberrant Wnt5a/Ror2 signaling depend on cell/tissue-tropic contexts.
Trends in cell biology 03/2010; 20(6):346-54. · 12.12 Impact Factor
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ABSTRACT: The maturation of primary microRNAs (pri-miRNAs) to precursor miRNAs (pre-miRNAs) is mediated by the "microprocessor" complex minimally comprimising two core components, Drosha and DGCR8. However, the roles of RNA-binding proteins associated with these core units in the large Drosha complex remain to be defined. While signal-dependent regulation of miRNA biogenesis is assumed, such regulation remains to be described. Here, we provide a short review based on our recent findings of hormonally-regulated pri-miRNA processing by nuclear estrogen receptor.
Advances in experimental medicine and biology 01/2010; 700:43-55. · 1.09 Impact Factor
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ABSTRACT: Steroid hormones and their cognate nuclear receptors exert a wide spectrum of biological actions through regulation of transcriptional and posttranscriptional processes. However, the underlying molecular mechanism by which steroid hormones control posttranscriptional processes is largely unknown. We now report that estrogen receptor alpha (ERalpha) inhibits the maturation of a particular microRNA (miRNA) and thereby stabilizes the mRNA of an ERalpha target gene through the 3'UTR. Estrogen-bound ERalpha downregulated expression of a set of miRNAs in both animals and cultured cells. Activated ERalpha attenuated the processing of primary miRNAs into pre-miRNAs through estrogen-dependent association with the Drosha complex, resulting in stabilization of the transcript of an ERalpha target gene through its 3'UTR. Thus, a steroid hormone achieves posttranscriptional control by regulating the maturation of miRNA.
Molecular cell 10/2009; 36(2):340-7. · 14.61 Impact Factor
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ABSTRACT: MicroRNAs (miRNAs) have emerged as key post-transcriptional regulators of gene expression, involved in diverse physiological and pathological processes. Although miRNAs can function as both tumour suppressors and oncogenes in tumour development, a widespread downregulation of miRNAs is commonly observed in human cancers and promotes cellular transformation and tumorigenesis. This indicates an inherent significance of small RNAs in tumour suppression. However, the connection between tumour suppressor networks and miRNA biogenesis machineries has not been investigated in depth. Here we show that a central tumour suppressor, p53, enhances the post-transcriptional maturation of several miRNAs with growth-suppressive function, including miR-16-1, miR-143 and miR-145, in response to DNA damage. In HCT116 cells and human diploid fibroblasts, p53 interacts with the Drosha processing complex through the association with DEAD-box RNA helicase p68 (also known as DDX5) and facilitates the processing of primary miRNAs to precursor miRNAs. We also found that transcriptionally inactive p53 mutants interfere with a functional assembly between Drosha complex and p68, leading to attenuation of miRNA processing activity. These findings suggest that transcription-independent modulation of miRNA biogenesis is intrinsically embedded in a tumour suppressive program governed by p53. Our study reveals a previously unrecognized function of p53 in miRNA processing, which may underlie key aspects of cancer biology.
Nature 08/2009; 460(7254):529-33. · 36.28 Impact Factor
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Eriko Suzuki,
Yue Zhao,
Saya Ito,
Shun Sawatsubashi,
Takuya Murata,
Takashi Furutani,
Yuko Shirode, Kaoru Yamagata,
Masahiko Tanabe,
Shuhei Kimura,
Takashi Ueda,
Sally Fujiyama,
Jinseon Lim,
Hiroyuki Matsukawa,
Alexander P Kouzmenko,
Toshiro Aigaki,
Tetsuya Tabata,
Ken-ichi Takeyama,
Shigeaki Kato
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ABSTRACT: Spinal and bulbar muscular atrophy (SBMA) is a neurodegenerative disorder caused by a polyglutamine repeat (polyQ) expansion within the human androgen receptor (AR). Unlike other neurodegenerative diseases caused by abnormal polyQ expansion, the onset of SBMA depends on androgen binding to mutant human polyQ-AR proteins. This is also observed in Drosophila eyes ectopically expressing the polyQ-AR mutants. We have genetically screened mediators of androgen-induced neurodegeneration caused by polyQ-AR mutants in Drosophila eyes. We identified Rbf (Retinoblastoma-family protein), the Drosophila homologue of human Rb (Retinoblastoma protein), as a neuroprotective factor. Androgen-dependent association of Rbf or Rb with AR was remarkably potentiated by aberrant polyQ expansion. Such potentiated Rb association appeared to attenuate recruitment of histone deacetyltransferase 1 (HDAC1), a corepressor of E2F function. Either overexpression of Rbf or E2F deficiency in fly eyes reduced the neurotoxicity of the polyQ-AR mutants. Induction of E2F function by polyQ-AR-bound androgen was suppressed by Rb in human neuroblastoma cells. We conclude that abnormal expansion of polyQ may potentiate innate androgen-dependent association of AR with Rb. This appears to lead to androgen-dependent onset of SBMA through aberrant E2F transactivation caused by suppressed histone deacetylation.
Proceedings of the National Academy of Sciences 03/2009; 106(10):3818-22. · 9.68 Impact Factor
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Yue Zhao,
Ken-ichi Takeyama,
Shun Sawatsubashi,
Saya Ito,
Eriko Suzuki, Kaoru Yamagata,
Masahiko Tanabe,
Shuhei Kimura,
Sally Fujiyama,
Takashi Ueda,
Takuya Murata,
Hiroyuki Matsukawa,
Yuko Shirode,
Alexander P Kouzmenko,
Feng Li,
Testuya Tabata,
Shigeaki Kato
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ABSTRACT: Ligand-bound nuclear receptors (NR) activate transcription of the target genes. This activation is coupled with histone modifications and chromatin remodeling through the function of various coregulators. However, the nature of the dependence of a NR coregulator action on the presence of the chromatin environment at the target genes is unclear. To address this issue, we have developed a modified position effect variegation experimental model system that includes an androgen-dependent reporter transgene inserted into either a pericentric heterochromatin region or a euchromatic region of Drosophila chromosome. Human androgen receptor (AR) and its constitutively active truncation mutant (AR AF-1) were transcriptionally functional in both chromosomal regions. Predictably, the level of AR-induced transactivation was lower in the pericentric heterochromatin. In genetic screening for AR AF-1 coregulators, Drosophila CREB binding protein (dCBP) was found to corepress AR transactivation at the pericentric region whereas it led to coactivation in the euchromatic area. Mutations of Sir2 acetylation sites or deletion of the CBP acetyltransferase domain abrogated dCBP corepressive action for AR at heterochromatic areas in vivo. Such a CBP corepressor function for AR was observed in the transcriptionally silent promoter of an AR target gene in cultured mammalian cells. Thus, our findings suggest that the action of NR coregulators may depend on the state of chromatin at the target loci.
Molecular and cellular biology 01/2009; 29(4):1017-34. · 6.06 Impact Factor
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Masahiko Tanabe,
Alexander P Kouzmenko,
Saya Ito,
Shun Sawatsubashi,
Eriko Suzuki,
Sally Fujiyama, Kaoru Yamagata,
Yue Zhao,
Shuhei Kimura,
Takashi Ueda,
Takuya Murata,
Hiroyuki Matsukawa,
Ken-ichi Takeyama,
Shigeaki Kato
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ABSTRACT: H2A.Z is an evolutionarily highly conserved non-allelic variant of histone H2A. H2A.Z and its homologues have been shown to involve in both chromatin silencing and activation. Although much of our knowledge of H2A.Z biological activity has come from studies on its yeast homologue Htz1, H2A.Z appears to have more complex and diverse functions in higher eukaryotes. To investigate the involvement of H2AvD, a Drosophila homologue of mammalian H2A.Z, in mechanisms of conditional activation of facultatively silenced genes, we generated transgenic Drosophila lines expressing H2AvD fused at the C- or N-terminus with the green fluorescent protein (GFP). Using heat shock-induced gene activation and polytene chromosome puff formation as an in vivo model system, we analyzed effects of H2AvD termini modifications on transcription. We found that N-terminally fused GFP inhibited H2AvD acetylation and impaired heat shock-induced puff formation and hsp70 gene activation. Our data suggest that the N-terminal region of H2AvD plays a pivotal role in transcriptional activation and that induction of transiently silenced Drosophila loci associates with increased acetylation of H2AvD.
Genes to Cells 12/2008; 13(12):1279-88. · 2.68 Impact Factor
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Takuya Murata,
Eriko Suzuki,
Saya Ito,
Shun Sawatsubashi,
Yue Zhao, Kaoru Yamagata,
Masahiko Tanabe,
Sally Fujiyama,
Shuhei Kimura,
Takashi Ueda,
Hiroyuki Matsukawa,
Alexander Kouzmenko,
Takashi Furutani,
Erina Kuranaga,
Masayuki Miura,
Ken-ichi Takeyama,
Shigeaki Kato
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ABSTRACT: Abnormal polyglutamine (polyQ) expansion in the N-terminal domain of the human androgen receptor (hAR) is known to cause spinobulbar muscular atrophy (SBMA), a hereditary human neurodegenerative disorder. To explore the molecular mechanisms of neurodegeneration in SBMA, we genetically screened modulators of neurodegeneration in a Drosophila SBMA experimental model system. We identified hoip as an accelerator of polyQ-induced neurodegeneration. We found that hoip forms a complex with 18s rRNA together nop56 and nop5 proteins, whose human homologs are known to form a snoRNP complex involved in ribosomal RNA processing. Significantly, the levels of mutant polyQ-hAR were up-regulated in a mutant line overexpressing hoip. Consistently, severe neurodegeneration phenotype (rough eye) was also observed in both nop56 and nop5 overexpression mutant lines. These findings suggest that the process of neurodegeneration induced by abnormal polyQ expansion in the hAR may be regulated by the activity of snoRNP complex.
Bioscience Biotechnology and Biochemistry 10/2008; 72(9):2255-61. · 1.28 Impact Factor
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Shuhei Kimura,
Shun Sawatsubashi,
Saya Ito,
Alexander Kouzmenko,
Eriko Suzuki,
Yue Zhao, Kaoru Yamagata,
Masahiko Tanabe,
Takashi Ueda,
Sari Fujiyama,
Takuya Murata,
Hiroyuki Matsukawa,
Ken-Ichi Takeyama,
Nobuo Yaegashi,
Shigeaki Kato
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ABSTRACT: Histone arginine methylation is an epigenetic marker that regulates gene expression by defining the chromatin state. Arginine methyltransferases, therefore, serve as transcriptional co-regulators. However, unlike other transcriptional co-regulators, the physiological roles of arginine methyltransferases are poorly understood. Drosophila arginine methyltransferase 1 (DART1), the mammalian PRMT1 homologue, methylates the arginine residue of histone H4 (H4R3me2). Disruption of DART1 in Drosophila by imprecise P-element excision resulted in low viability during metamorphosis in the pupal stages. In the pupal stage, an ecdysone hormone signal is critical for developmental progression. DART1 interacted with the nuclear ecdysone receptor (EcR) in a ligand-dependent manner, and co-repressed EcR in intact flies. These findings suggest that DART1, a histone arginine methyltransferase, is a co-repressor of EcR that is indispensable for normal pupal development in the intact fly.
Biochemical and Biophysical Research Communications 08/2008; 371(4):889-93. · 2.48 Impact Factor
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Toru Fukuda, Kaoru Yamagata,
Sally Fujiyama,
Takahiro Matsumoto,
Iori Koshida,
Kimihiro Yoshimura,
Masatomo Mihara,
Masanori Naitou,
Hideki Endoh,
Takashi Nakamura,
Chihiro Akimoto,
Yoko Yamamoto,
Takenobu Katagiri,
Charles Foulds,
Shinichiro Takezawa,
Hirochika Kitagawa,
Ken-ichi Takeyama,
Bert W O'Malley,
Shigeaki Kato
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ABSTRACT: MicroRNAs (miRNAs) control cell proliferation, differentiation and fate through modulation of gene expression by partially base-pairing with target mRNA sequences. Drosha is an RNase III enzyme that is the catalytic subunit of a large complex that cleaves pri-miRNAs with distinct structures into pre-miRNAs. Here, we show that both the p68 and p72 DEAD-box RNA helicase subunits in the mouse Drosha complex are indispensable for survival in mice, and both are required for primary miRNA and rRNA processing. Gene disruption of either p68 or p72 in mice resulted in early lethality, and in both p68(-/-) and p72(-/-) embryos, expression levels of a set of, but not all, miRNAs and 5.8S rRNA were significantly lowered. In p72(-/-) MEF cells, expression of p72, but not a mutant lacking ATPase activity, restored the impaired expression of miRNAs and 5.8S rRNA. Furthermore, we purified the large complex of mouse Drosha and showed it could generate pre-miRNA and 5.8S rRNA in vitro. Thus, we suggest that DEAD-box RNA helicase subunits are required for recognition of a subset of primary miRNAs in mDrosha-mediated processing.
Nature Cell Biology 06/2007; 9(5):604-11. · 19.49 Impact Factor
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Toru Fukuda, Kaoru Yamagata,
Sally Fujiyama,
Takahiro Matsumoto,
Iori Koshida,
Kimihiro Yoshimura,
Masatomo Mihara,
Masanori Naitou,
Hideki Endoh,
Takashi Nakamura,
Chihiro Akimoto,
Yoko Yamamoto,
Takenobu Katagiri,
Charles Foulds,
Shinichiro Takezawa,
Hirochika Kitagawa,
Ken-ichi Takeyama,
Bert W. O'Malley,
Shigeaki Kato
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ABSTRACT: MicroRNAs (miRNAs) control cell proliferation, differentiation and fate through modulation of gene expression by partially base-pairing with target mRNA sequences
Nature Cell Biology 04/2007; 9(5):604-611. · 19.49 Impact Factor
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ABSTRACT: While the ecdysone dependency of puff formation in giant polytene chromosomes from fly salivary glands has been well documented, the molecular mechanisms underlying this process remain unknown. However, it does appear to involve chromatin remodeling and modification mediated by ecdysone receptor (EcR). As Drosophila poly(ADP-ribose) polymerase (dPARP) has recently been reported to be involved in ecdysone-induced puff formation, we decided to test the possible role of dPARP in ligand-induced dEcR transactivation in an insect system. dPARP co-activated the ligand-induced transactivation function of EcR in the insect cell line S2, and appeared to physically interact with EcR in a ligand-dependent manner. ChIP analysis of an EcR target gene promoter revealed ligand-dependent recruitment of dPARP with poly(ADP-ribosyl)ation of histones in the EcR binding site and, surprisingly, also in a distal region of the promoter. Our results indicated that EcR-mediated gene regulation may be coupled with chromatin modification through poly(ADP-ribosyl)ation.
Biochemical and Biophysical Research Communications 08/2004; 320(1):268-72. · 2.48 Impact Factor
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ABSTRACT: Insect development is controlled by the combined actions of ecdysteroid and juvenile hormones. Transcriptional control by ecdysteroid hormones is mediated via two nuclear receptor superfamily members, ecdysone receptor (EcR) and its heterodimeric partner, ultraspiracle (USP). Although the ecdysteroid hormone 20-hydroxyecdysone acts as an EcR ligand and activates transcription through EcR/USP heterodimers, the activity of juvenile hormones, such as Juvenile hormone III (JH III), and methoprenic acid (MA) via USP remains unclear. Here, we demonstrate that juvenile hormones act as USP ligands and exhibit suppressive effects on ecdysone-dependent EcR transactivation. JH III- and MA-bound USP markedly repressed ecdysone-dependent EcR transactivation through shifting of the USP ligand-binding domain alpha-helix 12 without affecting EcR/USP heterodimerization or DNA binding. Moreover, transcriptional repression by USP ligands was attenuated by a histone deacetylation inhibitor. Our results suggested that juvenile hormones serve as USP ligands that antagonize EcR-mediated ecdysone actions through the recruitment of histone deacetylase complexes.
Biochemical and Biophysical Research Communications 08/2004; 320(1):262-7. · 2.48 Impact Factor
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Ken-Ichi Takeyama,
Saya Ito,
Shun Sawatsubashi,
Yuko Shirode,
Ayako Yamamoto,
Eriko Suzuki,
Akio Maki, Kaoru Yamagata,
Yue Zhao,
Alexandre Kouzmenko,
Tetsuya Tabata,
Shigeaki Kato
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ABSTRACT: Androgen receptor (hAR) regulates transcription of target genes in a ligand-dependent manner and recruits a number of co-activators for the ligand-induced transactivation via the N-terminal, activation function-1 (AF-1), and C-terminal, AF-2, transactivation domains. But the co-regulator functions on each of AR domains have not yet been fully understood. We have established a Drosophila transgenic system in which hAR and its deletion mutants are ectopically expressed in fly tissues together with an AR response element (ARE)-GFP reporter gene, and have confirmed that hAR was functional in ARE transactivation without affecting the expression of endogenous genes. We found that transcriptional activity of the hAR AF-1 domain was markedly reduced in Drosophila deficiency mutants of homologs for known mammalian co-activators of the AR ligand-dependent AF-2 domain. This suggests that hAR AF-1 recruits co-activators previously known only to interact with the AF-2 domain. Therefore, Drosophila with the hAR AF-1 transgene provides a relevant genetic system in which to uncover novel functions of vertebrate steroid hormone receptors and to screen for novel AF-1 co-regulators.
Bioscience Biotechnology and Biochemistry 07/2004; 68(6):1209-15. · 1.28 Impact Factor
