Analysis of the infection dynamics and the host gene expression by the viral latency specific genes
| Project/Area Number |
16390136
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| Research Category |
Grant-in-Aid for Scientific Research (B)
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
Virology
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| Research Institution | Osaka University |
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Principal Investigator |
UEDA Keiji Osaka University, Graduate School of Medicine, Associate Professor, 医学系研究科, 助教授 (00221797)
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| Co-Investigator(Kenkyū-buntansha) |
YAMANISHI Koichi Osaka University, Graduate School of Medicine, Professor, 医学系研究科, 教授 (10029811)
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| Project Period (FY) |
2004 – 2005
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| Project Status |
Completed (Fiscal Year 2005)
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| Budget Amount *help |
¥15,100,000 (Direct Cost: ¥15,100,000)
Fiscal Year 2005: ¥4,700,000 (Direct Cost: ¥4,700,000)
Fiscal Year 2004: ¥10,400,000 (Direct Cost: ¥10,400,000)
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| Keywords | KSHV / latency / replication / maintenance / heterochromatin / Suv39H1 / PARP1 / LANA / RTA / RTA誘導因子 / PEL / 複製・分配・維持 / インスレーター / マトリックス / γヘルペスウイルス / カポジ肉腫関連ヘルペスウイルス / ヘテロクロマチン因子 / 末端反復配列 |
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| Research Abstract |
We analyzed why the viral gene expression was extremely limited to a few genes in the latency, and how the viral genome replicated and was maintained at the same copy number according to the host cell cycles.
We cloned the almost full length of terminal repeats (TR) of the Kaposi's sarcoma-associated herpesvirus (KSHV) in the bacmid, and tested whether the genome replicated and was maintained in the cells with or without viral latency specific gene products such as LANA (ORF73). We found that the bacmid replicated only in the presence of LANA and was maintained at the same copy number only under the selective pressure using G418, the resistant gene against which is encoded in the vector. This suggested that TR and LANA should be enough for the viral genome replication but not for the viral genome maintenance. Then, are there elements in the genome other than TR for the viral genome maintenance? We tried whether the full viral genome cloned in a bacmid was maintained or not. The results
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showed that there was no element responsible for the viral genome maintenance. The selective pressure, hygromycin selection in this case, was always required for the maintenance. Thus, there could be other reason why the KSHV genome was maintained at the same copy number in the infected cells, especially in the primary effusion lymphoma originated cell lines.
KSHV gene expression is quite limited in the latency. There are only four genes such as LANA, v-cyc, v-FLIP and kaposin expressing in the latency. These genes are clustered side by side. We found that LANA interacted with a cellular gene called Suv39H1, which is a histone H3 methyl transferase and recruits heterochromatin factor such as HP1α. Chromatin immunoprecipitation analysis (ChIP) showed that Suv39H1 and HP1α were located on the TR as well as LANA. Thus, the viral genome could be in the heterochromatin state by LANA and this could be one of the mechanisms why the viral gene expression was quite limited to the several genes.
TR and LANA seem to have an important role for the viral replication and genome maintenance. LANA binds with a specific sequence within TR and executes the viral replication and maintenance and regulates the viral gene expression in the latency. We focused on the TR and tried to identify TR binding proteins with an exhaustive approach by preparing a TR DNA column. We identified several proteins binding to the column, several of which were infected cell specific and the other were still TR DNA specific but not infected cell specific. One of them was poly (ADP-ribose) polymerase 1 (PARP1). We characterized this factor and found that this bound to the specific sequence within TR very near the LANA binding site. The direct interaction between PARP1 and LANA with the immunoprecipitaion analysis but LANA was proved to be ADP-ribosylated by PARP1. PARP1 activity is modulated by some drugs. Hydroxy Urea at the low concentration increases PARP1 activity and 3-aminobenzamide (3-ABA) decreases the activity. Using these drugs, it was shown the higher ribosylation of LANA resulted in the lower copy number of the viral genome and the lower ribosylation did in the higher copy number in the infected cells. Less
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Report
(3 results)
Research Products
(35 results)
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[Journal Article] Alteration in gene expression profile by fill-length hepatitis B virus genome.2005
Author(s)
Nakanishi, F., Ohkawa, K., Ishida, H., Hosui, A., Sato, A., Hiramatsu, N., Ueda, K., Takehara, T., Kasahara, A., Sasaki, Y., Hori, M., Hayashi, N.
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Journal Title
Inervirology 48(2-3)
Pages: 77-83
Description
「研究成果報告書概要(欧文)」より
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[Journal Article] Involvement of p38 signaling pathway in interferon-a-mediated antiviral activity toward hepatitis C virus.2004
Author(s)
Ishida, H., Ohkawa, K., Hosui, A., Hiramatsu, N., Kanto, T., Ueda, K., Takehara, T., Hayshi, N.
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Journal Title
Biochem.Biophys.Res.Comm. 321
Pages: 722-727
Description
「研究成果報告書概要(欧文)」より
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[Journal Article] PARP1 (poly[ADP-ribose] polymerase 1) binds with Kaposi's sarcoma-associated herpesvirus (KSHV) terminal repeat sequence and modulated KSHV replication in latency.2004
Author(s)
Ohsaki, E., Ueda, K., Sakakibara, S., Do, E., Yada, K., Yamanishi, K.
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Journal Title
J.Virol. 78(18)
Pages: 9936-9946
Description
「研究成果報告書概要(欧文)」より
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[Journal Article] Posttranscriptional regulator of Kaposi's sarcoma-associated herpesvirus interacts with RNA-binding protein PCBP1 and controls gene expression through the IRES.2004
Author(s)
Nishimura, K., Ueda, K., Guwanan, E., Sakakibara, S., Do, E., Ohsaki, E., Yada, K., Okuno, T., Yamanishi, K.
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Journal Title
Virology 325
Pages: 364-378
Description
「研究成果報告書概要(欧文)」より
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[Journal Article] Hepatitis C virus core functions as a suppressor of cyclin-dependent kinase-activating kinase and impairs cell cycle progression.2004
Author(s)
Ohkawa, K., Ishida, H., Nakanishi, F., Hosui, A., Ueda, K., Takahara, T., Hori, M., Hayashi, N.
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Journal Title
J.Biol.Chem. 279(12)
Pages: 11719-11726
Description
「研究成果報告書概要(欧文)」より
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[Journal Article] Accumulation of heterochromatin components on the terminal repeat sequence of Kaposi's sarcoma-associated herpesvirus mediated by the latency-associated nuclear antigen.2004
Author(s)
Sakakibara, S., Ueda, K., Nishimura, K., Ohsaki, E., Do, E., Yamanishi, K.
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Journal Title
J.Virol. 78(14)
Pages: 7299-7310
Description
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[Journal Article] KSHV RTA induces a transcriptional repressor, HEY1 that represses rta promoter.
Author(s)
Yada, K., Do, E., Sakakibara, S., Ohsaki, E., Ito, E., Watanabe, S., Ueda, K.
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Journal Title
Biochem, Biophys.Res.Comm. (In press)
Description
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