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Sea urchin insulator protects lentiviral vector from silencing by maintaining active chromatin structure

Gene Therapy volume 11, pages 819–828 (2004)Cite this article

Abstract

Suppressed expression of transgenes in vivo is the major obstacle in the gene therapy. For the long-term expression, we utilized a chromatin insulator from sea urchin arylsulfatase (Ars) gene locus (Ars insulator, ArsI), which has been shown to epigenetically regulate gene expression across species. ArsI was able to prevent silencing of the transgene in a myeloid cell line, HL-60, and a murine embryonic stem cell line, CCE, in an orientation-dependent manner, but not in Huh-7, K562 and MCF-7 cells, indicating that the effect of ArsI on gene silencing was cell type dependent. Although anti-silencing effect of ArsI was almost equivalent to that of chicken β-globin insulator, incorporation of ArsI into lentiviral vector had little effect on the virus titer compared with chicken β-globin insulator. Clonal analysis of transduced HL-60 cells revealed that ArsI protects the lentiviral vector from position effects regardless of its orientation. Furthermore, chromatin immunoprecipitation assays revealed that a high acetylation level was observed in the promoter of the insulated vector, whereas that of ArsI was independent of its anti-silencing capacity. In addition to it having little deteriorative effect on the virus titer, the identified anti-silencing effect of ArsI suggested its possibility for application in gene therapy.

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Acknowledgements

We would like to thank Dr Takumi Era for his technical advice in ES cell manipulation. We thank Dr Inder M Verma for supplying pHR′ vector. We also thank Dr Hiroyuki Miyoshi for supplying pCSII and pcDNA-VSV. We would like to express gratitude to Drs Koichi Ikuta and Sang-Kyu Ye for instructions with regard to ChIP assay. Finally, we would like to thank Jun-ichiro Yasunaga and Yuko Taniguchi for valuable suggestions and discussions. This work was supported by a grant from Bio-oriented Technology Research Advancement Institution, Japanese Ministry of Agriculture, Forestry and Fishery. SH is supported by the 21st Century COE Program of the Ministry of Education, Culture, Sports, Science amd Technology.

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Authors and Affiliations

  1. Laboratory of Virus Immunology, Institute for Virus Research, Kyoto University, Kyoto, Japan

    S Hino, J Fan, S Taguwa & M Matsuoka

  2. Graduate Department of Gene Science, Faculty of Science, Hiroshima University, Higashi-Hiroshima, Japan

    K Akasaka

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Hino, S., Fan, J., Taguwa, S. et al. Sea urchin insulator protects lentiviral vector from silencing by maintaining active chromatin structure. Gene Ther 11, 819–828 (2004). https://doi.org/10.1038/sj.gt.3302227

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