Retrovirus vector silencing is de novo methylase independent and marked by a repressive histone code

Pannell, Dylan; Osborne, Cameron S.; Yao, Shuyuan; Sukonnik, Tanya; Pasceri, Peter; Karaiskakis, Angelo; Okano, Masaki; Li, En; Lipshitz, Howard D.; Ellis, James

doi:10.1093/emboj/19.21.5884

Cited by 149 publications

(137 citation statements)

References 49 publications

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“…It has also been suggested that retroviral silencing may act through methylation-independent mechanisms, and methylation is only a secondary step in this pathway. [16][17][18][19] Consistent with this, retroviral silencing was reported in mouse embryonic stem cells that are de novo methyltransferase (dnmt3a and dnmt3b) null. 16,20 Up until now, when a UIGD technique is used, the only way to confirm whether embryos or adult mice have received the transgene has been to analyze brains taken from killed animals.…”

Section: Discussionsupporting

confidence: 69%

“…[16][17][18][19] Consistent with this, retroviral silencing was reported in mouse embryonic stem cells that are de novo methyltransferase (dnmt3a and dnmt3b) null. 16,20 Up until now, when a UIGD technique is used, the only way to confirm whether embryos or adult mice have received the transgene has been to analyze brains taken from killed animals. In order to construct adult mouse models using the UIGD technique, therefore, gene-transferred mice should be discriminated against other littermates without killing.…”

Section: Discussionsupporting

confidence: 69%

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A retroviral vector suitable for ultrasound image-guided gene delivery to mouse brain

et al. 2011

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Gene transfer to the early-stage embryonic brain using the ultrasound image-guided gene delivery (UIGD) technique has proven to be valuable for investigating brain development. Thus far, this technology has been restricted to the study of embryonic neurogenesis. When this technique is designed to be employed for the study in adult animals, a long-term stable gene expression will be required. We attempted to develop a retroviral vector suitable for expressing exogenous genes in the brains of postnatal and adult mice in the context of the UIGD technique. Retroviral vectors containing four different long terminal repeats (LTRs) (each from Moloney murine leukemia virus (MoMLV), murine stem cell virus (MSCV), myeloproliferative sarcoma virus (MPSV) and spleen focus-forming virus (SFFV)) were compared using the well-known CE vector having the EF1a internal promoter as a control. The MS vector containing MSCV LTR produced a higher viral titer and a higher level of gene expression than other vectors including CE. The MS vector drove the gene expression in cultured neural stem cells for 3 weeks. Furthermore, the MS vector could efficiently deliver the gene to the mouse central nervous system, as transgene expression was found in various regions of the brains and spinal cords as well as in all major neural cell types. The data from an in vivo luciferase imaging analysis showed that the gene expression from the MS vector was sustainable for almost 3 months. Our data suggested that the MS vector would be suitable to construct mice containing the transgene expressed in the brain or spinal cord in a quick and cost-effective manner.

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Section: Discussionsupporting

confidence: 69%

Section: Discussionsupporting

confidence: 69%

A retroviral vector suitable for ultrasound image-guided gene delivery to mouse brain

et al. 2011

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“…2,5,6 Especially, acetylation status of lysine residues within histone tails are generally accepted as markers of chromatin activation. 27 For this purpose, we performed ChIP assay to detect histone acetylation levels of proviral ArsI and CMV promoter ( Figure 7).…”

Section: Effect Of Arsi On Histone Acetylationmentioning

confidence: 99%

“…The mechanism of transgene silencing remains to be elucidated; however, accumulating evidence suggest that epigenetic alterations of the transgene, such as DNA methylation and/or histone modifications, are likely components. 3,5,6 Engineering retroviral vectors to avoid epigenetic alterations is needed for successful application of gene therapy. Retroviral vectors currently in experimental and/or clinical practice are categorized into two groups: oncoretrovirus-based (eg Moloney-murine leukemia virus, MoMLV) and lentivirus-based (eg human immunodeficiency virus type-1, HIV-1) vectors.…”

Section: Introductionmentioning

confidence: 99%

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

et al. 2004

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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 b-globin insulator, incorporation of ArsI into lentiviral vector had little effect on the virus titer compared with chicken b-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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“…28,29 Another report described a methylase-independent mechanism of proviral silencing through MoMuLV and immunodeficiency virus type 1 (HIV-1) cis-acting sequences. 30 Facing this problem we decided to transduce the CD34 + progenitor cells with titers ranging from 100 to 150 MOI, which resulted in a high tyrosinase mRNA and concomitant EGFP expression in the first 3 days after transduction. Nevertheless, we revealed a strong silencing of transgenes in the cells during the expansion phase of the progenitor cells.…”

Section: Discussionmentioning

confidence: 99%

Efficient transduction and long-term retroviral expression of the melanoma-associated tumor antigen tyrosinase in CD34+ cord blood-derived dendritic cells

et al. 2002

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Retrovirus vector silencing is de novo methylase independent and marked by a repressive histone code

Cited by 149 publications

References 49 publications

A retroviral vector suitable for ultrasound image-guided gene delivery to mouse brain

A retroviral vector suitable for ultrasound image-guided gene delivery to mouse brain

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

Efficient transduction and long-term retroviral expression of the melanoma-associated tumor antigen tyrosinase in CD34+ cord blood-derived dendritic cells

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