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CCCTC-binding factor recruitment to the early region of the human papillomavirus 18 genome regulates viral oncogene expression.

Paris C, Pentland I, Groves I, Roberts DC, Powis SJ, Coleman N, Roberts S, Parish JL - J. Virol. (2015)

Bottom Line: Loss of CTCF binding results in a reduction of a specific alternatively spliced transcript expressed from the early gene region concomitant with an increase in the abundance of unspliced early transcripts.In this study, we show that the essential host cell protein CTCF, which functions in genome-wide chromatin organization and gene regulation, is recruited to the HPV genome and plays an essential role in the regulation of early viral gene expression and transcript processing.These data highlight a novel virus-host interaction important for HPV pathogenicity.

View Article: PubMed Central - PubMed

Affiliation: University of St. Andrews, School of Medicine, St. Andrews, Fife, United Kingdom.

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Morphology and differentiation of HPV18 ΔCTCF organotypic raft cultures. (A, upper) Organotypic raft cultures of HFK, WT HPV18, and ΔCTCF HPV18 lines were fixed at day 14, and sections were stained with hematoxylin and eosin to assess morphology (upper). (Lower) Sections were stained by C-ISH to qualitatively assess viral genome amplification. Brown nuclear staining is present in cells with amplified HPV18 genomes. Scale bar, 10 μm. (B) The number of cells positive for C-ISH in wild-type and ΔCTCF HPV18 sections was counted in 10 fields of vision from sections of three independent raft cultures from each line (n = 30). The data are shown as means and standard errors. (C) Sections were stained with antibodies specific for keratin 5 (green; upper), keratin 1 (green; middle), or loricrin (red; lower). Sections were counterstained with Hoechst to highlight the nuclei (blue) and E1^E4 antibody to highlight productive areas of each section (red in upper and middle panels [rabbit antibody r424], green in the lower panel [mouse antibody 1D11]). Scale bar, 10 μm.
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Figure 6: Morphology and differentiation of HPV18 ΔCTCF organotypic raft cultures. (A, upper) Organotypic raft cultures of HFK, WT HPV18, and ΔCTCF HPV18 lines were fixed at day 14, and sections were stained with hematoxylin and eosin to assess morphology (upper). (Lower) Sections were stained by C-ISH to qualitatively assess viral genome amplification. Brown nuclear staining is present in cells with amplified HPV18 genomes. Scale bar, 10 μm. (B) The number of cells positive for C-ISH in wild-type and ΔCTCF HPV18 sections was counted in 10 fields of vision from sections of three independent raft cultures from each line (n = 30). The data are shown as means and standard errors. (C) Sections were stained with antibodies specific for keratin 5 (green; upper), keratin 1 (green; middle), or loricrin (red; lower). Sections were counterstained with Hoechst to highlight the nuclei (blue) and E1^E4 antibody to highlight productive areas of each section (red in upper and middle panels [rabbit antibody r424], green in the lower panel [mouse antibody 1D11]). Scale bar, 10 μm.

Mentions: To assess the biological function of CTCF recruitment to the HPV18 genome in differentiating epithelium, WT and ΔCTCF HPV18 HFK lines were grown in organotypic raft culture. Formaldehyde-fixed rafts were paraffin embedded and sectioned. Sections were stained with hematoxylin and eosin to assess morphology (Fig. 6A). As previously described, the WT HPV18 genome-containing rafts were increased in thickness, and mitotic cells were visible in the lower and upper suprabasal layers of the rafts compared to rafts derived from HFKs that did not contain HPV18 genomes (13). This phenotype was enhanced in ΔCTCF HPV18 rafts, which were consistently thicker, indicating increased cellular proliferation. Alongside these experiments, viral genome amplification was assessed by chromogenic in situ hybridization (C-ISH). No consistent differences were observed in the number of cells with amplified HPV genomes between WT and ΔCTCF HPV18 rafts, demonstrating that CTCF recruitment has a minimal role in viral genome amplification (Fig. 6A and B).


CCCTC-binding factor recruitment to the early region of the human papillomavirus 18 genome regulates viral oncogene expression.

Paris C, Pentland I, Groves I, Roberts DC, Powis SJ, Coleman N, Roberts S, Parish JL - J. Virol. (2015)

Morphology and differentiation of HPV18 ΔCTCF organotypic raft cultures. (A, upper) Organotypic raft cultures of HFK, WT HPV18, and ΔCTCF HPV18 lines were fixed at day 14, and sections were stained with hematoxylin and eosin to assess morphology (upper). (Lower) Sections were stained by C-ISH to qualitatively assess viral genome amplification. Brown nuclear staining is present in cells with amplified HPV18 genomes. Scale bar, 10 μm. (B) The number of cells positive for C-ISH in wild-type and ΔCTCF HPV18 sections was counted in 10 fields of vision from sections of three independent raft cultures from each line (n = 30). The data are shown as means and standard errors. (C) Sections were stained with antibodies specific for keratin 5 (green; upper), keratin 1 (green; middle), or loricrin (red; lower). Sections were counterstained with Hoechst to highlight the nuclei (blue) and E1^E4 antibody to highlight productive areas of each section (red in upper and middle panels [rabbit antibody r424], green in the lower panel [mouse antibody 1D11]). Scale bar, 10 μm.
© Copyright Policy - open-access
Related In: Results  -  Collection

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Figure 6: Morphology and differentiation of HPV18 ΔCTCF organotypic raft cultures. (A, upper) Organotypic raft cultures of HFK, WT HPV18, and ΔCTCF HPV18 lines were fixed at day 14, and sections were stained with hematoxylin and eosin to assess morphology (upper). (Lower) Sections were stained by C-ISH to qualitatively assess viral genome amplification. Brown nuclear staining is present in cells with amplified HPV18 genomes. Scale bar, 10 μm. (B) The number of cells positive for C-ISH in wild-type and ΔCTCF HPV18 sections was counted in 10 fields of vision from sections of three independent raft cultures from each line (n = 30). The data are shown as means and standard errors. (C) Sections were stained with antibodies specific for keratin 5 (green; upper), keratin 1 (green; middle), or loricrin (red; lower). Sections were counterstained with Hoechst to highlight the nuclei (blue) and E1^E4 antibody to highlight productive areas of each section (red in upper and middle panels [rabbit antibody r424], green in the lower panel [mouse antibody 1D11]). Scale bar, 10 μm.
Mentions: To assess the biological function of CTCF recruitment to the HPV18 genome in differentiating epithelium, WT and ΔCTCF HPV18 HFK lines were grown in organotypic raft culture. Formaldehyde-fixed rafts were paraffin embedded and sectioned. Sections were stained with hematoxylin and eosin to assess morphology (Fig. 6A). As previously described, the WT HPV18 genome-containing rafts were increased in thickness, and mitotic cells were visible in the lower and upper suprabasal layers of the rafts compared to rafts derived from HFKs that did not contain HPV18 genomes (13). This phenotype was enhanced in ΔCTCF HPV18 rafts, which were consistently thicker, indicating increased cellular proliferation. Alongside these experiments, viral genome amplification was assessed by chromogenic in situ hybridization (C-ISH). No consistent differences were observed in the number of cells with amplified HPV genomes between WT and ΔCTCF HPV18 rafts, demonstrating that CTCF recruitment has a minimal role in viral genome amplification (Fig. 6A and B).

Bottom Line: Loss of CTCF binding results in a reduction of a specific alternatively spliced transcript expressed from the early gene region concomitant with an increase in the abundance of unspliced early transcripts.In this study, we show that the essential host cell protein CTCF, which functions in genome-wide chromatin organization and gene regulation, is recruited to the HPV genome and plays an essential role in the regulation of early viral gene expression and transcript processing.These data highlight a novel virus-host interaction important for HPV pathogenicity.

View Article: PubMed Central - PubMed

Affiliation: University of St. Andrews, School of Medicine, St. Andrews, Fife, United Kingdom.

Show MeSH
Related in: MedlinePlus