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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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Association of CTCF with HPV genomes. Chromatin extracted from HPV16-positive W12 cells (A) and HPV18-positive HFKs (B) was immunoprecipitated with control antibody (rabbit IgG for W12 and FLAG M2 antibody for HPV18 HFKs) or CTCF-specific antibody. Coprecipitating DNA was analyzed by qPCR. The x axes indicate the position in the HPV genome amplified, and each data point represents the central point in each amplicon. A graphical representation of the HPV genome is shown above each data set, which has been linearized for ease of presentation (Enh, enhancer; Ep, early promoter). The CTCF binding sites verified by EMSA (Fig. 1 and Table 2) are indicated (dark gray ovals). Binding efficiency was normalized to negative-control antibody using the ΔΔCT method. The data represent the means and standard errors from three independent repeats.
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Figure 3: Association of CTCF with HPV genomes. Chromatin extracted from HPV16-positive W12 cells (A) and HPV18-positive HFKs (B) was immunoprecipitated with control antibody (rabbit IgG for W12 and FLAG M2 antibody for HPV18 HFKs) or CTCF-specific antibody. Coprecipitating DNA was analyzed by qPCR. The x axes indicate the position in the HPV genome amplified, and each data point represents the central point in each amplicon. A graphical representation of the HPV genome is shown above each data set, which has been linearized for ease of presentation (Enh, enhancer; Ep, early promoter). The CTCF binding sites verified by EMSA (Fig. 1 and Table 2) are indicated (dark gray ovals). Binding efficiency was normalized to negative-control antibody using the ΔΔCT method. The data represent the means and standard errors from three independent repeats.

Mentions: We next used HPV16 and HPV18 genome-containing cells to ascertain whether CTCF associates with the viral genome in cells. W12 cells, derived from a low-grade cervical squamous epithelial lesion, contain ∼100 episomal HPV16 genome copies/cell (39, 40), and HPV18-transfected HFKs contain ∼200 episomal HPV18 copies/cell (see Fig. 5B). CTCF association with the HPV genomes was determined by ChIP followed by qPCR. In both HPV16 and HPV18 genome-containing cells grown in monolayer, we noted a significant enrichment of CTCF binding within the E2 ORF, coinciding with the CTCF binding site conserved in high-risk HPV types but not in low-risk types (Fig. 3). In contrast, we failed to detect CTCF binding to the late gene region in either HPV16 or HPV18 genome-containing model systems.


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)

Association of CTCF with HPV genomes. Chromatin extracted from HPV16-positive W12 cells (A) and HPV18-positive HFKs (B) was immunoprecipitated with control antibody (rabbit IgG for W12 and FLAG M2 antibody for HPV18 HFKs) or CTCF-specific antibody. Coprecipitating DNA was analyzed by qPCR. The x axes indicate the position in the HPV genome amplified, and each data point represents the central point in each amplicon. A graphical representation of the HPV genome is shown above each data set, which has been linearized for ease of presentation (Enh, enhancer; Ep, early promoter). The CTCF binding sites verified by EMSA (Fig. 1 and Table 2) are indicated (dark gray ovals). Binding efficiency was normalized to negative-control antibody using the ΔΔCT method. The data represent the means and standard errors from three independent repeats.
© Copyright Policy - open-access
Related In: Results  -  Collection

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Figure 3: Association of CTCF with HPV genomes. Chromatin extracted from HPV16-positive W12 cells (A) and HPV18-positive HFKs (B) was immunoprecipitated with control antibody (rabbit IgG for W12 and FLAG M2 antibody for HPV18 HFKs) or CTCF-specific antibody. Coprecipitating DNA was analyzed by qPCR. The x axes indicate the position in the HPV genome amplified, and each data point represents the central point in each amplicon. A graphical representation of the HPV genome is shown above each data set, which has been linearized for ease of presentation (Enh, enhancer; Ep, early promoter). The CTCF binding sites verified by EMSA (Fig. 1 and Table 2) are indicated (dark gray ovals). Binding efficiency was normalized to negative-control antibody using the ΔΔCT method. The data represent the means and standard errors from three independent repeats.
Mentions: We next used HPV16 and HPV18 genome-containing cells to ascertain whether CTCF associates with the viral genome in cells. W12 cells, derived from a low-grade cervical squamous epithelial lesion, contain ∼100 episomal HPV16 genome copies/cell (39, 40), and HPV18-transfected HFKs contain ∼200 episomal HPV18 copies/cell (see Fig. 5B). CTCF association with the HPV genomes was determined by ChIP followed by qPCR. In both HPV16 and HPV18 genome-containing cells grown in monolayer, we noted a significant enrichment of CTCF binding within the E2 ORF, coinciding with the CTCF binding site conserved in high-risk HPV types but not in low-risk types (Fig. 3). In contrast, we failed to detect CTCF binding to the late gene region in either HPV16 or HPV18 genome-containing model systems.

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