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Differences in Transcriptional Activity of Human Papillomavirus Type 6 Molecular Variants in Recurrent Respiratory Papillomatosis.

Measso do Bonfim C, Simão Sobrinho J, Lacerda Nogueira R, Salgado Kupper D, Cardoso Pereira Valera F, Lacerda Nogueira M, Villa LL, Rahal P, Sichero L - PLoS ONE (2015)

Bottom Line: Furthermore, our results support a crucial role for ELF1 on transcriptional downregulation.We identified TFs implicated in the regulation of HPV-6 early gene expression.Many of these factors are mutated in cancer or are putative cancer biomarkers, and must be further studied.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Genomic Studies, Universidade do Estado de São Paulo, UNESP, São José do Rio Preto, SP, Brazil.

ABSTRACT
A significant proportion of recurrent respiratory papillomatosis (RRP) is caused by human papillomavirus type 6 (HPV-6). The long control region (LCR) contains cis-elements for regulation of transcription. Our aim was to characterize LCR HPV-6 variants in RRP cases, compare promoter activity of these isolates and search for cellular transcription factors (TFs) that could explain the differences observed. The complete LCR from 13 RRP was analyzed. Transcriptional activity of 5 variants was compared using luciferase assays. Differences in putative TFs binding sites among variants were revealed using the TRANSFAC database. Chromatin immunoprecipation (CHIP) and luciferase assays were used to evaluate TF binding and impact upon transcription, respectively. Juvenile-onset RRP cases harbored exclusively HPV-6vc related variants, whereas among adult-onset cases HPV-6a variants were more prevalent. The HPV-6vc reference was more transcriptionally active than the HPV-6a reference. Active FOXA1, ELF1 and GATA1 binding sites overlap variable nucleotide positions among isolates and influenced LCR activity. Furthermore, our results support a crucial role for ELF1 on transcriptional downregulation. We identified TFs implicated in the regulation of HPV-6 early gene expression. Many of these factors are mutated in cancer or are putative cancer biomarkers, and must be further studied.

No MeSH data available.


Related in: MedlinePlus

Binding and activity of ELF1, GATA1 and FOXA1 to HPV-6a-ref and HPV-6vc-ref.(A) Luciferase reporter assay for GATA1. Amplification of LCR fragments following chromatin immunoprecipitation (ChIP) using primers surrounding nucleotide position (B) 7631/33 and (C) 7762 and that differ between HPV-6a-ref and HPV-6vc-var1 variants. Input-nonimmunoprecipated samples.
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pone.0132325.g004: Binding and activity of ELF1, GATA1 and FOXA1 to HPV-6a-ref and HPV-6vc-ref.(A) Luciferase reporter assay for GATA1. Amplification of LCR fragments following chromatin immunoprecipitation (ChIP) using primers surrounding nucleotide position (B) 7631/33 and (C) 7762 and that differ between HPV-6a-ref and HPV-6vc-var1 variants. Input-nonimmunoprecipated samples.

Mentions: Nucleotide differences in positions 7631/33 leads to a substitution of a putative CEBP for an ELF1 binding site in HPV-6vc-ref as compared to HPV-6a-ref (Fig 1B). Additionally, alteration in nucleotide position 7762 determines the replacement of a putative HNF4a for a GATA1 binding site in HPV-6vc-ref as compared to HPV-6a-ref. ELF1 significantly repressed both HPV-6a-ref and HPV-6vc-ref (Figs 2A and 3A). However, ELF1 binding to HPV-6vc-ref LCR was not observed. In this case, ELF1 is most probably bound to the the ELF1 binding site at nucleotide position 7626 within the HPV-6a-ref LCR. Furthermore, we observed that GATA1 represses transcription of both HPV-6a-ref and HPV-6vc-ref (Figs 2A and 4A), although repression was more pronounced for the HPV-6vc-ref. GATA1 was shown to bind the LCR of both variants using ChIP (Fig 4B). We observed that ELF1, GATA1 and FOXA1 are detected not only in PHFK, but also in keratinocytes transducing E6/E7 from both HPV-16 and HPV-6b (S1 Fig).


Differences in Transcriptional Activity of Human Papillomavirus Type 6 Molecular Variants in Recurrent Respiratory Papillomatosis.

Measso do Bonfim C, Simão Sobrinho J, Lacerda Nogueira R, Salgado Kupper D, Cardoso Pereira Valera F, Lacerda Nogueira M, Villa LL, Rahal P, Sichero L - PLoS ONE (2015)

Binding and activity of ELF1, GATA1 and FOXA1 to HPV-6a-ref and HPV-6vc-ref.(A) Luciferase reporter assay for GATA1. Amplification of LCR fragments following chromatin immunoprecipitation (ChIP) using primers surrounding nucleotide position (B) 7631/33 and (C) 7762 and that differ between HPV-6a-ref and HPV-6vc-var1 variants. Input-nonimmunoprecipated samples.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4494706&req=5

pone.0132325.g004: Binding and activity of ELF1, GATA1 and FOXA1 to HPV-6a-ref and HPV-6vc-ref.(A) Luciferase reporter assay for GATA1. Amplification of LCR fragments following chromatin immunoprecipitation (ChIP) using primers surrounding nucleotide position (B) 7631/33 and (C) 7762 and that differ between HPV-6a-ref and HPV-6vc-var1 variants. Input-nonimmunoprecipated samples.
Mentions: Nucleotide differences in positions 7631/33 leads to a substitution of a putative CEBP for an ELF1 binding site in HPV-6vc-ref as compared to HPV-6a-ref (Fig 1B). Additionally, alteration in nucleotide position 7762 determines the replacement of a putative HNF4a for a GATA1 binding site in HPV-6vc-ref as compared to HPV-6a-ref. ELF1 significantly repressed both HPV-6a-ref and HPV-6vc-ref (Figs 2A and 3A). However, ELF1 binding to HPV-6vc-ref LCR was not observed. In this case, ELF1 is most probably bound to the the ELF1 binding site at nucleotide position 7626 within the HPV-6a-ref LCR. Furthermore, we observed that GATA1 represses transcription of both HPV-6a-ref and HPV-6vc-ref (Figs 2A and 4A), although repression was more pronounced for the HPV-6vc-ref. GATA1 was shown to bind the LCR of both variants using ChIP (Fig 4B). We observed that ELF1, GATA1 and FOXA1 are detected not only in PHFK, but also in keratinocytes transducing E6/E7 from both HPV-16 and HPV-6b (S1 Fig).

Bottom Line: Furthermore, our results support a crucial role for ELF1 on transcriptional downregulation.We identified TFs implicated in the regulation of HPV-6 early gene expression.Many of these factors are mutated in cancer or are putative cancer biomarkers, and must be further studied.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Genomic Studies, Universidade do Estado de São Paulo, UNESP, São José do Rio Preto, SP, Brazil.

ABSTRACT
A significant proportion of recurrent respiratory papillomatosis (RRP) is caused by human papillomavirus type 6 (HPV-6). The long control region (LCR) contains cis-elements for regulation of transcription. Our aim was to characterize LCR HPV-6 variants in RRP cases, compare promoter activity of these isolates and search for cellular transcription factors (TFs) that could explain the differences observed. The complete LCR from 13 RRP was analyzed. Transcriptional activity of 5 variants was compared using luciferase assays. Differences in putative TFs binding sites among variants were revealed using the TRANSFAC database. Chromatin immunoprecipation (CHIP) and luciferase assays were used to evaluate TF binding and impact upon transcription, respectively. Juvenile-onset RRP cases harbored exclusively HPV-6vc related variants, whereas among adult-onset cases HPV-6a variants were more prevalent. The HPV-6vc reference was more transcriptionally active than the HPV-6a reference. Active FOXA1, ELF1 and GATA1 binding sites overlap variable nucleotide positions among isolates and influenced LCR activity. Furthermore, our results support a crucial role for ELF1 on transcriptional downregulation. We identified TFs implicated in the regulation of HPV-6 early gene expression. Many of these factors are mutated in cancer or are putative cancer biomarkers, and must be further studied.

No MeSH data available.


Related in: MedlinePlus