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Characterization and whole genome analysis of human papillomavirus type 16 e1-1374^63nt variants.

Sabol I, Matovina M, Si-Mohamed A, Grce M - PLoS ONE (2012)

Bottom Line: On the other hand, changes within the E1 region were the major differences from the A2 sub-lineage, which has been historically but inconclusively associated with high grade cervical disease.However, this study shows that the variations within the E1 region could possibly affect cervical disease, since the E1-1374^63nt E-G350 variant is significantly associated with lower grade cervical lesions, in comparison to the A1 and A2 sub-lineage variants.Furthermore, it appears that the silent variation 109T>C of the E-C109/G350 variant might have a significant role in the viral life cycle and warrants further study.

View Article: PubMed Central - PubMed

Affiliation: Division of Molecular Medicine, Rudjer Boskovic Institute, Zagreb, Croatia.

ABSTRACT

Background: The variation of the most common Human papillomavirus (HPV) type found in cervical cancer, the HPV16, has been extensively investigated in almost all viral genes. The E1 gene variation, however, has been rarely studied. The main objective of the present investigation was to analyze the variability of the E6 and E1 genes, focusing on the recently identified E1-1374^63nt variant.

Methodology/principal findings: Variation within the E6 of 786 HPV16 positive cervical samples was analyzed using high-resolution melting, while the E1-1374^63nt duplication was assayed by PCR. Both techniques were supplemented with sequencing. The E1-1374^63nt duplication was linked with the E-G350 and the E-C109/G350 variants. In comparison to the referent HPV16, the E1-1374^63nt E-G350 variant was significantly associated with lower grade cervical lesions (p = 0.029), while the E1-1374^63nt E-C109/G350 variant was equally distributed between high and low grade lesions. The E1-1374^63nt variants were phylogenetically closest to E-G350 variant lineage (A2 sub-lineage based on full genome classification). The major differences between E1-1374^63nt variants were within the LCR and the E6 region. On the other hand, changes within the E1 region were the major differences from the A2 sub-lineage, which has been historically but inconclusively associated with high grade cervical disease. Thus, the shared variations cannot explain the particular association of the E1-1374^63nt variant with lower grade cervical lesions.

Conclusions/significance: The E1 region has been thus far considered to be well conserved among all HPVs and therefore uninteresting for variability studies. However, this study shows that the variations within the E1 region could possibly affect cervical disease, since the E1-1374^63nt E-G350 variant is significantly associated with lower grade cervical lesions, in comparison to the A1 and A2 sub-lineage variants. Furthermore, it appears that the silent variation 109T>C of the E-C109/G350 variant might have a significant role in the viral life cycle and warrants further study.

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Prediction of E1 protein structure.Panel A shows 3D structure comparison between superposed monomer of the previously solved BPV1 E1 helicase domain hexamer structure (cyan; PDB ID:2GXA) [25], HPV18 E1 helicase domain structure (yellow; PDB ID:1TUE) [26], BPV1 E1 DNA binding domain structure (red; PDB ID:1F08) [27] and the predicted referent E1 model (white). The sequence at the position where the duplication occurs within the E1-1374∧63nt variants is highlighted in green, as is the amino acid 63 that is also changed in the E1-1374∧63nt variants. Panel B depicts superposed structures of the referent (white) and E1-1374∧63nt variant (magenta) E1 models. As before the referent sequence is highlighted in green and the changes specific to E1-1374∧63nt variant are highlighted in red. It can be seen that the newly added 21 amino acids (highlighted in red) partially overlap the potentially phosphorylated threonines (highlighted in green) at positions 153 and 155 and possibly influence the phosphorylation regulation of the E1 function. Panels C and D show the hexamer structure derived from the predicted E1 reference or E1-1374∧63nt variant proteins, respectively. The individual monomers are colored differently. The original referent sequence is colored in green within the white referent E1 monomer and the E1 variant magenta monomer. The sequence duplication in the E1-1374∧63nt variants is located at the junction between 2 adjacent E1 monomers within the structure and is highlighted in red on panel D. Positions of potentially phosphorylated threonines and amino acid substitution at position 63 are only highlighted on the white and magenta monomers.
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pone-0041045-g002: Prediction of E1 protein structure.Panel A shows 3D structure comparison between superposed monomer of the previously solved BPV1 E1 helicase domain hexamer structure (cyan; PDB ID:2GXA) [25], HPV18 E1 helicase domain structure (yellow; PDB ID:1TUE) [26], BPV1 E1 DNA binding domain structure (red; PDB ID:1F08) [27] and the predicted referent E1 model (white). The sequence at the position where the duplication occurs within the E1-1374∧63nt variants is highlighted in green, as is the amino acid 63 that is also changed in the E1-1374∧63nt variants. Panel B depicts superposed structures of the referent (white) and E1-1374∧63nt variant (magenta) E1 models. As before the referent sequence is highlighted in green and the changes specific to E1-1374∧63nt variant are highlighted in red. It can be seen that the newly added 21 amino acids (highlighted in red) partially overlap the potentially phosphorylated threonines (highlighted in green) at positions 153 and 155 and possibly influence the phosphorylation regulation of the E1 function. Panels C and D show the hexamer structure derived from the predicted E1 reference or E1-1374∧63nt variant proteins, respectively. The individual monomers are colored differently. The original referent sequence is colored in green within the white referent E1 monomer and the E1 variant magenta monomer. The sequence duplication in the E1-1374∧63nt variants is located at the junction between 2 adjacent E1 monomers within the structure and is highlighted in red on panel D. Positions of potentially phosphorylated threonines and amino acid substitution at position 63 are only highlighted on the white and magenta monomers.

Mentions: Structural prediction of the variant and the referent E1 proteins using the SAM-T08 [21], I-TASSER [22] and Phyre2 [23] services resulted in several models. However, only the E1 Phyre2 model of the E1-1374∧63nt variant could be reconciled with the solved structure of the E1 hexamer (PDB ID:2GXA) [25], without any overlaps between individual E1 monomer structures. The E1 reference protein model was refined by the I-TASSER server using the Phyre2 E1-1374∧63nt variant structure as a constraint. Figure 2 presents the predicted full length structural models of the reference and the E1-1374∧63nt variant proteins. From the structural prediction, it appears that both E1-C1053 and E1-1374∧63nt are positioned on the surface of the E1 protein. Additional three dimensional representations of the predicted models are presented in Supplement material (Figure S1).


Characterization and whole genome analysis of human papillomavirus type 16 e1-1374^63nt variants.

Sabol I, Matovina M, Si-Mohamed A, Grce M - PLoS ONE (2012)

Prediction of E1 protein structure.Panel A shows 3D structure comparison between superposed monomer of the previously solved BPV1 E1 helicase domain hexamer structure (cyan; PDB ID:2GXA) [25], HPV18 E1 helicase domain structure (yellow; PDB ID:1TUE) [26], BPV1 E1 DNA binding domain structure (red; PDB ID:1F08) [27] and the predicted referent E1 model (white). The sequence at the position where the duplication occurs within the E1-1374∧63nt variants is highlighted in green, as is the amino acid 63 that is also changed in the E1-1374∧63nt variants. Panel B depicts superposed structures of the referent (white) and E1-1374∧63nt variant (magenta) E1 models. As before the referent sequence is highlighted in green and the changes specific to E1-1374∧63nt variant are highlighted in red. It can be seen that the newly added 21 amino acids (highlighted in red) partially overlap the potentially phosphorylated threonines (highlighted in green) at positions 153 and 155 and possibly influence the phosphorylation regulation of the E1 function. Panels C and D show the hexamer structure derived from the predicted E1 reference or E1-1374∧63nt variant proteins, respectively. The individual monomers are colored differently. The original referent sequence is colored in green within the white referent E1 monomer and the E1 variant magenta monomer. The sequence duplication in the E1-1374∧63nt variants is located at the junction between 2 adjacent E1 monomers within the structure and is highlighted in red on panel D. Positions of potentially phosphorylated threonines and amino acid substitution at position 63 are only highlighted on the white and magenta monomers.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC3404080&req=5

pone-0041045-g002: Prediction of E1 protein structure.Panel A shows 3D structure comparison between superposed monomer of the previously solved BPV1 E1 helicase domain hexamer structure (cyan; PDB ID:2GXA) [25], HPV18 E1 helicase domain structure (yellow; PDB ID:1TUE) [26], BPV1 E1 DNA binding domain structure (red; PDB ID:1F08) [27] and the predicted referent E1 model (white). The sequence at the position where the duplication occurs within the E1-1374∧63nt variants is highlighted in green, as is the amino acid 63 that is also changed in the E1-1374∧63nt variants. Panel B depicts superposed structures of the referent (white) and E1-1374∧63nt variant (magenta) E1 models. As before the referent sequence is highlighted in green and the changes specific to E1-1374∧63nt variant are highlighted in red. It can be seen that the newly added 21 amino acids (highlighted in red) partially overlap the potentially phosphorylated threonines (highlighted in green) at positions 153 and 155 and possibly influence the phosphorylation regulation of the E1 function. Panels C and D show the hexamer structure derived from the predicted E1 reference or E1-1374∧63nt variant proteins, respectively. The individual monomers are colored differently. The original referent sequence is colored in green within the white referent E1 monomer and the E1 variant magenta monomer. The sequence duplication in the E1-1374∧63nt variants is located at the junction between 2 adjacent E1 monomers within the structure and is highlighted in red on panel D. Positions of potentially phosphorylated threonines and amino acid substitution at position 63 are only highlighted on the white and magenta monomers.
Mentions: Structural prediction of the variant and the referent E1 proteins using the SAM-T08 [21], I-TASSER [22] and Phyre2 [23] services resulted in several models. However, only the E1 Phyre2 model of the E1-1374∧63nt variant could be reconciled with the solved structure of the E1 hexamer (PDB ID:2GXA) [25], without any overlaps between individual E1 monomer structures. The E1 reference protein model was refined by the I-TASSER server using the Phyre2 E1-1374∧63nt variant structure as a constraint. Figure 2 presents the predicted full length structural models of the reference and the E1-1374∧63nt variant proteins. From the structural prediction, it appears that both E1-C1053 and E1-1374∧63nt are positioned on the surface of the E1 protein. Additional three dimensional representations of the predicted models are presented in Supplement material (Figure S1).

Bottom Line: On the other hand, changes within the E1 region were the major differences from the A2 sub-lineage, which has been historically but inconclusively associated with high grade cervical disease.However, this study shows that the variations within the E1 region could possibly affect cervical disease, since the E1-1374^63nt E-G350 variant is significantly associated with lower grade cervical lesions, in comparison to the A1 and A2 sub-lineage variants.Furthermore, it appears that the silent variation 109T>C of the E-C109/G350 variant might have a significant role in the viral life cycle and warrants further study.

View Article: PubMed Central - PubMed

Affiliation: Division of Molecular Medicine, Rudjer Boskovic Institute, Zagreb, Croatia.

ABSTRACT

Background: The variation of the most common Human papillomavirus (HPV) type found in cervical cancer, the HPV16, has been extensively investigated in almost all viral genes. The E1 gene variation, however, has been rarely studied. The main objective of the present investigation was to analyze the variability of the E6 and E1 genes, focusing on the recently identified E1-1374^63nt variant.

Methodology/principal findings: Variation within the E6 of 786 HPV16 positive cervical samples was analyzed using high-resolution melting, while the E1-1374^63nt duplication was assayed by PCR. Both techniques were supplemented with sequencing. The E1-1374^63nt duplication was linked with the E-G350 and the E-C109/G350 variants. In comparison to the referent HPV16, the E1-1374^63nt E-G350 variant was significantly associated with lower grade cervical lesions (p = 0.029), while the E1-1374^63nt E-C109/G350 variant was equally distributed between high and low grade lesions. The E1-1374^63nt variants were phylogenetically closest to E-G350 variant lineage (A2 sub-lineage based on full genome classification). The major differences between E1-1374^63nt variants were within the LCR and the E6 region. On the other hand, changes within the E1 region were the major differences from the A2 sub-lineage, which has been historically but inconclusively associated with high grade cervical disease. Thus, the shared variations cannot explain the particular association of the E1-1374^63nt variant with lower grade cervical lesions.

Conclusions/significance: The E1 region has been thus far considered to be well conserved among all HPVs and therefore uninteresting for variability studies. However, this study shows that the variations within the E1 region could possibly affect cervical disease, since the E1-1374^63nt E-G350 variant is significantly associated with lower grade cervical lesions, in comparison to the A1 and A2 sub-lineage variants. Furthermore, it appears that the silent variation 109T>C of the E-C109/G350 variant might have a significant role in the viral life cycle and warrants further study.

Show MeSH
Related in: MedlinePlus