Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

  • Letter
  • Published:

Genome-wide profiling of HPV integration in cervical cancer identifies clustered genomic hot spots and a potential microhomology-mediated integration mechanism

Nature Genetics volume 47, pages 158–163 (2015)Cite this article

Subjects

Abstract

Human papillomavirus (HPV) integration is a key genetic event in cervical carcinogenesis1. By conducting whole-genome sequencing and high-throughput viral integration detection, we identified 3,667 HPV integration breakpoints in 26 cervical intraepithelial neoplasias, 104 cervical carcinomas and five cell lines. Beyond recalculating frequencies for the previously reported frequent integration sites POU5F1B (9.7%), FHIT (8.7%), KLF12 (7.8%), KLF5 (6.8%), LRP1B (5.8%) and LEPREL1 (4.9%), we discovered new hot spots HMGA2 (7.8%), DLG2 (4.9%) and SEMA3D (4.9%). Protein expression from FHIT and LRP1B was downregulated when HPV integrated in their introns. Protein expression from MYC and HMGA2 was elevated when HPV integrated into flanking regions. Moreover, microhomologous sequence between the human and HPV genomes was significantly enriched near integration breakpoints, indicating that fusion between viral and human DNA may have occurred by microhomology-mediated DNA repair pathways2. Our data provide insights into HPV integration-driven cervical carcinogenesis.

This is a preview of subscription content, access via your institution

Access options

Buy this article

  • Purchase on Springer Link
  • Instant access to full article PDF
Buy now

Prices may be subject to local taxes which are calculated during checkout

Figure 1: Distribution of breakpoints in the human and HPV genomes in 135 samples.
Figure 2: Clinical annotation of HPV integration sites in 104 cervical carcinomas, 26 CINs and five cell lines.
Figure 3: Mapping five HPV integration hot spots in 135 samples.
Figure 4: Effects of HPV integration on gene expression in samples assessed by immunohistochemistry.
Figure 5: MH sequences are significantly enriched in the regions flanking integration sites.

Similar content being viewed by others

Accession codes

Primary accessions

Sequence Read Archive

Referenced accessions

NCBI Reference Sequence

References

  1. Pett, M. & Coleman, N. Integration of high-risk human papillomavirus: a key event in cervical carcinogenesis? J. Pathol. 212, 356–367 (2007).

    Article  CAS  Google Scholar 

  2. Lawson, A.R. et al. RAF gene fusion breakpoints in pediatric brain tumors are characterized by significant enrichment of sequence microhomology. Genome Res. 21, 505–514 (2011).

    Article  CAS  Google Scholar 

  3. zur Hausen, H. Papillomaviruses and cancer: from basic studies to clinical application. Nat. Rev. Cancer 2, 342–350 (2002).

    Article  CAS  Google Scholar 

  4. Crosbie, E.J., Einstein, M.H., Franceschi, S. & Kitchener, H.C. Human papillomavirus and cervical cancer. Lancet 382, 889–899 (2013).

    Article  Google Scholar 

  5. Shi, Y. et al. A genome-wide association study identifies two new cervical cancer susceptibility loci at 4q12 and 17q12. Nat. Genet. 45, 918–922 (2013).

    Article  CAS  Google Scholar 

  6. Stanley, M.A., Pett, M.R. & Coleman, N. HPV: from infection to cancer. Biochem. Soc. Trans. 35, 1456–1460 (2007).

    Article  CAS  Google Scholar 

  7. Wentzensen, N., Vinokurova, S. & von Knebel Doeberitz, M. Systematic review of genomic integration sites of human papillomavirus genomes in epithelial dysplasia and invasive cancer of the female lower genital tract. Cancer Res. 64, 3878–3884 (2004).

    Article  CAS  Google Scholar 

  8. Hudelist, G. et al. Physical state and expression of HPV DNA in benign and dysplastic cervical tissue: different levels of viral integration are correlated with lesion grade. Gynecol. Oncol. 92, 873–880 (2004).

    Article  CAS  Google Scholar 

  9. Arias-Pulido, H., Peyton, C.L., Joste, N.E., Vargas, H. & Wheeler, C.M. Human papillomavirus type 16 integration in cervical carcinoma in situ and in invasive cervical cancer. J. Clin. Microbiol. 44, 1755–1762 (2006).

    Article  CAS  Google Scholar 

  10. el Awady, M.K., Kaplan, J.B., O'Brien, S.J. & Burk, R.D. Molecular analysis of integrated human papillomavirus 16 sequences in the cervical cancer cell line SiHa. Virology 159, 389–398 (1987).

    Article  CAS  Google Scholar 

  11. Thorland, E.C. et al. Human papillomavirus type 16 integrations in cervical tumors frequently occur in common fragile sites. Cancer Res. 60, 5916–5921 (2000).

    CAS  PubMed  Google Scholar 

  12. Luft, F. et al. Detection of integrated papillomavirus sequences by ligation-mediated PCR (DIPS-PCR) and molecular characterization in cervical cancer cells. Int. J. Cancer 92, 9–17 (2001).

    Article  CAS  Google Scholar 

  13. Kalantari, M., Blennow, E., Hagmar, B. & Johansson, B. Physical state of HPV16 and chromosomal mapping of the integrated form in cervical carcinomas. Diagn. Mol. Pathol. 10, 46–54 (2001).

    Article  CAS  Google Scholar 

  14. Xu, B. et al. Multiplex identification of human papillomavirus 16 DNA integration sites in cervical carcinomas. PLoS ONE 8, e66693 (2013).

    Article  CAS  Google Scholar 

  15. Klaes, R. et al. Detection of high-risk cervical intraepithelial neoplasia and cervical cancer by amplification of transcripts derived from integrated papillomavirus oncogenes. Cancer Res. 59, 6132–6136 (1999).

    CAS  PubMed  Google Scholar 

  16. Li, W. et al. HIVID: an efficient method to detect HBV integration using low coverage sequencing. Genomics 102, 338–344 (2013).

    Article  CAS  Google Scholar 

  17. Dall, K.L. et al. Characterization of naturally occurring HPV16 integration sites isolated from cervical keratinocytes under noncompetitive conditions. Cancer Res. 68, 8249–8259 (2008).

    Article  CAS  Google Scholar 

  18. Ferber, M.J. et al. Preferential integration of human papillomavirus type 18 near the c-myc locus in cervical carcinoma. Oncogene 22, 7233–7242 (2003).

    Article  CAS  Google Scholar 

  19. Schmitz, M. et al. Loss of gene function as a consequence of human papillomavirus DNA integration. Int. J. Cancer 131, E593–E602 (2012).

    Article  CAS  Google Scholar 

  20. Peter, M. et al. MYC activation associated with the integration of HPV DNA at the MYC locus in genital tumors. Oncogene 25, 5985–5993 (2006).

    Article  CAS  Google Scholar 

  21. Boulet, G.A. et al. Human papillomavirus 16 load and E2/E6 ratio in HPV16-positive women: biomarkers for cervical intraepithelial neoplasia >or=2 in a liquid-based cytology setting? Cancer Epidemiol. Biomarkers Prev. 18, 2992–2999 (2009).

    Article  CAS  Google Scholar 

  22. Gradíssimo Oliveira, A., Delgado, C., Verdasca, N. & Pista, A. Prognostic value of human papillomavirus types 16 and 18 DNA physical status in cervical intraepithelial neoplasia. Clin. Microbiol. Infect. 19, E447–E450 (2013).

    Article  Google Scholar 

  23. Adey, A. et al. The haplotype-resolved genome and epigenome of the aneuploid HeLa cancer cell line. Nature 500, 207–211 (2013).

    Article  CAS  Google Scholar 

  24. Ojesina, A.I. et al. Landscape of genomic alterations in cervical carcinomas. Nature 506, 371–375 (2014).

    Article  CAS  Google Scholar 

  25. Schmitz, M., Driesch, C., Jansen, L., Runnebaum, I.B. & Durst, M. Non-random integration of the HPV genome in cervical cancer. PLoS ONE 7, e39632 (2012).

    Article  CAS  Google Scholar 

  26. Lee, J.A., Carvalho, C.M. & Lupski, J.R.A. DNA replication mechanism for generating nonrecurrent rearrangements associated with genomic disorders. Cell 131, 1235–1247 (2007).

    Article  CAS  Google Scholar 

  27. Zhang, F. et al. The DNA replication FoSTeS/MMBIR mechanism can generate genomic, genic and exonic complex rearrangements in humans. Nat. Genet. 41, 849–853 (2009).

    Article  CAS  Google Scholar 

  28. Verdin, H. et al. Microhomology-mediated mechanisms underlie non-recurrent disease-causing microdeletions of the FOXL2 gene or its regulatory domain. PLoS Genet. 9, e1003358 (2013).

    Article  CAS  Google Scholar 

  29. Campitelli, M. et al. Human papillomavirus mutational insertion: specific marker of circulating tumor DNA in cervical cancer patients. PLoS ONE 7, e43393 (2012).

    Article  CAS  Google Scholar 

  30. Das, P. et al. HPV genotyping and site of viral integration in cervical cancers in Indian women. PLoS ONE 7, e41012 (2012).

    Article  CAS  Google Scholar 

  31. Karlsen, F. et al. Use of multiple PCR primer sets for optimal detection of human papillomavirus. J. Clin. Microbiol. 34, 2095–2100 (1996).

    CAS  PubMed  PubMed Central  Google Scholar 

  32. Baay, M.F. et al. Comprehensive study of several general and type-specific primer pairs for detection of human papillomavirus DNA by PCR in paraffin-embedded cervical carcinomas. J. Clin. Microbiol. 34, 745–747 (1996).

    CAS  PubMed  PubMed Central  Google Scholar 

  33. Walboomers, J.M. et al. Human papillomavirus is a necessary cause of invasive cervical cancer worldwide. J. Pathol. 189, 12–19 (1999).

    Article  CAS  Google Scholar 

  34. Woodman, C.B., Collins, S.I. & Young, L.S. The natural history of cervical HPV infection: unresolved issues. Nat. Rev. Cancer 7, 11–22 (2007).

    Article  CAS  Google Scholar 

  35. Stanley, M.A., Browne, H.M., Appleby, M. & Minson, A.C. Properties of a non-tumorigenic human cervical keratinocyte cell line. Int. J. Cancer 43, 672–676 (1989).

    Article  CAS  Google Scholar 

  36. Li, H. & Durbin, R. Fast and accurate short read alignment with Burrows-Wheeler transform. Bioinformatics 25, 1754–1760 (2009).

    Article  CAS  Google Scholar 

  37. Wang, K., Li, M. & Hakonarson, H. ANNOVAR: functional annotation of genetic variants from high-throughput sequencing data. Nucleic Acids Res. 38, e164 (2010).

    Article  Google Scholar 

  38. Lettice, L.A. et al. A long-range Shh enhancer regulates expression in the developing limb and fin and is associated with preaxial polydactyly. Hum. Mol. Genet. 12, 1725–1735 (2003).

    Article  CAS  Google Scholar 

  39. Li, L. et al. A far downstream enhancer for murine Bcl11b controls its T-cell specific expression. Blood 122, 902–911 (2013).

    Article  CAS  Google Scholar 

  40. Sung, W.K. et al. Genome-wide survey of recurrent HBV integration in hepatocellular carcinoma. Nat. Genet. 44, 765–769 (2012).

    Article  CAS  Google Scholar 

  41. Burk, R.D., Harari, A. & Chen, Z. Human papillomavirus genome variants. Virology 445, 232–243 (2013).

    Article  CAS  Google Scholar 

  42. Cer, R.Z. et al. Non-B DB v2.0: a database of predicted non-B DNA-forming motifs and its associated tools. Nucleic Acids Res. 41, D94–D100 (2013).

    Article  CAS  Google Scholar 

  43. Abeysinghe, S.S., Chuzhanova, N., Krawczak, M., Ball, E.V. & Cooper, D.N. Translocation and gross deletion breakpoints in human inherited disease and cancer I: nucleotide composition and recombination-associated motifs. Hum. Mutat. 22, 229–244 (2003).

    Article  CAS  Google Scholar 

Download references

Acknowledgements

This work was supported by funds from the National Development Program (973) for the Key Basic Research of China (2015CB553903 and 2013CB911304) and the National Natural Science Funding of China (81230038, 81372805, 81172466, 81272859, 81372801, 81372804, 81402158, 81370469, 81372806, 81370469, 81172468, 81101964, 81230052 and 81302266). The study is also sponsored by the Chinese 863 Program (2012AA02A507, 2012AA02A201), Guangdong Enterprise Key Laboratory of Human Disease Genomics, ShenZhen Engineering Laboratory for Clinical Molecular Diagnostic and China National GeneBank–Shenzhen. We thank all participants recruited for this study. We thank our colleagues from BGI, H. Huang, L. Huang, X. Zhuang, L. Lin, H. Cao, X. Fang, X. Zhang, Y. Shuang and H. Yang for sequencing and analysis.

Author information

Author notes

  1. Zheng Hu, Da Zhu, Wei Wang, Weiyang Li, Wenlong Jia and Xi Zeng: These authors contributed equally to this work.

Authors and Affiliations

  1. Department of Obstetrics and Gynecology, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei, China.,

    Zheng Hu, Da Zhu, Wencheng Ding, Lan Yu, Xiaoli Wang, Liming Wang, Hui Shen, Changlin Zhang, Gang Chen, Qinglei Gao, Shuang Li, Ling Xi, Changyu Wang, Shujie Liao, Xiangyi Ma, Peng Wu, Kezhen Li, Shixuan Wang, Hui Wang & Ding Ma

  2. Department of Obstetrics and Gynecology, Nanfang Hospital, Southern Medical University, Guangzhou, China

    Wei Wang

  3. Beijing Genomics Institute (BGI),

    Weiyang Li, Wenlong Jia, Xi Zeng, Hongjie Liu, Xiao Liu, Yi Zhao, Xiaodong Fang, Wei Chen, Jun Wang & Xun Xu

  4. -Shenzhen, Shenzhen, China

    Weiyang Li, Wenlong Jia, Xi Zeng, Hongjie Liu, Xiao Liu, Yi Zhao, Xiaodong Fang, Wei Chen, Jun Wang & Xun Xu

  5. School of Bioscience and Bioengineering, South China University of Technology, Guangzhou, China

    Weiyang Li & Jun Wang

  6. Department of Computer Science, City University of Hong Kong, Hong Kong, China

    Wenlong Jia & Shuaicheng Li

  7. WuHan Frasergen Bioinformatics Co., Ltd , Wuhan, China,

    Tang Tang

  8. Wuhan Institute of Biotechnology, Wuhan, China

    Aisi Fu & Zou Wang

  9. Department of Hematology, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei, China.,

    Jianfeng Zhou

  10. Princess Al Jawhara Center of Excellence in the Research of Hereditary Disorders, King Abdulaziz University, Jeddah, Saudi Arabia

    Jun Wang

  11. The Novo Nordisk Foundation Center for Basic Metabolic Research, University of Copenhagen, Copenhagen, Denmark

    Jun Wang

  12. Centre for iSequencing, Aarhus University, Aarhus, Denmark

    Jun Wang

Authors
  1. Zheng Hu
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Da Zhu
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Wei Wang
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Weiyang Li
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Wenlong Jia
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Xi Zeng
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Wencheng Ding
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Lan Yu
    View author publications

    You can also search for this author in PubMed Google Scholar

  9. Xiaoli Wang
    View author publications

    You can also search for this author in PubMed Google Scholar

  10. Liming Wang
    View author publications

    You can also search for this author in PubMed Google Scholar

  11. Hui Shen
    View author publications

    You can also search for this author in PubMed Google Scholar

  12. Changlin Zhang
    View author publications

    You can also search for this author in PubMed Google Scholar

  13. Hongjie Liu
    View author publications

    You can also search for this author in PubMed Google Scholar

  14. Xiao Liu
    View author publications

    You can also search for this author in PubMed Google Scholar

  15. Yi Zhao
    View author publications

    You can also search for this author in PubMed Google Scholar

  16. Xiaodong Fang
    View author publications

    You can also search for this author in PubMed Google Scholar

  17. Shuaicheng Li
    View author publications

    You can also search for this author in PubMed Google Scholar

  18. Wei Chen
    View author publications

    You can also search for this author in PubMed Google Scholar

  19. Tang Tang
    View author publications

    You can also search for this author in PubMed Google Scholar

  20. Aisi Fu
    View author publications

    You can also search for this author in PubMed Google Scholar

  21. Zou Wang
    View author publications

    You can also search for this author in PubMed Google Scholar

  22. Gang Chen
    View author publications

    You can also search for this author in PubMed Google Scholar

  23. Qinglei Gao
    View author publications

    You can also search for this author in PubMed Google Scholar

  24. Shuang Li
    View author publications

    You can also search for this author in PubMed Google Scholar

  25. Ling Xi
    View author publications

    You can also search for this author in PubMed Google Scholar

  26. Changyu Wang
    View author publications

    You can also search for this author in PubMed Google Scholar

  27. Shujie Liao
    View author publications

    You can also search for this author in PubMed Google Scholar

  28. Xiangyi Ma
    View author publications

    You can also search for this author in PubMed Google Scholar

  29. Peng Wu
    View author publications

    You can also search for this author in PubMed Google Scholar

  30. Kezhen Li
    View author publications

    You can also search for this author in PubMed Google Scholar

  31. Shixuan Wang
    View author publications

    You can also search for this author in PubMed Google Scholar

  32. Jianfeng Zhou
    View author publications

    You can also search for this author in PubMed Google Scholar

  33. Jun Wang
    View author publications

    You can also search for this author in PubMed Google Scholar

  34. Xun Xu
    View author publications

    You can also search for this author in PubMed Google Scholar

  35. Hui Wang
    View author publications

    You can also search for this author in PubMed Google Scholar

  36. Ding Ma
    View author publications

    You can also search for this author in PubMed Google Scholar

Contributions

D.M. took full responsibility for the study, especially in conceiving, designing and supervising the research together with Z.H. and H.W. W.W., J.Z., J.W. and X.X. participated in the study design. G.C., Q.G., Shuang Li., L.X., C.W., S. Liao, X.M., P.W., K.L. and S.W. supervised the diagnosis of patients and subject recruitment. D.Z., H.S., W.J., W.L., X.Z., H.L., X.F., Shuaicheng Li and X.L. performed statistical analysis. W.L., X.Z. and Y.Z. performed HIVID and RNA-seq analysis. W.J. performed FuseSV (in-house software for MH analysis) and WGS analysis. W.D., L.Y., L.W., H.S., X.W., C.Z., W.C., T.T., A.F. and Z.W. performed the experiments. The manuscript was drafted by Z.H., D.Z., W.J., W.L. and X.Z. under the supervision of D.M., H.W. and X.X. All authors critically reviewed the article and approved the final manuscript.

Corresponding authors

Correspondence to Xun Xu, Hui Wang or Ding Ma.

Ethics declarations

Competing interests

The authors declare no competing financial interests.

Supplementary information

Supplementary Text and Figures

Supplementary Figures 1–29 and Supplementary Note (PDF 23284 kb)

Supplementary Tables 1–28

Supplementary Tables 1–28 (XLSX 3257 kb)

Source data

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Hu, Z., Zhu, D., Wang, W. et al. Genome-wide profiling of HPV integration in cervical cancer identifies clustered genomic hot spots and a potential microhomology-mediated integration mechanism. Nat Genet 47, 158–163 (2015). https://doi.org/10.1038/ng.3178

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/ng.3178

This article is cited by

Search

Advanced search

Quick links

Nature Briefing: Translational Research

Sign up for the Nature Briefing: Translational Research newsletter — top stories in biotechnology, drug discovery and pharma.

Get what matters in translational research, free to your inbox weekly. Sign up for Nature Briefing: Translational Research