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.

Genetics and Genomics

Multi-omics mapping of human papillomavirus integration sites illuminates novel cervical cancer target genes

Abstract

Background

Integration of human papillomavirus (HPV) into the host genome is a dominant feature of invasive cervical cancer (ICC), yet the tumorigenicity of cis genomic changes at integration sites remains largely understudied.

Methods

Combining multi-omics data from The Cancer Genome Atlas with patient-matched long-read sequencing of HPV integration sites, we developed a strategy for using HPV integration events to identify and prioritise novel candidate ICC target genes (integration-detected genes (IDGs)). Four IDGs were then chosen for in vitro functional studies employing small interfering RNA-mediated knockdown in cell migration, proliferation and colony formation assays.

Results

PacBio data revealed 267 unique human–HPV breakpoints comprising 87 total integration events in eight tumours. Candidate IDGs were filtered based on the following criteria: (1) proximity to integration site, (2) clonal representation of integration event, (3) tumour-specific expression (Z-score) and (4) association with ICC survival. Four candidates prioritised based on their unknown function in ICC (BNC1, RSBN1, USP36 and TAOK3) exhibited oncogenic properties in cervical cancer cell lines. Further, annotation of integration events provided clues regarding potential mechanisms underlying altered IDG expression in both integrated and non-integrated ICC tumours.

Conclusions

HPV integration events can guide the identification of novel IDGs for further study in cervical carcinogenesis and as putative therapeutic targets.

Access options

Rent or Buy article

Get time limited or full article access on ReadCube.

from$8.99

All prices are NET prices.

Fig. 1: Filtering and prioritisation results of four candidate IDGs chosen for validation and functional testing.
Fig. 2: Annotation of HPV integration affecting BNC1 expression in TCGA-C5-A2LV.
Fig. 3: Annotation of HPV integration affecting RSBN1 expression in TCGA-C5-A3HD.
Fig. 4: Annotation of HPV integration affecting USP36 expression in TCGA-C5-A8XH.
Fig. 5: Annotation of HPV integration affecting TAOK3 expression in TCGA-C5-A2LX.
Fig. 6: Functional testing of candidate IDGs in cervical cancer cell lines.

Data availability

The PacBio long-read sequencing data generated in this study have been submitted to the NCBI BioProject database under accession number PRJNA640649.

Code availability

All codes used to generate results presented are publicly available and cited with the first mention.

References

  1. 1.

    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. 2004;64:3878–84.

    CAS  PubMed  Article  Google Scholar 

  2. 2.

    Pett M, Coleman N. Integration of high-risk human papillomavirus: a key event in cervical carcinogenesis? J Pathol. 2007;212:356–67.

    CAS  PubMed  Article  Google Scholar 

  3. 3.

    Moody CA, Laimins LA. Human papillomavirus oncoproteins: pathways to transformation. Nat Rev Cancer. 2010;10:550–60.

    CAS  PubMed  Article  Google Scholar 

  4. 4.

    Bester AC, Roniger M, Oren YS, Im MM, Sarni D, Chaoat M, et al. Nucleotide deficiency promotes genomic instability in early stages of cancer development. Cell. 2011;145:435–46.

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  5. 5.

    Parfenov M, Pedamallu CS, Gehlenborg N, Freeman SS, Danilova L, Bristow CA, et al. Characterization of HPV and host genome interactions in primary head and neck cancers. Proc Natl Acad Sci USA. 2014;111:15544–9.

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  6. 6.

    Banister CE, Liu C, Pirisi L, Creek KE, Buckhaults PJ. Identification and characterization of HPV-independent cervical cancers. Oncotarget. 2017;8:13375–86.

    PubMed  PubMed Central  Article  Google Scholar 

  7. 7.

    Yuan H, Krawczyk E, Blancato J, Albanese C, Zhou D, Wang N, et al. HPV positive neuroendocrine cervical cancer cells are dependent on Myc but not E6/E7 viral oncogenes. Sci Rep. 2017;7:45617.

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  8. 8.

    Adey A, Burton JN, Kitzman JO, Hiatt JB, Lewis AP, Martin BK, et al. The haplotype-resolved genome and epigenome of the aneuploid HeLa cancer cell line. Nature. 2013;500:207–11.

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  9. 9.

    Warburton A, Redmond CJ, Dooley KE, Fu H, Gillison ML, Akagi K, et al. HPV integration hijacks and multimerizes a cellular enhancer to generate a viral-cellular super-enhancer that drives high viral oncogene expression. PLoS Genet. 2018;14:e1007179.

    PubMed  PubMed Central  Article  CAS  Google Scholar 

  10. 10.

    Kadaja M, Isok-Paas H, Laos T, Ustav E, Ustav M. Mechanism of genomic instability in cells infected with the high-risk human papillomaviruses. PLoS Pathog. 2009;5:e1000397.

    PubMed  PubMed Central  Article  CAS  Google Scholar 

  11. 11.

    Cancer Genome Atlas Research, N., Albert Einstein College of, M., Analytical Biological, S., Barretos Cancer, H., Baylor College of, M., Beckman Research Institute of City of, H. et al. Integrated genomic and molecular characterization of cervical cancer. Nature. 2017;543:378–84.

    Article  CAS  Google Scholar 

  12. 12.

    McBride AA, Warburton A. The role of integration in oncogenic progression of HPV-associated cancers. PLoS Pathog. 2017;13:e1006211.

    PubMed  PubMed Central  Article  CAS  Google Scholar 

  13. 13.

    Ojesina AI, Lichtenstein L, Freeman SS, Pedamallu CS, Imaz-Rosshandler I, Pugh TJ, et al. Landscape of genomic alterations in cervical carcinomas. Nature. 2014;506:371–5.

    CAS  PubMed  Article  Google Scholar 

  14. 14.

    Tang KW, Alaei-Mahabadi B, Samuelsson T, Lindh M, Larsson E. The landscape of viral expression and host gene fusion and adaptation in human cancer. Nat Commun. 2013;4:2513.

    PubMed  Article  CAS  Google Scholar 

  15. 15.

    Berger AC, Korkut A, Kanchi RS, Hegde AM, Lenoir W, Liu W, et al. A comprehensive pan-cancer molecular study of gynecologic and breast cancers. Cancer Cell. 2018;33:690–705. e699

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  16. 16.

    Sack LM, Davoli T, Li MZ, Li Y, Xu Q, Naxerova K, et al. Profound tissue specificity in proliferation control underlies cancer drivers and aneuploidy patterns. Cell. 2018;173:499–514. e423

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  17. 17.

    Schneider G, Schmidt-Supprian M, Rad R, Saur D. Tissue-specific tumorigenesis: context matters. Nat Rev Cancer. 2017;17:239–53.

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  18. 18.

    Zhang Z, Borecki I, Nguyen L, Ma D, Smith K, Huettner PC, et al. CD83 gene polymorphisms increase susceptibility to human invasive cervical cancer. Cancer Res. 2007;67:11202–8.

    CAS  PubMed  Article  Google Scholar 

  19. 19.

    Liu P, Iden M, Fye S, Huang YW, Hopp E, Chu C, et al. Targeted, deep sequencing reveals full methylation profiles of multiple HPV types and potential biomarkers for cervical cancer progression. Cancer Epidemiol Biomark Prev. 2017;26:642–50.

    CAS  Article  Google Scholar 

  20. 20.

    Nguyen ND, Deshpande V, Luebeck J, Mischel PS, Bafna V. ViFi: accurate detection of viral integration and mRNA fusion reveals indiscriminate and unregulated transcription in proximal genomic regions in cervical cancer. Nucleic Acids Res. 2018;46:3309–25.

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  21. 21.

    Chen X, Kost J, Sulovari A, Wong N, Liang WS, Cao J, et al. A virome-wide clonal integration analysis platform for discovering cancer viral etiology. Genome Res. 2019;29:819–30.

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  22. 22.

    Guo Y, Long J, He J, Li CI, Cai Q, Shu XO, et al. Exome sequencing generates high quality data in non-target regions. BMC Genomics. 2012;13:194.

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  23. 23.

    R Foundation for Statistical Computing. R: a language and environment for statistical computing. Vienna: R Foundation for Statistical Computing; 2013.

  24. 24.

    Hadley W. Ggplot2. New York: Springer Science+Business Media, LLC; 2016.

  25. 25.

    Martin MP, Borecki IB, Zhang Z, Nguyen L, Ma D, Gao X, et al. HLA-Cw group 1 ligands for KIR increase susceptibility to invasive cervical cancer. Immunogenetics. 2010;62:761–5.

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  26. 26.

    Yu KJ, Rader JS, Borecki I, Zhang Z, Hildesheim A. CD83 polymorphisms and cervical cancer risk. Gynecol Oncol. 2009;114:319–22.

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  27. 27.

    Kent WJ, Sugnet CW, Furey TS, Roskin KM, Pringle TH, Zahler AM, et al. The human genome browser at UCSC. Genome Res. 2002;12:996–1006.

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  28. 28.

    Fishilevich S, Nudel R, Rappaport N, Hadar R, Plaschkes I, Iny Stein T, et al. GeneHancer: genome-wide integration of enhancers and target genes in GeneCards. Database. 2017; 2017: bax028. https://doi.org/10.1093/database/bax028.

  29. 29.

    Davis CA, Hitz BC, Sloan CA, Chan ET, Davidson JM, Gabdank I, et al. The Encyclopedia of DNA elements (ENCODE): data portal update. Nucleic Acids Res. 2018;46:D794–D801.

    CAS  PubMed  Article  Google Scholar 

  30. 30.

    Zheng R, Wan C, Mei S, Qin Q, Wu Q, Sun H, et al. Cistrome Data Browser: expanded datasets and new tools for gene regulatory analysis. Nucleic Acids Res. 2019;47:D729–D735.

    CAS  PubMed  Article  Google Scholar 

  31. 31.

    Robinson JT, Thorvaldsdottir H, Winckler W, Guttman M, Lander ES, Getz G, et al. Integrative genomics viewer. Nat Biotechnol. 2011;29:24–26.

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  32. 32.

    Thorvaldsdottir H, Robinson JT, Mesirov JP. Integrative Genomics Viewer (IGV): high-performance genomics data visualization and exploration. Brief Bioinform. 2013;14:178–92.

    CAS  PubMed  Article  Google Scholar 

  33. 33.

    Nattestad M, Aboukhalil R, Chin CS, Schatz MC. Ribbon: intuitive visualization for complex genomic variation. Bioinformatics. 2020. https://doi.org/10.1093/bioinformatics/btaa680

    Article  PubMed  PubMed Central  Google Scholar 

  34. 34.

    Koch A, Jeschke J, Van Criekinge W, van Engeland M, De Meyer T. MEXPRESS update 2019. Nucleic Acids Res. 2019;47:W561–65.

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  35. 35.

    Rau A, Flister M, Rui H, Auer PL. Exploring drivers of gene expression in the Cancer Genome Atlas. Bioinformatics. 2018. https://doi.org/10.1093/bioinformatics/bty551.

  36. 36.

    Iden M, Fye S, Li K, Chowdhury T, Ramchandran R, Rader JS. The lncRNA PVT1 contributes to the cervical cancer phenotype and associates with poor patient prognosis. PLoS ONE. 2016;11:e0156274.

    PubMed  PubMed Central  Article  CAS  Google Scholar 

  37. 37.

    Cai Z, Chattopadhyay N, Liu WJ, Chan C, Pignol JP, Reilly RM. Optimized digital counting colonies of clonogenic assays using ImageJ software and customized macros: comparison with manual counting. Int J Radiat Biol. 2011;87:1135–46.

    CAS  PubMed  Article  PubMed Central  Google Scholar 

  38. 38.

    Durst M, Croce CM, Gissmann L, Schwarz E, Huebner K. Papillomavirus sequences integrate near cellular oncogenes in some cervical carcinomas. Proc Natl Acad Sci USA. 1987;84:1070–4.

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  39. 39.

    Bodelon C, Untereiner ME, Machiela MJ, Vinokurova S, Wentzensen N. Genomic characterization of viral integration sites in HPV-related cancers. Int J Cancer. 2016;139:2001–11.

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  40. 40.

    Cannizzaro LA, Durst M, Mendez MJ, Hecht BK, Hecht F. Regional chromosome localization of human papillomavirus integration sites near fragile sites, oncogenes, and cancer chromosome breakpoints. Cancer Genet Cytogenet. 1988;33:93–98.

    CAS  PubMed  Article  Google Scholar 

  41. 41.

    Rao SS, Huntley MH, Durand NC, Stamenova EK, Bochkov ID, Robinson JT, et al. A 3D map of the human genome at kilobase resolution reveals principles of chromatin looping. Cell. 2014;159:1665–80.

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  42. 42.

    Schuijers J, Manteiga JC, Weintraub AS, Day DS, Zamudio AV, Hnisz D, et al. Transcriptional dysregulation of MYC reveals common enhancer-docking mechanism. Cell Rep. 2018;23:349–60.

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  43. 43.

    Dixon JR, Selvaraj S, Yue F, Kim A, Li Y, Shen Y, et al. Topological domains in mammalian genomes identified by analysis of chromatin interactions. Nature. 2012;485:376–80.

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  44. 44.

    Belokopytova PS, Nuriddinov MA, Mozheiko EA, Fishman D, Fishman V. Quantitative prediction of enhancer-promoter interactions. Genome Res. 2020;30:72–84.

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  45. 45.

    Stelzer G, Rosen N, Plaschkes I, Zimmerman S, Twik M, Fishilevich S. et al. The GeneCards Suite: from gene data mining to disease genome sequence analyses. Curr Protoc Bioinform. 2016;54:1.30.1–33.

    Article  Google Scholar 

  46. 46.

    Turner KM, Deshpande V, Beyter D, Koga T, Rusert J, Lee C, et al. Extrachromosomal oncogene amplification drives tumour evolution and genetic heterogeneity. Nature. 2017;543:122–5.

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  47. 47.

    Holmes A, Lameiras S, Jeannot E, Marie Y, Castera L, Sastre-Garau X, et al. Mechanistic signatures of HPV insertions in cervical carcinomas. NPJ Genom Med. 2016;1:16004.

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  48. 48.

    Ren G, Jin W, Cui K, Rodrigez J, Hu G, Zhang Z. et al. CTCF-mediated enhancer-promoter interaction is a critical regulator of cell-to-cell variation of gene expression. Mol Cell. 2017;67:1049–58. e1046.

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  49. 49.

    Ron G, Globerson Y, Moran D, Kaplan T. Promoter-enhancer interactions identified from Hi-C data using probabilistic models and hierarchical topological domains. Nat Commun. 2017;8:2237.

    PubMed  PubMed Central  Article  CAS  Google Scholar 

  50. 50.

    Javierre BM, Burren OS, Wilder SP, Kreuzhuber R, Hill SM, Sewitz S. et al. Lineage-specific genome architecture links enhancers and non-coding disease variants to target gene promoters. Cell. 2016;167:1369–84. e1319.

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  51. 51.

    Kron KJ, Bailey SD, Lupien M. Enhancer alterations in cancer: a source for a cell identity crisis. Genome Med. 2014;6:77.

    PubMed  PubMed Central  Article  CAS  Google Scholar 

  52. 52.

    Webb E, Adams JM, Cory S. Variant (6; 15) translocation in a murine plasmacytoma occurs near an immunoglobulin kappa gene but far from the myc oncogene. Nature. 1984;312:777–9.

    CAS  PubMed  Article  Google Scholar 

  53. 53.

    Graham M, Adams JM. Chromosome 8 breakpoint far 3’ of the c-myc oncogene in a Burkitt’s lymphoma 2;8 variant translocation is equivalent to the murine pvt-1 locus. EMBO J. 1986;5:2845–51.

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  54. 54.

    Guan Y, Kuo WL, Stilwell JL, Takano H, Lapuk AV, Fridlyand J, et al. Amplification of PVT1 contributes to the pathophysiology of ovarian and breast cancer. Clin Cancer Res. 2007;13:5745–55.

    CAS  PubMed  Article  Google Scholar 

  55. 55.

    Haynes WA, Tomczak A, Khatri P. Gene annotation bias impedes biomedical research. Sci Rep. 2018;8:1362.

    PubMed  PubMed Central  Article  CAS  Google Scholar 

  56. 56.

    Maertens A, Tran VH, Maertens M, Kleensang A, Luechtefeld TH, Hartung T, et al. Functionally enigmatic genes in cancer: using TCGA data to map the limitations of annotations. Sci Rep. 2020;10:4106.

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  57. 57.

    Akagi K, Li J, Broutian TR, Padilla-Nash H, Xiao W, Jiang B, et al. Genome-wide analysis of HPV integration in human cancers reveals recurrent, focal genomic instability. Genome Res. 2014;24:185–99.

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  58. 58.

    Kantidze OL, Gurova KV, Studitsky VM, Razin SV. The 3D genome as a target for anticancer therapy. Trends Mol Med. 2020;26:141–9.

    CAS  PubMed  Article  Google Scholar 

  59. 59.

    Cao C, Hong P, Huang X, Lin D, Cao G, Wang L, et al. HPV-CCDC106 integration alters local chromosome architecture and hijacks an enhancer by three-dimensional genome structure remodeling in cervical cancer. J Genet Genomics. 2020;47:437–50.

    PubMed  Article  Google Scholar 

  60. 60.

    Feuerborn A, Mathow D, Srivastava PK, Gretz N, Grone HJ. Basonuclin-1 modulates epithelial plasticity and TGF-beta1-induced loss of epithelial cell integrity. Oncogene. 2015;34:1185–95.

    CAS  PubMed  Article  Google Scholar 

  61. 61.

    Abu-Jamous B, Buffa FM, Harris AL, Nandi AK. In vitro downregulated hypoxia transcriptome is associated with poor prognosis in breast cancer. Mol Cancer. 2017;16:105.

    PubMed  PubMed Central  Article  CAS  Google Scholar 

  62. 62.

    Kim SY, Choi J, Lee DH, Park JH, Hwang YJ, Baek KH. PME-1 is regulated by USP36 in ERK and Akt signaling pathways. FEBS Lett. 2018;592:1575–88.

    CAS  PubMed  Article  Google Scholar 

  63. 63.

    Bian Y, Teper Y, Mathews Griner LA, Aiken TJ, Shukla V, Guha R, et al. Target deconvolution of a multikinase inhibitor with antimetastatic properties identifies TAOK3 as a key contributor to a cancer stem cell-like phenotype. Mol Cancer Ther. 2019;18:2097–110.

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  64. 64.

    Ormonde JVS, Li Z, Stegen C, Madrenas J. TAOK3 regulates canonical TCR signaling by preventing early SHP-1-mediated inactivation of LCK. J Immunol. 2018;201:3431–42.

    CAS  PubMed  Article  Google Scholar 

  65. 65.

    Ormonde JVS, Nie Y, Madrenas J. TAOK3, a regulator of LCK-SHP-1 crosstalk during TCR signaling. Crit Rev Immunol. 2019;39:59–81.

    PubMed  Article  Google Scholar 

Download references

Acknowledgements

We thank the Genomic Sciences and Precision Medicine Center (GSPMC) at the Medical College of Wisconsin for their assistance and technical support with PacBio sequencing.

Funding

This work was supported by the NCI R21CA241013 (to JSR) and the Women’s Health Research Programme in the Department of Obstetrics and Gynecology, Medical College of Wisconsin.

Author information

Affiliations

Authors

Contributions

MI, Y-WH, PL, MJF and JSR designed the study concept and experiments; MI, Y-WH and MX performed the experiments; MI, S-WT, Y-WH and PL acquired, analysed and interpreted the data; MI, MJF and JSR wrote the paper. All authors discussed the results and had final approval of the submitted manuscript.

Corresponding author

Correspondence to Janet S. Rader.

Ethics declarations

Competing interests

The authors declare no competing interests.

Ethics approval and consent to participate

The study protocol was approved by the Medical College of Wisconsin’s Institutional Review Board. All the cervical cell lines used in this study were purchased from the American Type Cell Collection (ATCC, Manassas, VA).

Consent to publish

Not applicable.

Additional information

Publisher’s note Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Supplementary information

Rights and permissions

Reprints and Permissions

About this article

Verify currency and authenticity via CrossMark

Cite this article

Iden, M., Tsaih, SW., Huang, YW. et al. Multi-omics mapping of human papillomavirus integration sites illuminates novel cervical cancer target genes. Br J Cancer (2021). https://doi.org/10.1038/s41416-021-01545-0

Download citation

Search

Quick links