Merkel Cell Polyomavirus Large T Antigen is Dispensable in G2 and M-Phase to Promote Proliferation of Merkel Cell Carcinoma Cells
Abstract
:1. Introduction
2. Materials and Methods
2.1. Cell Culture
2.2. Vectors
2.3. Retroviral Infection
2.4. Time Lapse Microscopy
2.5. Mixed Cell Culture Assay
2.6. Cell Cycle Analysis
3. Results and Discussion
Supplementary Materials
Author Contributions
Funding
Conflicts of Interest
References
- Miller, R.W.; Rabkin, C.S. Merkel cell carcinoma and melanoma: Etiological similarities and differences. Cancer Epidemiol. Biomark. Prev. 1999, 8, 153–158. [Google Scholar]
- Rodig, S.J.; Cheng, J.W.; Wardzala, J.; DoRosario, A.; Scanlon, J.J.; Laga, A.C.; Martinez-Fernandez, A.; Barletta, J.A.; Bellizzi, A.M.; Sadasivam, S.; et al. Improved detection suggests all Merkel cell carcinomas harbor Merkel polyomavirus. J. Clin. Investig. 2012, 122, 4645–4653. [Google Scholar] [CrossRef] [Green Version]
- Houben, R.; Shuda, M.; Weinkam, R.; Schrama, D.; Feng, H.; Chang, Y.; Moore, P.S.; Becker, J.C. Merkel cell polyomavirus-infected Merkel cell carcinoma cells require expression of viral T antigens. J. Virol. 2010, 84, 7064–7072. [Google Scholar] [CrossRef] [PubMed] [Green Version]
- An, P.; Saenz Robles, M.T.; Pipas, J.M. Large T antigens of polyomaviruses: Amazing molecular machines. Annu. Rev. Microbiol. 2012, 66, 213–236. [Google Scholar] [CrossRef] [PubMed]
- Becker, J.C.; Stang, A.; Hausen, A.Z.; Fischer, N.; DeCaprio, J.A.; Tothill, R.W.; Lyngaa, R.; Hansen, U.K.; Ritter, C.; Nghiem, P.; et al. Epidemiology, biology and therapy of Merkel cell carcinoma: Conclusions from the EU project IMMOMEC. Cancer Immunol. Immunother. CII 2018, 67, 341–351. [Google Scholar] [CrossRef] [PubMed]
- Shuda, M.; Kwun, H.J.; Feng, H.C.; Chang, Y.; Moore, P.S. Human Merkel cell polyomavirus small T antigen is an oncoprotein targeting the 4E-BP1 translation regulator. J. Clin. Investig. 2011, 121, 3623–3634. [Google Scholar] [CrossRef]
- Verhaegen, M.E.; Mangelberger, D.; Harms, P.W.; Eberl, M.; Wilbert, D.M.; Meireles, J.; Bichakjian, C.K.; Saunders, T.L.; Wong, S.Y.; Dlugosz, A.A. Merkel Cell Polyomavirus Small T Antigen Initiates Merkel Cell Carcinoma-like Tumor Development in Mice. Cancer Res. 2017, 77, 3151–3157. [Google Scholar] [CrossRef] [Green Version]
- Angermeyer, S.; Hesbacher, S.; Becker, J.C.; Schrama, D.; Houben, R. Merkel cell polyomavirus-positive Merkel cell carcinoma cells do not require expression of the viral small T antigen. J. Investig. Dermatol. 2013, 133, 2059–2064. [Google Scholar] [CrossRef] [PubMed] [Green Version]
- Verhaegen, M.E.; Mangelberger, D.; Harms, P.W.; Vozheiko, T.D.; Weick, J.W.; Wilbert, D.M.; Saunders, T.L.; Ermilov, A.N.; Bichakjian, C.K.; Johnson, T.M.; et al. Merkel cell polyomavirus small T antigen is oncogenic in transgenic mice. J. Investig. Dermatol. 2015, 135, 1415–1424. [Google Scholar] [CrossRef] [Green Version]
- Hesbacher, S.; Pfitzer, L.; Wiedorfer, K.; Angermeyer, S.; Borst, A.; Haferkamp, S.; Scholz, C.J.; Wobser, M.; Schrama, D.; Houben, R. RB1 is the crucial target of the Merkel cell polyomavirus Large T antigen in Merkel cell carcinoma cells. Oncotarget 2016, 7, 32956–32968. [Google Scholar] [CrossRef]
- Shuda, M.; Feng, H.; Kwun, H.J.; Rosen, S.T.; Gjoerup, O.; Moore, P.S.; Chang, Y. T antigen mutations are a human tumor-specific signature for Merkel cell polyomavirus. Proc. Natl. Acad. Sci. USA 2008, 105, 16272–16277. [Google Scholar] [CrossRef] [Green Version]
- Schrama, D.; Sarosi, E.-M.; Adam, C.; Ritter, C.; Kaemmerer, U.; Klopocki, E.; König, E.-M.; Utikal, J.; Becker, J.C.; Houben, R. Characterization of six Merkel cell polyomavirus-positive Merkel cell carcinoma cell lines: Integration pattern suggest that large T antigen truncating events occur before or during integration. Int. J. Cancer 2019, 145, 1020–1032. [Google Scholar] [CrossRef] [PubMed]
- McIlroy, D.; Halary, F.; Bressollette-Bodin, C. Intra-patient viral evolution in polyomavirus-related diseases. Philos. Trans. R. Soc. Lond B Biol. Sci. 2019, 374, 20180301. [Google Scholar] [CrossRef] [PubMed] [Green Version]
- DeCaprio, J.A. How the Rb tumor suppressor structure and function was revealed by the study of Adenovirus and SV40. Virology 2009, 384, 274–284. [Google Scholar] [CrossRef] [PubMed] [Green Version]
- Rosen, S.T.; Gould, V.E.; Salwen, H.R.; Herst, C.V.; Le Beau, M.M.; Lee, I.; Bauer, K.; Marder, R.J.; Andersen, R.; Kies, M.S.; et al. Establishment and characterization of a neuroendocrine skin carcinoma cell line. Lab. Investig. 1987, 56, 302–312. [Google Scholar]
- Scherer, W.F.; Syverton, J.T.; Gey, G.O. Studies on the propagation in vitro of poliomyelitis viruses. IV. Viral multiplication in a stable strain of human malignant epithelial cells (strain HeLa) derived from an epidermoid carcinoma of the cervix. J. Exp. Med. 1953, 97, 695–710. [Google Scholar] [CrossRef] [Green Version]
- DuBridge, R.B.; Tang, P.; Hsia, H.C.; Leong, P.M.; Miller, J.H.; Calos, M.P. Analysis of mutation in human cells by using an Epstein-Barr virus shuttle system. Mol. Cell. Biol. 1987, 7, 379–387. [Google Scholar] [CrossRef]
- Houben, R.; Adam, C.; Baeurle, A.; Hesbacher, S.; Grimm, J.; Angermeyer, S.; Henzel, K.; Hauser, S.; Elling, R.; Brocker, E.B.; et al. An intact retinoblastoma protein-binding site in Merkel cell polyomavirus large T antigen is required for promoting growth of Merkel cell carcinoma cells. Int. J. Cancer 2012, 130, 847–856. [Google Scholar] [CrossRef]
- Henley, S.A.; Dick, F.A. The retinoblastoma family of proteins and their regulatory functions in the mammalian cell division cycle. Cell Div. 2012, 7, 10. [Google Scholar] [CrossRef] [Green Version]
- Dimova, D.K.; Dyson, N.J. The E2F transcriptional network: Old acquaintances with new faces. Oncogene 2005, 24, 2810–2826. [Google Scholar] [CrossRef] [Green Version]
- Eguchi, T.; Takaki, T.; Itadani, H.; Kotani, H. RB silencing compromises the DNA damage-induced G2/M checkpoint and causes deregulated expression of the ECT2 oncogene. Oncogene 2007, 26, 509–520. [Google Scholar] [CrossRef] [PubMed] [Green Version]
- Flatt, P.M.; Tang, L.J.; Scatena, C.D.; Szak, S.T.; Pietenpol, J.A. p53 regulation of G(2) checkpoint is retinoblastoma protein dependent. Mol. Cell. Biol. 2000, 20, 4210–4223. [Google Scholar] [CrossRef] [PubMed] [Green Version]
- Niculescu, A.B., 3rd; Chen, X.; Smeets, M.; Hengst, L.; Prives, C.; Reed, S.I. Effects of p21(Cip1/Waf1) at both the G1/S and the G2/M cell cycle transitions: pRb is a critical determinant in blocking DNA replication and in preventing endoreduplication. Mol. Cell. Biol. 1998, 18, 629–643. [Google Scholar] [CrossRef] [PubMed] [Green Version]
- Dyson, N.J. RB1: A prototype tumor suppressor and an enigma. Genes Dev. 2016, 30, 1492–1502. [Google Scholar] [CrossRef] [Green Version]
- Chen, X.; Zaro, J.L.; Shen, W.C. Fusion protein linkers: Property, design and functionality. Adv. Drug Deliv. Rev. 2013, 65, 1357–1369. [Google Scholar] [CrossRef] [Green Version]
- Sakaue-Sawano, A.; Kurokawa, H.; Morimura, T.; Hanyu, A.; Hama, H.; Osawa, H.; Kashiwagi, S.; Fukami, K.; Miyata, T.; Miyoshi, H.; et al. Visualizing spatiotemporal dynamics of multicellular cell-cycle progression. Cell 2008, 132, 487–498. [Google Scholar] [CrossRef] [Green Version]
- Bradford, J.A.; Whitney, P.; Huang, T.; Pinson, P.; Cheung, C.-Y.; Yue, S.; Godfrey, W.L. Novel Vybrant® DyeCycle ™ Stains Provide Cell Cycle Analysis in Live Cells Using Flow Cytometry with Violet, Blue, and Green Excitation. Blood 2006, 108, 4234. [Google Scholar] [CrossRef]
- Houben, R.; Angermeyer, S.; Haferkamp, S.; Aue, A.; Goebeler, M.; Schrama, D.; Hesbacher, S. Characterization of functional domains in the Merkel cell polyoma virus Large T antigen. Int. J. Cancer. J. Int. Du Cancer 2015, 136, E290–E300. [Google Scholar] [CrossRef]
- Ludlow, J.W.; DeCaprio, J.A.; Huang, C.M.; Lee, W.H.; Paucha, E.; Livingston, D.M. SV40 large T antigen binds preferentially to an underphosphorylated member of the retinoblastoma susceptibility gene product family. Cell 1989, 56, 57–65. [Google Scholar] [CrossRef]
- DeCaprio, J.A.; Ludlow, J.W.; Lynch, D.; Furukawa, Y.; Griffin, J.; Piwnica-Worms, H.; Huang, C.M.; Livingston, D.M. The product of the retinoblastoma susceptibility gene has properties of a cell cycle regulatory element. Cell 1989, 58, 1085–1095. [Google Scholar] [CrossRef]
Publisher’s Note: MDPI stays neutral with regard to jurisdictional claims in published maps and institutional affiliations. |
© 2020 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (http://creativecommons.org/licenses/by/4.0/).
Share and Cite
Houben, R.; Ebert, M.; Hesbacher, S.; Kervarrec, T.; Schrama, D. Merkel Cell Polyomavirus Large T Antigen is Dispensable in G2 and M-Phase to Promote Proliferation of Merkel Cell Carcinoma Cells. Viruses 2020, 12, 1162. https://doi.org/10.3390/v12101162
Houben R, Ebert M, Hesbacher S, Kervarrec T, Schrama D. Merkel Cell Polyomavirus Large T Antigen is Dispensable in G2 and M-Phase to Promote Proliferation of Merkel Cell Carcinoma Cells. Viruses. 2020; 12(10):1162. https://doi.org/10.3390/v12101162
Chicago/Turabian StyleHouben, Roland, Marlies Ebert, Sonja Hesbacher, Thibault Kervarrec, and David Schrama. 2020. "Merkel Cell Polyomavirus Large T Antigen is Dispensable in G2 and M-Phase to Promote Proliferation of Merkel Cell Carcinoma Cells" Viruses 12, no. 10: 1162. https://doi.org/10.3390/v12101162
APA StyleHouben, R., Ebert, M., Hesbacher, S., Kervarrec, T., & Schrama, D. (2020). Merkel Cell Polyomavirus Large T Antigen is Dispensable in G2 and M-Phase to Promote Proliferation of Merkel Cell Carcinoma Cells. Viruses, 12(10), 1162. https://doi.org/10.3390/v12101162