scholarly journals Merkel cell polyomavirus T antigens promote cell proliferation and inflammatory cytokine gene expression

2015 ◽  
Vol 96 (12) ◽  
pp. 3532-3544 ◽  
Author(s):  
Kathleen F. Richards ◽  
Anna Guastafierro ◽  
Masahiro Shuda ◽  
Tuna Toptan ◽  
Patrick S. Moore ◽  
...  
Keyword(s):  
Gene Expression ◽  
Cell Cycle ◽  
Cellular Proliferation ◽  
Human Fibroblasts ◽  
T Antigen ◽  
Merkel Cell Polyomavirus ◽  
Cytokine Gene Expression ◽  
Merkel Cell ◽  
T Antigens ◽  
Gene Expression Alterations

Merkel cell polyomavirus (MCV) is clonally integrated in over 80 % of Merkel cell carcinomas and mediates tumour development through the expression of viral oncoproteins, the large T (LT) and small T antigens (sT). Viral integration is associated with signature mutations in the T-antigen locus that result in deletions of C-terminal replicative functions of the LT antigen. Despite these truncations, the LT LXCXE retinoblastoma (Rb) pocket protein family binding domain is retained, and the entire sT isoform is maintained intact. To investigate the ability of MCV oncoproteins to regulate host gene expression, we performed microarray analysis on cells stably expressing tumour-derived LT, tumour-derived LT along with sT, and tumour-derived LT with a mutated Rb interaction domain. Gene expression alterations in the presence of tumour-derived LT could be classified into three main groups: genes that are involved in the cell cycle (specifically the G1/S transition), genes involved in DNA replication and genes involved in cellular movement. The LXCXE mutant LT largely reversed gene expression alterations detected with the WT tumour-derived LT, while co-expression of sT did not significantly affect these patterns of gene expression. LXCXE-dependent upregulation of cyclin E and CDK2 correlated with increased proliferation in tumour-derived LT-expressing cells. Tumour-derived LT and tumour-derived LT plus sT increased expression of multiple cytokines and chemokines, which resulted in elevated levels of secreted IL-8. We concluded that, in human fibroblasts, the LXCXE motif of tumour-derived LT enhances cellular proliferation and upregulates cell cycle and immune signalling gene transcription.

scholarly journals Merkel Cell Polyomavirus Downregulates N-myc Downstream-Regulated Gene 1, Leading to Cellular Proliferation and Migration

2019 ◽  
Vol 94 (3) ◽  
Author(s):  
Purnima Gupta ◽  
Naveed Shahzad ◽  
Alexis Harold ◽  
Masahiro Shuda ◽  
Assunta Venuti ◽  
...  
Keyword(s):  
Gene Expression ◽  
Cell Carcinoma ◽  
Cellular Proliferation ◽  
Expression Profiles ◽  
Gene Expression Profiles ◽  
T Antigen ◽  
Merkel Cell Polyomavirus ◽  
Early Gene ◽  
Merkel Cell ◽  
Gene Expressing

ABSTRACT Merkel cell polyomavirus (MCPyV) is the first human polyomavirus etiologically associated with Merkel cell carcinoma (MCC), a rare and aggressive form of skin cancer. Similar to other polyomaviruses, MCPyV encodes early T antigen genes, viral oncogenes required for MCC tumor growth. To identify the unique oncogenic properties of MCPyV, we analyzed the gene expression profiles in human spontaneously immortalized keratinocytes (NIKs) expressing the early genes from six distinct human polyomaviruses (PyVs), including MCPyV. A comparison of the gene expression profiles revealed 28 genes specifically deregulated by MCPyV. In particular, the MCPyV early gene downregulated the expression of the tumor suppressor gene N-myc downstream-regulated gene 1 (NDRG1) in MCPyV gene-expressing NIKs and hTERT-MCPyV gene-expressing human keratinocytes (HK) compared to their expression in the controls. In MCPyV-positive MCC cells, the expression of NDRG1 was downregulated by the MCPyV early gene, as T antigen knockdown rescued the level of NDRG1. In addition, NDRG1 overexpression in hTERT-MCPyV gene-expressing HK or MCC cells resulted in a decrease in the number of cells in S phase and cell proliferation inhibition. Moreover, a decrease in wound healing capacity in hTERT-MCPyV gene-expressing HK was observed. Further analysis revealed that NDRG1 exerts its biological effect in Merkel cell lines by regulating the expression of the cyclin-dependent kinase 2 (CDK2) and cyclin D1 proteins. Overall, NDRG1 plays an important role in MCPyV-induced cellular proliferation. IMPORTANCE Merkel cell carcinoma was first described in 1972 as a neuroendocrine tumor of skin, most cases of which were reported in 2008 to be caused by a PyV named Merkel cell polyomavirus (MCPyV), the first PyV linked to human cancer. Thereafter, numerous studies have been conducted to understand the etiology of this virus-induced carcinogenesis. However, it is still a new field, and much work is needed to understand the molecular pathogenesis of MCC. In the current work, we sought to identify the host genes specifically deregulated by MCPyV, as opposed to other PyVs, in order to better understand the relevance of the genes analyzed on the biological impact and progression of the disease. These findings open newer avenues for targeted drug therapies, thereby providing hope for the management of patients suffering from this highly aggressive cancer.

scholarly journals Defective DNA repair and cell cycle arrest in cells expressing Merkel cell polyomavirus T antigen

2012 ◽  
Vol 131 (8) ◽  
pp. 1818-1827 ◽  
Author(s):  
Stephanie K. Demetriou ◽  
Katherine Ona-Vu ◽  
Erin M. Sullivan ◽  
Tiffany K. Dong ◽  
Shu-Wei Hsu ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Dna Repair ◽  
Cell Cycle Arrest ◽  
T Antigen ◽  
Merkel Cell Polyomavirus ◽  
Merkel Cell ◽  
Cycle Arrest ◽  
Defective Dna Repair ◽  
In Cells

scholarly journals The Merkel Cell Polyomavirus T Antigens Function as Tumor Promoters in Murine Skin

Cancers ◽  
2021 ◽  
Vol 13 (2) ◽  
pp. 222
Author(s):  
Megan E. Spurgeon ◽  
Amy Liem ◽  
Darya Buehler ◽  
Jingwei Cheng ◽  
James A. DeCaprio ◽  
...  
Keyword(s):  
Transgenic Mice ◽  
Constitutive Expression ◽  
T Antigen ◽  
Tumor Formation ◽  
Merkel Cell Polyomavirus ◽  
Tumor Promoter ◽  
Tumor Promoters ◽  
Merkel Cell ◽  
T Antigens

Merkel cell polyomavirus (MCPyV) causes the majority of human Merkel cell carcinomas (MCC), a rare but highly aggressive form of skin cancer. We recently reported that constitutive expression of MCC tumor-derived MCPyV tumor (T) antigens in the skin of transgenic mice leads to hyperplasia, increased proliferation, and spontaneous epithelial tumor development. We sought to evaluate how the MCPyV T antigens contribute to tumor formation in vivo using a classical, multi-stage model for squamous cell carcinoma development. In this model, two chemical carcinogens, DMBA and TPA, contribute to two distinct phases of carcinogenesis—initiation and promotion, respectively—that are required for tumors to develop. By treating the MCPyV transgenic mice with each chemical carcinogen, we determined how the viral oncogenes contributed to carcinogenesis. We observed that the MCPyV T antigens synergized with the tumor initiator DMBA, but not with the tumor promoter TPA, cause tumors. Therefore, the MCPyV tumor antigens function primarily as tumor promoters, similar to that seen with human papillomavirus (HPV) oncoproteins. These studies provide insight into the role of MCPyV T antigen expression in tumor formation in vivo and contribute to our understanding of how MCPyV may function as a human DNA tumor virus.

scholarly journals Merkel Cell Polyomavirus-Infected Merkel Cell Carcinoma Cells Require Expression of Viral T Antigens

2010 ◽  
Vol 84 (14) ◽  
pp. 7064-7072 ◽  
Author(s):  
Roland Houben ◽  
Masahiro Shuda ◽  
Rita Weinkam ◽  
David Schrama ◽  
Huichen Feng ◽  
...  
Keyword(s):  
Cell Carcinoma ◽  
Cell Lines ◽  
Merkel Cell Carcinoma ◽  
Annexin V ◽  
T Antigen ◽  
Merkel Cell Polyomavirus ◽  
Merkel Cell ◽  
T Antigens ◽  
Negative Cell ◽  
Exon 1

ABSTRACT Merkel cell carcinoma (MCC) is the most aggressive skin cancer. Recently, it was demonstrated that human Merkel cell polyomavirus (MCV) is clonally integrated in ∼80% of MCC tumors. However, direct evidence for whether oncogenic viral proteins are needed for the maintenance of MCC cells is still missing. To address this question, we knocked down MCV T-antigen (TA) expression in MCV-positive MCC cell lines using three different short hairpin RNA (shRNA)-expressing vectors targeting exon 1 of the TAs. The MCC cell lines used include three newly generated MCV-infected cell lines and one MCV-negative cell line from MCC tumors. Notably, all MCV-positive MCC cell lines underwent growth arrest and/or cell death upon TA knockdown, whereas the proliferation of MCV-negative cell lines remained unaffected. Despite an increase in the number of annexin V-positive, 7-amino-actinomycin D (7-AAD)-negative cells upon TA knockdown, activation of caspases or changes in the expression and phosphorylation of Bcl-2 family members were not consistently detected after TA suppression. Our study provides the first direct experimental evidence that TA expression is necessary for the maintenance of MCV-positive MCC and that MCV is the infectious cause of MCV-positive MCC.

scholarly journals Merkel cell polyomavirus infection of animal dermal fibroblasts

2017 ◽  
pp. JVI.01610-17 ◽  
Author(s):  
Wei Liu ◽  
Nathan A. Krump ◽  
Margo MacDonald ◽  
Jianxin You
Keyword(s):  
Gene Expression ◽  
Animal Models ◽  
Tree Shrew ◽  
T Antigen ◽  
Dermal Fibroblasts ◽  
Merkel Cell Polyomavirus ◽  
Early Gene ◽  
Merkel Cell ◽  
Woolly Monkey ◽  
Chimeric Viruses

Merkel cell polyomavirus (MCPyV) is the first polyomavirus to be associated with human cancer. Mechanistic studies attempting to fully elucidate MCPyV oncogenic mechanisms have been hampered by the lack of animal models for MCPyV infection. In this study, we examined the infectability of MCPyV-GFP pseudovirus, MCPyV recombinant virion, and several MCPyV chimeric viruses in dermal fibroblasts isolated from various model animals, including mouse (Mus musculus), rabbit (Oryctolagus cuniculus), rat (Rattus norvegicus), chimpanzee (Pan troglodytes), rhesus macaque (Macaca mulatta), patas monkey (Erythrocebus patas), common woolly monkey (Lagothrix lagotricha), red-chested mustached tamarin (Saguinus labiatus), and tree shrew (Tupaia Belangeri). We found that MCPyV-GFP pseudovirus is able to enter the dermal fibroblasts of all species tested. Chimpanzee dermal fibroblasts were the only type that supported vigorous MCPyV gene expression and viral replication, and did so to a level beyond that of human dermal fibroblasts. We further demonstrated that both human and chimpanzee dermal fibroblasts produce infectious MCPyV virions that can successfully infect new cells. In addition, rat dermal fibroblasts supported robust MCPyV large T antigen expression after infection with an MCPyV chimeric virus, in which the entire enhancer region of MCPyV early promoter has been replaced with the SV40 analog. Our results suggest that viral transcription and/or replication events represent the major hurdle for MCPyV cross-species transmission. The capacity of rat dermal fibroblasts to support MCPyV early gene expression suggests that the rat is a candidate model organism for studying the viral oncogene function during MCC oncogenic progression.IMPORTANCEMCPyV plays an important role in the development of a highly aggressive form of skin cancer, Merkel cell carcinoma (MCC). With the increasing number of MCC diagnoses, there is a need to better understand the virus and its oncogenic potential. However, studies attempting to fully elucidate MCPyV oncogenic mechanisms have been hampered by the lack of animal models for MCPyV infection. To pinpoint the best candidate for developing an MCPyV infection animal model, we examined the MCPyV infectability of dermal fibroblasts isolated from various established model animals. Of the animal cell types we tested, chimpanzee dermal fibroblasts were the only isolates that support the full MCPyV infectious cycle. To overcome the infection blockade in the other model animals, we constructed chimeric viruses that achieved robust MCPyV entry and oncogene expression in rat fibroblasts. Our results suggest that the rat may serve as anin vivomodel to study MCV oncogenesis.

scholarly journals Merkel Cell Polyomavirus Large T Antigen Disrupts Host Genomic Integrity and Inhibits Cellular Proliferation

2013 ◽  
Vol 87 (16) ◽  
pp. 9173-9188 ◽  
Author(s):  
J. Li ◽  
X. Wang ◽  
J. Diaz ◽  
S. H. Tsang ◽  
C. B. Buck ◽  
...  
Keyword(s):  
Cellular Proliferation ◽  
T Antigen ◽  
Merkel Cell Polyomavirus ◽  
Merkel Cell ◽  
Genomic Integrity ◽  
Large T Antigen

scholarly journals Merkel Cell Polyomavirus Large T Antigen Unique Domain Regulates Its Own Protein Stability and Cell Growth

Viruses ◽  
2020 ◽  
Vol 12 (9) ◽  
pp. 1043
Author(s):  
Nnenna Nwogu ◽  
Luz E. Ortiz ◽  
Hyun Jin Kwun
Keyword(s):  
Cell Proliferation ◽  
Cell Growth ◽  
Viral Replication ◽  
Cellular Proliferation ◽  
E3 Ligase ◽  
T Antigen ◽  
Merkel Cell Polyomavirus ◽  
Merkel Cell ◽  
E3 Ligases ◽  
Unique Domain

Merkel cell polyomavirus (MCV) is the only known human oncogenic virus in the polyomaviridae family and the etiological agent of most Merkel cell carcinomas (MCC). MCC is an aggressive and highly metastatic skin cancer with a propensity for recurrence and poor prognosis. Large tumor antigen (LT), is an essential oncoprotein for MCV transcription, viral replication, and cancer cell proliferation. MCV LT is a short-lived protein that encodes a unique domain: MCV LT unique regions (MURs). These domains consist of phosphorylation sites that interact with multiple E3 ligases, thus limiting LT expression and consequently, viral replication. In this study, we show that MURs are necessary for regulating LT stability via multiple E3 ligase interactions, resulting in cell growth arrest. While expression of wild-type MCV LT induced a decrease in cellular proliferation, deletion of the MUR domains resulted in increased LT stability and cell proliferation. Conversely, addition of MURs to SV40 LT propagated E3 ligase interactions, which in turn, reduced SV40 LT stability and decreased cell growth activity. Our results demonstrate that compared to other human polyomaviruses (HPyVs), MCV LT has evolved to acquire the MUR domains that are essential for MCV LT autoregulation, potentially leading to viral latency and MCC.

scholarly journals The Simian Virus 40 Small-t and Large-T Antigens Jointly Regulate Cell Cycle Reentry in Human Fibroblasts

1999 ◽  
Vol 73 (4) ◽  
pp. 3102-3107 ◽  
Author(s):  
Analía Porrás ◽  
Stéphanie Gaillard ◽  
Kathleen Rundell
Keyword(s):  
Cell Cycle ◽  
Human Fibroblasts ◽  
Simian Virus 40 ◽  
Simian Virus ◽  
T Antigen ◽  
Large T Antigen ◽  
T Antigens ◽  
Cell Cycle Reentry ◽  
Small T Antigen ◽  
Hdf Cells

ABSTRACT Focus formation in human diploid fibroblasts (HDF cells) is known to require both the simian virus 40 (SV40) large-T and small-t antigens. Similarly, both SV40 proteins were required to stimulate confluent, density-arrested HDF cells to reenter the cell cycle. This study used defective recombinant adenoviruses to examine the roles of the individual SV40 proteins in altering specific steps in the cell cycle. Small-t antigen and, to a lesser extent, large-T antigen increased the level of the S phase cyclin cyclin A but without increasing the activity of associated cyclin kinases unless the two SV40 proteins were coexpressed. The absence of kinase activity reflected the presence in density-arrested cells of high levels of the cyclin-dependent kinase inhibitors p21WAF1 and p27KIP1. We report here that expression of SV40 large-T antigen reduced levels of p21WAF1, while expression of small-t antigen was required to decrease p27KIP1. The separate effects of large-T and small-t antigens on these two inhibitors may explain the joint requirement for the two proteins to drive cell cycle reentry of HDF cells and ultimately transform these cells.

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