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

Abstract

Merkel cell carcinoma (MCC) is the most aggressive skin cancer. Recently, it was demonstrated that human Merkel cell polyomavirus (MCV) is clonally integrated inMerkel cell carcinoma (MCC) is a highly aggressive neuroendocrine skin cancer. Although it is rare, its reported incidence is increasing (19). MCC is associated with UV exposure and affects primarily elderly and immune-suppressed patients (5,11,17,26). The susceptibility of MCC to immune surveillance is similar to that of known virus-induced cancers and suggests that MCC has an infectious trigger (9). Recently, a new human polyomavirus, termed Merkel cell polyomavirus (MCV), was discovered to be clonally integrated into MCC tumor genomes (14). While MCV integration occurs at distinct sites in MCC tumors from different individuals, primary tumors and corresponding metastases have identical integration sites, consistent with the occurrence of MCV infection and integration prior to clonal expansion and metastasis (14,37). A number of studies have confirmed that MCV is present in 69 to 85% of MCC tumors collected from Europe and the United States (4, 15, 21, 41). Surveys of control non-MCC skin, hematolymphoid, and neuroendocrine tumors are generally negative for MCV, although incidental low-level infection can be detected (4,14,22,33,34,39,42,44).All polyomaviruses encode alternatively spliced large T (LT) and small T (sT) antigen transcripts that share exon 1 of the T-antigen (TA) locus. Additional multiply spliced TA transcripts have been described for different polyomaviruses, including the 17kT and 57kT antigens in simian virus 40 (SV40) and MCV, respectively (40,46). Research on viral proteins encoded by the TA locus has been central to uncovering cell signaling networks important in cancer biology (10, 38). The targeting of cellular proteins, such as retinoblastoma protein (Rb), p53, and protein phosphatase 2A (PP2A), by TAs contributes to polyomavirusinduced cell transformation (for reviews, see references 1 and 2). MCV TAs that are expressed in MCC tumors lack a putative p53 binding domain because of tumor-associated T-antigen deletion mutations (37,40). Other conserved tumor suppressor-targeting motifs, including the Rb binding domain (LXCXE motif), the J domain (HPDK) in LT/57kT, and a putative PP2A interaction domain in sT, remain intact (40).Current data point toward MCV as the infectious cause for most Merkel cell cancers: the virus is associated with MCC tumors and, when present, expresses T antigen in tumor cells but not in healthy surrounding tissues (7,20,39). MCV is specific to MCC and is not detected at significant levels in other cancers or in healthy skin examined to date, despite widespread circulation of MCV among human populations (8,23,29,42). Clonal analysis of MCC tumors also supports the correct temporal relationship for causality; i.e., MCV infection occurs prior to MCC tumor development (18). If MCV is a direct cause of MCC tumorigenesis, it is expected that MCC tumors will require MCV protein expression to maintain the tumor ...

Restricted Protein Phosphatase 2A Targeting by Merkel Cell Polyomavirus Small T Antigen

“…SV40 sT activates Akt (43), a protein serine/threonine kinase, causing increased mTORC1 activation associated with decreased inhibitory PP2A dephosphorylation (67). MCV sT targets an end-effector of this pathway to promote cap-dependent translation but does so through a novel mTOR-independent mechanism (38,39). We find that both viruses disrupt PP2A B56␣ phosphatase activity, suggesting that inhibition of this portion of PP2A activity may be important for polyomavirus survival in cells.…”

“…Merkel cell polyomavirus (MCV) (31) was the first human pathogen discovered by nondirected tissue transcriptome sequencing, an approach called digital transcriptome subtraction (DTS) (32), and it is the cause of most Merkel cell carcinomas (31,(33)(34)(35)(36)(37). Similar to other polyomaviruses, MCV encodes large T (LT) and sT early proteins that are required for MCV-positive Merkel cell carcinoma (MCC) cell survival and proliferation (38,39). Unlike SV40 sT, however, MCV sT is a transforming oncoprotein in immortalized rodent fibroblast cells (38,39) and has unique functions that have not been identified in other polyomavirus small T antigens.…”

“…Similar to other polyomaviruses, MCV encodes large T (LT) and sT early proteins that are required for MCV-positive Merkel cell carcinoma (MCC) cell survival and proliferation (38,39). Unlike SV40 sT, however, MCV sT is a transforming oncoprotein in immortalized rodent fibroblast cells (38,39) and has unique functions that have not been identified in other polyomavirus small T antigens. For example, MCV sT enhances LT-mediated MCV viral genome replication by stabilizing LT through sequestration of Fbw7, an E3 ligase that ubiquitinates LT for rapid turnover (40,41).…”

Protein-mediated viral latency is a novel mechanism for Merkel cell polyomavirus persistence

“…MCV sT protein inhibits MCV LT turnover, at least in part, by sequestering Fbw7 (22), which led us to search for Fbw7 phosphodegron motifs encoded by LT protein. In tumors, in which MCV replication is abrogated by integration and mutation, MCV sT serves as an oncoprotein required for cell proliferation (49); however, in the natural setting of the virus, it is likely to be a replication accessory protein that promotes LT stabilization (22) and genome replication (17). The factors controlling alternative splicing to generate LT or sT mRNAs remain unknown, but this reifies the importance of LT stability in controlling MCV replication.…”

Retinoblastoma gene mutations detected by whole exome sequencing of Merkel cell carcinoma

“…13,14 Emerging data implicate maintenance and expression of the polyomavirus large T antigen in cell cycle dysregulation and the pathogenesis of viral transformation leading to Merkel cell carcinoma. 12,[15][16][17][18][19][20][21] Although the molecular role of Merkel cell polyomavirus is quickly evolving, the overall biology of Merkel cell carcinoma is poorly understood. Moreover, the presence of polyomavirus alone is not sufficient for carcinogenesis, namely in those Merkel cell carcinoma cases considered as polyomavirusnegative.…”