Merkel Cell Polyomavirus Large T Antigen Has Growth-Promoting and Inhibitory Activities

Abstract

cMerkel cell carcinoma (MCC) is a rare and aggressive form of skin cancer. In at least 80% of all MCC, Merkel cell polyomavirus (MCPyV) DNA has undergone clonal integration into the host cell genome, and most tumors express the MCPyV large and small T antigens. In all cases of MCC reported to date, the integrated MCPyV genome has undergone mutations in the large T antigen. These mutations result in expression of a truncated large T antigen that retains the Rb binding or LXCXE motif but deletes the DNA binding and helicase domains. However, the transforming functions of full-length and truncated MCPyV large T antigen are unknown. We compared the transforming activities of full-length, truncated, and alternatively spliced 57kT forms of MCPyV large T antigen. MCPyV large T antigen could bind to Rb but was unable to bind to p53. Furthermore, MCPyV-truncated large T antigen was more effective than full-length and 57kT large T antigen in promoting the growth of human and mouse fibroblasts. In contrast, expression of the MCPyV large T antigen C-terminal 100 residues could inhibit the growth of several different cell types. These data imply that the deletion of the C terminus of MCPyV large T antigen found in MCC serves not only to disrupt viral replication but also results in the loss of a distinct growth-inhibitory function intrinsic to this region. Merkel cell carcinoma (MCC) is an aggressive skin cancer with an annual incidence of 3 per million in the United States (1). Risk factors for developing MCC include advanced age, prolonged UV exposure, and immunosuppression due to HIV, hematologic malignancy, or solid-organ transplantation (2, 3). Recently, Merkel cell polyomavirus (MCPyV) was discovered to be clonally integrated in at least 80% of MCC, raising the possibility that this pathogen contributes to carcinogenesis (4, 5).MCPyV is a typical polyomavirus with a circular, doublestranded DNA genome containing an early region that expresses large and small T antigens, a late region that encodes 3 viral coat proteins, VP1, VP2, and VP3, and a regulatory region that contains the origin of replication and a bidirectional promoter for the early and late genes. MCPyV was the fifth polyomavirus identified in humans, preceded by BKPyV, JCPyV, KIPyV, and WUPyV (6-9). Since then, 6 additional human polyomaviruses have been discovered, including HPyV6, HPyV7, TSPyV, HPyV9, MWPyV, and STLPyV (10-15). Although JCPyV and BKPyV have been detected in a variety of human cancers (16, 17), Merkel cell polyomavirus is the only polyomavirus DNA clonally integrated in human cancer. Expression of Merkel large and small T antigens can be detected in most MCC specimens (5, 18, 19).The T antigens from several polyomaviruses have oncogenic activity. Notably, the simian virus 40 (SV40) large and small T antigens can transform a variety of rodent and human cells. Expression of SV40 large and small T antigens, together with human telomerase reverse transcriptase and an oncogenic form of H-RAS, can fully transform normal human fibroblasts (20,21)...

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.…”

Phosphorylation of Merkel Cell Polyomavirus Large Tumor Antigen at Serine 816 by ATM Kinase Induces Apoptosis in Host Cells

“…The identification of a tumor with intact full-length LT but a mutated viral origin sequence supports this hypothesis (26). Later studies have since suggested that the C-terminal helicase domain may contain other functions that oppose tumorigenesis (16,24). Our previous work indicated that expression of full-length MCV LT activates a dramatic DNA damage response (DDR) that is antagonistic to tumorigenesis; this activity activates p53 and induces a growthinhibiting phenotype (16).…”

“…Our previous study demonstrated that the C-terminal portion of LT activates p53 and promotes growth inhibition (16). Cheng et al (24) also reported that the C-terminal 100 amino acids of MCV LT have a cell growth-inhibiting effect. The underlying mechanism of these findings was not completely established.…”

“…MCV LT can bind specifically to pRb (3,23). Although there are two potential p53-binding motifs in the MCV LT C-terminal domain, there appears to be no direct interaction between MCV LT and p53 (24,25). Interestingly, the MCV genome is commonly clonally integrated into MCC tumor cell genomes.…”

“…Our previous work indicated that expression of full-length MCV LT activates a dramatic DNA damage response (DDR) that is antagonistic to tumorigenesis; this activity activates p53 and induces a growthinhibiting phenotype (16). Additionally, Cheng et al (24) reported that expression of the C-terminal 100 residues of MCV LT inhibits the growth of several different cell types. These studies support a model in which the C-terminal domain must be deleted in tumor cells to both limit viral replication from the integrated viral genomes and eliminate growth-arresting properties intrinsic to the C-terminal domain of LT. How the MCV C-terminal 100 residues accomplish this growth-arresting function is not clearly understood.…”