Detection of Merkel Cell Polyomavirus (MCPyV) DNA and Transcripts in Merkel Cell Carcinoma (MCC)

Abstract

Merkel cell polyomavirus (MCPyV) is the etiological agent of the majority of Merkel cell carcinoma (MCC): a rare skin tumor. To improve our understanding of the role of MCPyV in MCCs, the detection and analysis of MCPyV DNA and transcripts were performed on primary tumors and regional lymph nodes from two MCC patients: one metastatic and one non-metastatic. MCPyV-DNA was searched by a quantitative polymerase chain reaction (qPCR), followed by the amplification of a Large T Antigen (LTAg), Viral Protein 1 (VP1) and Non-Coding Control Region (NCCR). LTAg and VP1 transcripts were investigated by reverse-transcription PCR (RT-PCR). Viral integration was also studied, and full-length LTAg sequencing was performed. qPCR revealed that the primary tumor of both patients and the lymph node of one patient was positive for the small t-antigen, with an average value of 7.0 × 102 copies/µg. The same samples harbored LTAg, NCCR and VP1 DNA. Sequencing results showed truncated LTAg with the conserved retinoblastoma (Rb) protein binding motif and VP1 and NCCR sequences identical to the MCC350 strain. RT-PCR detected LTAg but not VP1 transcripts. The MCPyV genome was integrated into the primary tumor of both patients. The results confirmed the connection between MCPyV and MCC, assuming integration, LTAg truncation and Rb sequestration as key players in MCPyV-mediated oncogenesis.

Merkel Cell Polyomavirus in the Context of Oral Squamous Cell Carcinoma and Oral Potentially Malignant Disorders

“…However, since the detection and quantitation of viral DNA as well as the characterization of NCCR and VP1 regions are not sufficient to define the MCPyV role in oral malignancies, further analyses have been performed. In spite of what has been reported for virus-positive MCCs, where viral replication is hampered, late genes are not expressed [39,48], and transcript analysis revealed both LT and VP1 gene expression in all virus-positive samples, supporting viral replication activity in our samples.…”

“…To date, several studies reported viral integration in the host genome and the expression of a truncated LT gene with a preserved LXCXE motif, able to bind and sequestrate pRb as key players in MCPyV-mediated oncogenesis [24,39,48]. In our MCPyV-positive samples, viral integration was not reported and a full-length LT, retaining both pRb and p53 binding domains, was observed.…”