Probing the potential origin of virus-driven merkel cell carcinoma using a novel mouse model

Summary: Over 80% of Merkel cell carcinomas (MCCs) carry integrated Merkel cell polyomavirus (MCPyV), leading to expression of oncogenic small T (sT) and truncated large T (tLT) viral antigens which drive tumorigenesis. The MCPyV non-coding control region (NCCR) contains DNA regulatory elements that drive expression of sT and tLT viral oncogenes in MCC tumor cells, and presumably also in skin cells that give rise to MCC. To map NCCR-driven gene expression <i>in vivo</i> we generated mice carrying a randomly integrated <i>NCCR-RFP</i> transgene with a red fluorescent protein reporter molecule to identify cells in which the MCPyV viral promoter is active. <i>NCCR-RFP</i> mice exhibited RFP in rare dermal cells, but these were infrequent, and RFP expression was weak. To improve our ability to detect and manipulate NCCR+ cells, we generated a modified construct containing i) the MCPyV NCCR, ii) a nuclear-localized RFP (nRFP), and iii) an rtTA3 cDNA for inducible expression of tetO-driven transgenes. The <i>NCCR-nRFP-rtTA3</i> construct was inserted into the ROSA26 safe harbor genomic locus to mimic the open chromatin landscape associated with integrated NCCRs in human MCC tumor cells. In contrast to <i>NCCR-RFP</i> mice, <i>R26-NCCR-nRFP-rtTA3</i> mice consistently exhibited nRFP expression in a subset of epidermal and dermal cells, and most strikingly and consistently, in hair follicle stem cells localized to the bulge region. NCCR activity/nRFP expression in follicle stem cells is particularly interesting since early-stage tumors in our MCC mouse models are first detected in the bulge region. These findings point to follicle stem cells as potential cells of origin for MCC, addressing a long-standing question that has been difficult to tackle using other approaches. In addition, our data show that targeted insertion of promoters into the ROSA26 locus can override the confounding effects of flanking sequences on faithful expression of randomly-integrated transgenes, providing a more reliable way to map promoter activity. Li-Jyun Syu¹, Pei-Wei Huang¹, Shreya Mishra¹, Dawn Wilbert¹, Honglai Zhang¹, Zach Freeman¹, Monique Verhaegen¹, Andrzej Dlugosz¹ 1. University of Michigan, Ann Arbor, MI, United States. UV Biology/Injury and Non-melanoma Cancers