ALV INDUCED TUMORIGENESIS: Identifying Clonal Progression of Tumors and Novel Cancer Associated Genes

Avian leukosis virus (ALV) is a simple retrovirus that infects and causes cancer in chickens. The virus integrates into the chicken genome and it can alter the expression of surrounding genes via its strong promoter and enhancer elements. This perturbation of gene expression near ALV integration sites induces a wide range of tumors. Chickens infected with ALV subgroup A mostly develop B-cell lymphomas, while those infected with ALV subgroup J develop myeloid tumors as well as hemangiomas. Since ALV acts as a mutagen by integrating into the chicken genome, it serves as a good insertional mutagenesis tool. We analyzed the preferences of ALV integrations by infecting different cells (CEF, DT40 and HeLa) in tissue culture. We also analyzed integrations in early and late ALV-induced neoplasms. We infected 5- and 10-day old chicken embryos with ALV subgroup A and J to induce tumorigenesis, and then identified viral integration sites using an Illumina high-throughput sequencing platform. This allowed us to determine how the ALV integration pattern is altered due to oncogenic selection during tumorigenesis. We were also able to empirically define the clonal progression of tumors and identify the underlying biological pathways and gene players implicated in oncogenesis. We identified clonally expanded integrations in the MET gene in ALV subgroup A and J induced hemangiomas. We also showed that ALV induces strong overexpression of MET in the tumors that harbor these integrations. MET is a known proto-oncogene, involved in angiogenesis but has not been previously implicated in the development of hemangiomas. We also identified the TERT promoter region as a common ALV integration hotspot in many B-cell lymphomas. This led us to observe the activation of a novel lncRNA in the TERT promoter region, named TERT antisense promoter-associated (TAPAS) RNA. We showed that ALV drives the overexpression of TAPAS and contributes to the development of lymphomas. Lastly, we reported a novel transcript in the human TERT promoter region, named human TAPAS (hTAPAS) lncRNA. hTAPAS expression inversely correlates with TERT expression in tumors, and its expression is not affected by the TERT promoter mutation. Knock down of hTAPAS results in increased TERT expression, and its ectopic overexpression down regulates TERT expression. With a role in regulating TERT expression, hTAPAS is implicated in regulating TERT homeostasis