The role of Merkel cell polyomavirus in Merkel cell carcinoma
Merkel cell carcinoma is a skin cancer with a high rate of mortality. Factors that increase the risk for developing MCC include excessive exposure to sunlight, advanced age and an immunocompromised state. Recognition that immunodeficiency increased the risk for developing Merkel cell carcinoma prompted a focused search for pathogens and the discovery of Merkel cell polyomavirus. In at least 80% of all Merkel cell carcinomas, the Merkel cell polyomavirus small T antigen (ST) is intact, while the large T antigen (LT) is truncated. Given the experience of our laboratory in studying polyomaviruses especially SV40, we initiated studies of Merkel cell polyomavirus LT and ST. We generated monoclonal antibodies specific for LT and ST that had improved specificity and sensitivity for immunohistochemical detection of MCPyV in MCC tumor specimens that indicated that most cases of Merkel cell carcinoma express the viral T antigens. We generated a mouse knock-in model capable of tissue specific expression of the Merkel virus T antigens and demonstrated their oncogenic potential in vivo. We continue to use advanced proteomics, genomics and bioinformatic approaches to identify the tumorigenic properties of the Merkel cell polyomavirus T antigens in patient samples, mouse models and cell lines.
The DREAM (DP, RB-related, E2F and MuvB) complex
Our laboratory demonstrated that the Retinoblastoma-related proteins p130 and p107 provide tumor suppressor activities. To determine how p130 contributed to growth suppression, we performed a large-scale immunoprecipitation followed by mass spectrometric identification of associated proteins. We identified an 8-protein complex that we termed DREAM based on its similarity to complexes previously identified by genetic and biochemical studies in C. elegans and D. melanogaster. We determined that the mammalian DREAM complex bound specifically to the promoters of all cell cycle regulated genes and repressed their expression during cellular quiescence. We determined that the DYRK1A kinase was required for assembly of the DREAM complex during quiescence. We also found that the DREAM complex underwent a metamorphosis during cell cycle progression with the MuvB component being released from p130, E2F4 and DP1 during the G1 phase of the cell cycle and subsequently binding to B-MYB (MYBL2) and FOXM1 to specifically activate expression of several hundred genes required for progression during G2 and M phase. Our work demonstrated that the DREAM complex serves as a master coordinator of cell cycle gene expression. We continue to study the DREAM complex to understand how cell cycle dependent gene expression is controlled during the major transition phases of the cell cycle including G0, G1/S and G2/M.
Identification of human disease genes by study of Viral-Host cell interactions
Our laboratory identified CUL7, CUL9, FBXW8, GLMN and FAM111A as specific interacting proteins with SV40 LT and generated knockout mouse models for Cul7, Fbxw8, Cul9 and Glmn genes. We demonstrated that knockout of Cul7 or Fbxw8 led to severe growth retardation that was recognized by an independent genetic study that determined that homozygous mutations in CUL7 were responsible for the human 3M short stature syndrome. We demonstrated that GLMN binds directly to RBX1 and inhibits the ubiquitin ligase activity of cullin RING ligases and revealed the molecular basis for Glomuvenous malformation, a human hereditary vascular malformation syndrome. Most recently, we identified FAM111A as an SV40 host range restriction factor. Remarkably, mutations in FAM111A have recently been described in the short stature Kenny-Caffey and osteocraniostenosis syndromes. These studies of viral-host cell protein interactions have led to the molecular characterization of genes that are mutated in a variety of human diseases and suggest that LT targets cellular and organismal growth-promoting activities. Our laboratory focuses on deciphering the molecular functions of these viral-targeted host genes. We have performed a large-scale viral interactome and transcriptome screen that has fueled several new research projects and are actively pursuing new viral protein interactions with other disease related genes in addition to FAM111A and GLMN.