Richard J. Frisque Professor of Molecular Virology and Department Head 433 S. Frear Laboratory, University Park, PA 16802 Phone: (814) 863-3523 Fax: (814) 863-7024 E-mail: rjf6@psu.edu B.S. and Ph.D. in medical microbiology, University of Wisconsin Frisque Lab Web Site

Pathogenic and Oncogenic Potential of a Human Polyomavirus

Our laboratory's research interests center on the unique biology of the human polyomavirus, JC virus (JCV). JCV infects most individuals early in life and persists in their body. Most infections are asymptomatic, but in an immunocompromised host, JCV may cause the fatal demyelinating brain disease called progressive multifocal leukoencephalopathy (PML); ~5% of AIDS patients now succumb to this disease. Recently PML has surfaced in multiple sclerosis and Crohn’s disease patients treated with Tysabri, a monoclonal antibody designed to inhibit lymphocyte migration into the tissues. Similar drugs are in clinical trials, elevating concerns that such treatments will lead to other life-threatening events. JCV is also an oncogenic agent, with the ability to induce a wide variety of tumors in rodents and in non-human primates. Recent reports link JCV to certain human cancers.

Our sequence analysis of the JCV genome has revealed that this virus shares approximately 70% homology with the human virus BKV and the monkey virus SV40. Major sequence differences between these viruses occur within the promoter/enhancer signals and the coding region for the multifunctional regulatory protein, T antigen. To determine how these sequence alterations translate into specific biological differences, our initial studies involved genetic approaches; the construction and phenotypic analysis of hybrid polyomaviruses and the use of site-directed mutagenesis techniques to alter cis-acting replication signals and specific functional domains of the T protein. These experiments began to define those sequences that contribute to the restricted lytic and transforming activities of this human pathogen. Particularly interesting were the findings that binding of the JCV T antigen to the viral replication origin and to the cellular "anti-oncogene products", pRB and p53, differs with that of the corresponding SV40 protein. In addition, we have identified 3 truncated forms of the T antigen (T'135, T'136 and T'165) in transformed and lytically-infected cells that arise via a differential splicing mechanism. We are intrigued by the observation that T' proteins exhibit unique functions even though they are closely related at the sequence level. We have shown that JCV mutants lacking the T' proteins exhibit significantly reduced DNA replication activity. To complement our genetic approaches, the JCV T/T' proteins have been overproduced in eukaryotic expression systems to facilitate analyses at the biochemical level. In addition, cell lines expressing individual JCV tumor proteins have been generated. We have found that T antigen and the 3 T' proteins exhibit differential binding to the pRB family of cellular tumor suppressor proteins both in vivo and in vitro.  Furthermore, this binding causes differential release of pRB-bound members of the E2F transcription factor family. Using a ras cooperation assay, we have demonstrated that the JCV early proteins vary in their ability to induce transformation and immortalization of rodent cells. Currently we are attempting to identify the basis for functional differences among the closely related JCV early proteins and to understand how the alternative splicing process, which generates the five JCV early transcripts, is regulated.

Representative Publications:

• R.J. Frisque, B. Bollag, S.K. Tyagarajan and L.H. Kilpatrick. 2003, T’ proteins influence JC virus biology.  J. NeuroVirol. 9: 15-20.
• Ricciardiello L, Baglioni M, Giovannini C, Pariali M, Cenacchi G, Ripalti A, Landini MP, Sawa H, Nagashima K, Frisque RJ, Goel A, Boland CR, Tagono M, Roda E and Bazzoli F. 2003. Induction of chromosomal instability in colonic cells by the human polyomavirus JC virus. Cancer Research 63: 7256-7262.
• Ziegler, K., T. Bui, R.J. Frisque, A. Grandinetti, and V.R. Nerurkar (2004). A rapid in-vitro polyomavirus DNA replication assay. J. Virol. Meth. 122: 123-127.
• Frisque,R.J., C. Hofstetter, and S. Tyagarajan. 2006. Transforming activities of JC virus early proteins. In Polyomaviruses and Human Diseases, (ed. N. Ahsan). Advances in Experimental Medicine and Biology, Vol. 577, pp. 288-309. Eurekah.com and Springer Science+Business Media, New York, NY.
• Verma, S., K. Ziegler, P. Ananthula, J.K.G Co, R.J. Frisque, R. Yanagihara, and V.R. Nerurkar. (2006). JC virus induces altered patterns of cellular gene expression: Interferon inducible genes as major transcriptional targets. Virology 345:457-467.
• Tyagarajan, S.K., and R.J. Frisque. (2006). The stability and function of JC virus large T antigen and T' proteins are altered by mutation of their phosphorylated threonine 125 residue. J. Virol. 80: 2083-2091.
• Bollag, B., Kilpatrick, L.H., S.K. Tyagarajan, M.J. Tevethia, and R.J. Frisque. (2006). JC Virus proteins T'135, T'136 and T'165 interact with cellular regulatory factors and influence viral transformation potential. J. NeuroVirol. 12: 428-442.
• Shiramizu, B., N. Hu, R.J. Frisque, and V.R. Nerurkar. (2007). High prevalence of human polyomavirus JC VP1 gene sequences in pediatric malignancies. Cell. Mol. Biol. 53: 4-12.

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