Associate Professor
Ph.D., University of Alabama at Birmingham, 1998
Post-doctoral Fellow, Washington University, St. Louis, 1998-2002

Program Affiliation: Molecular Biology & Genetics | Microbiology

Research Groups Affiliation: Biochemistry | Genomics & Bioinformatics | Microbiology

Phone: 812-856-0652
Fax: 812-855-6705
Email Richard

Work in my laboratory focuses on the role of cis-acting RNA elements in viral genomes and how these elements interact with viral and host trans-acting factors to regulate genome function. For these studies we use alphaviruses as they are relatively simple RNA viruses that are readily amenable to genetic manipulation. Sindbis virus is predominantly used in the lab, however work on Venezuelan equine encephalitis virus is also ongoing.

Cis-acting signals for replication. Alphavirus genomic RNA replication is a two-step process; initially the plus-sense genomic RNA is copied into a full-length minus-strand intermediate, this minus-strand RNA then functions as a template for the production of multiple copies of the genome RNA. The replication enzymes required for these two processes are different while being derived from the same source. Upon infection the genome RNA functions as a messenger RNA and is translated to produce a polyprotein. A partially processed form of this polyprotein functions as the replicase for minus-strand RNA synthesis while a more completely processed form functions as the replicase for plus-strand synthesis. One of the major tools used in my laboratory is an in vitro minus-strand synthesis system. This system allows us to separate minus-strand RNA synthesis from translation or plus-strand synthesis, thus facilitating the analysis of mutational effects on this process alone. Using this method we have been able to identify cis-acting elements at both the 5' and 3' end of the genome that influence RNA replication. Additionally detailed molecular work has defined binding sites for the viral RNA polymerase in the 3’ region of the genome, and defined promoter functions within the genome. Future work will focus on how these cis-acting elements communicate with one another to coordinate the various functions carried out by the genome.

Replicase composition and assembly. An active are of research in the lab focuses on determining the molecular interactions required to form a functional genome replication complex. Which viral proteins contact each other? What host factors are part of this complex? Do host factors contact specific cis-acting elements? Using a sub cellular fractionation, proteomics, and immunoprecipitation techniques we have been able to identify multiple host proteins enriched in fractions containing active viral replication complexes. Initial analyses indicate some of these proteins are important for efficient virus replication. Additionally we have been able to establish specific interactions between viral components of the replication complex and continued work will aim to verify the significance of these interactions.

A. J. Burnham, L. Gong, and R. W. Hardy. 2007. Heterogeneous nuclear ribonuclear protein K interacts with Sindbis virus nonstructural proteins and viral subgenomic mRNA. Virology. In press.

M. A. Thal, B. R. Wasik, J. Posto, and R. W. Hardy. 2007. Template requirements for recognition and copying by Sindbis virus RNA-dependent RNA polymerase. Virology. 358: p.221-232.

S. Tomar, R. W. Hardy, J. Smith, and R. J. Kuhn. 2006. Catalytic core of alphavirus nonstructural protein, nsP4 possesses a terminal adenyly transferase activity. J. Virol. 80(20): p.9962-9969.

R. W. Hardy. 2005. The role of the 3’ terminus of the Sindbis virus genome in minus-strand initiation site selection. Virology 345: p.520-531.

R. W. Hardy and C. M. Rice. 2005. Requirements at the 3’ end of the Sindbis virus genome for efficient synthesis of minus-strand RNA. J. Virol. 79 (8): p. 4630-4639.

R. Gorchakov, R. Hardy, C.M. Rice, and I. Frolov, 2004. Selection of functional 5' cis-acting elements promoting efficient Sindbis virus genome replication. J. Virol, 78(1): p. 61-75.

R. W. Hardy, J. Marcotrigiano, K. Blight, J. Majors, and C. M. Rice. 2003. Hepatitis C virus RNA synthesis in a cell-free system isolated from replicon-containing hepatoma cells. J. Virol. 77 (3). 2029-2037.

Frolov*, R. W. Hardy*, C. M. Rice. 2001. cis-acting RNA elements at the 5’ end of Sindbis virus genome RNA regulate minus and plus strand RNA synthesis. RNA. 7. 1638-1651. * co-first authors