My laboratory explores the molecular mechanisms of replication of positive-strand RNA animal viruses. Positive-strand RNA viruses are interesting because they replicate exclusively via RNA intermediates. Considering the "RNA World" view of the evolution of life, RNA viruses represent modern day organisms with evolutionarily ancient replication strategies.
Positive-strand viral RNA serves two important functions within the cytoplasm of infected host cells:
(1) as mRNA for the expression of the viral proteins and,
(2) as the template for negative-strand RNA synthesis.
Viral RNA cannot simultaneously serve as a mRNA and as a template for negative-strand RNA synthesis due to the 5' to 3' movement of translating ribosomes and the 3' to 5' movement of replicase during negative-strand RNA synthesis. We study two viruses in great detail: poliovirus and hepatitis C virus. We hypothesize that direct interactions occur between the 5'- and 3'-terminal non-translated regions of viral RNA to regulate the transformation of viral mRNA into a template for viral negative-strand RNA synthesis. Poliovirus RNA, as mRNA, first becomes part of a messenger ribonucleoprotein (mRNP) complex with communication between the 5'- and 3'-termini mediated by the cellular translation machinery (eIF4G, poly(A) binding protein, etc.). Following viral protein synthesis, the viral mRNP complex must transform into a preinitiation RNA replication complex to allow for the initiation of viral negative-strand RNA synthesis. The 5'-terminal ribonucleoprotein complex of poliovirus containing viral protein 3CD mediates, in part, the initiation of viral negative-strand RNA synthesis at the 3'-terminus of poliovirus RNA. We propose that this model of viral RNA replication, emphasizing communication between the 5'- and 3'-termini of the viral RNA, is broadly applicable to all positive-strand RNA animal viruses.
In another series of experiments, we discovered that hepatitis C virus RNA is detected and destroyed by an interferon-regulated antiviral pathway present in the cytoplasm of cells; the 2'-5' oligoadenylate synthetase/ribonuclease L pathway. Ribonuclease L cleaves viral RNA at single-stranded UA and UU dinucleotides. Relatively interferon-resistant genotype 1 hepatitis C viruses have fewer ribonuclease L cleavage sites than more interferon-sensitive genotype 2 and 3 viruses. These discoveries may help to explain the clinical outcome of interferon therapy in hepatitis C virus-infected patients where patients infected with genotype 1 viruses are cured less frequently by interferon therapy than patients infected with genotype 2 or 3 viruses.