Cis and trans-acting factors that regulate hepatitis A virus translation

Picornaviruses have been shown to translate their genomes by binding ribosomes to internal sequences withing the 5'-noncoding region (5'NCR) of the viral RNA genome. The internal ribosome binding sites (RES) has been mapped to the 5'NCR of all picornaviruses studies so far. The experiments presented in this dissertation have been designed to identify the cis- and trans- acting factors that effect hepatitis A virus (HAV) translation. Studies provided in chapter II describe the minimal sequence required for cap-independent translation of HAV monocistronic RNA in vitro. The region between nucleotides 347 and 734 is required for translation in this system. By utilizing bicistronic RNAs containing the HAV 5'NCR, Chapter II further defines those sequences by determining that HAV also translated by internal initiation and contains an IRES element that exists between nucleotides 45 and 734. Comparative analysis using capped and uncapped bi- and monocistronic RNAs showed that RNA synthesized with 5'-terminal methylated cap group allows for more efficient translation that RNA containing only the HAV IRES. Studies in Chapter IV describe the existence of a trans-acting activity fournd in liver cells that stimulates HAV translation in vitro. This activity appears to be specific to HAV RNA and could not be found in other cell of tissue culture origin or kidney, heart and brain extracts derived from the mouse. In combination, the studies presented in this dissertation may help elucidate the possible causes for the slow-growth and pathogenicity exhibited by HAV.