GENETIC-ANALYSIS OF INTERNAL RIBOSOMAL ENTRY SITE ON HEPATITIS-C VIRUS-RNA - IMPLICATION FOR INVOLVEMENT OF THE HIGHLY ORDERED STRUCTURE AND CELL-TYPE-SPECIFIC TRANSACTING FACTORS

Hepatitis C virus (HCV) carries an internal ribosomal entry site (IRES ) within the 5' portion of the RNA. To identify structures that influe nce efficiency of the translation initiation, relative activities of m odified IRESs were examined by using engineered bicistronic mRNAs, bet ween the two cistrons of which various mutant IRESs were inserted. An [RES derived from genotype 2b is at least two times more efficient tha n one from genotype Ib in cultured cells. Activity ratios of genotype 2b IRES to Ib IRES differ in magnification among cultured cells, sugge sting the difference in assortment of IRES-related host factors among individual cell types. Recombinant IRESs between the genotypes show si milar or higher activities compared with 2b IRES in cell-free systems and show intermediate activities in cultured cells. Patterns of relati ve activities of those IRESs indicate that the IRES activity is not re gulated by defined structure(s), although a cluster of different nucle otides is observed in the genome region of nucleotides 176-224 between the two alleles. The results suggest that a highly ordered structure formed by the entire 5' portion of the RNA is important for the IRES a ctivity. The 5' border of HCV IRES was examined by using a series of d eletion RNAs in Various systems. The results strongly suggest that the border resides between nucleotide positions 28 and 45. Patterns of re lative activities of the deletion IRESs differ in translation systems or cell types. These results imply that interactions of HCV RNA with t he related transacting factor(s) may differ in the translation systems or cell types. (C) 1997 Academic Press.