Modulation of the expression of poliovirus proteins in reticulocyte lysates.
The translation of poliovirus RNA into specific viral proteins in mRNA-dependent reticulocyte lysates (MDLs) was found to be highly dependent on individual lysate preparations. Under optimal conditions, the first polypeptide detected was always P3-1b (formerly NCVP 1b), the product of the 3' portion of the poliovirus genome; the formation of P1-1a (formerly NCVP 1a) followed as shown by time-course and pulse-chase experiments. However, some lysates synthesized little or no P1-1a despite their ability to synthesize P3-1b and to translate normally other cellular and viral mRNAs. When an MDL competent in synthesizing P1-1a was diluted ca. twofold, while maintaining optimal concentrations of salts, tRNA, DTT, creatine phosphate, and amino acids, P1-1a formation was virtually eliminated, while the synthesis of P3-1b, presumably as a consequence of a more downstream initiation, was maintained. The synthesis of P1-1a in a diluted MDL was restored, and P3-1b synthesis suppressed, by the addition of a S10 fraction prepared from uninfected or virus-infected HeLa cells. Nuclease treatment and dialysis of the S10 fraction did not inhibit its activity. These findings indicate that individual MDLs either possess limiting quantities of, or occasionally are deficient in, a factor(s) that promotes the utilization of the presumed 5' proximal initiation site (the AUG at nucleotide position 781-783) and that a homologous factor(s) exists in HeLa cells. The implication of these findings for the strategy of poliovirus replication is discussed.
D000818:Animals; D001483:Base Sequence; D002474:Cell-Free System; D005786:Gene Expression Regulation; D006367:HeLa Cells; D010448:Peptide Initiation Factors / Q000494:pharmacology; D017955:Poliovirus / Q000235:genetics / Q000378:metabolism*; D014176:Protein Biosynthesis / Q000187:drug effects; D011498:Protein Precursors / Q000096:biosynthesis; D012333:RNA, Messenger / Q000235:genetics; D012367:RNA, Viral / Q000235:genetics; D011817:Rabbits; D012156:Reticulocytes; D014764:Viral Proteins / Q000096:biosynthesis* / Q000235:genetics
D010448:Peptide Initiation Factors; D011498:Protein Precursors; D012333:RNA, Messenger; D012367:RNA, Viral; D014764:Viral Proteins