AN ELABORATED PSEUDOKNOT IS REQUIRED FOR HIGH-FREQUENCY FRAMESHIFTINGDURING TRANSLATION OF HCV 229E POLYMERASE MESSENGER-RNA

The RNA polymerase gene (gene 1) of the human coronavirus 229E is appr oximately 20 kb in length and is located at the 5' end of the positive -strand genomic RNA. The coding sequence of gene 1 is divided into two large open reading frames, ORF1a and ORF1b, that overlap by 43 nucleo tides. In the region of the ORF1a/ORF1b overlap, the genomic RNA displ ays two elements that are known to mediate (-1) ribosomal frameshiftin g. These are the slippery sequence, UUUAAAC, and a 3' pseudoknot struc ture. By introducing site-specific mutations into synthetic mRNAs, we have analysed the predicted structure of the HCV 229E pseudoknot and s hown that besides the well-known stem structures, S1 and S2, a third s tem structure, S3, is required for a high frequency of frameshifting. The requirement for an S3 stem is independent of the length of loop 2.