ANALYSES OF FRAMESHIFTING AT UUU-PYRIMIDINE SITES

Others have recently shown that the UUU phenylalanine codon is highly frameshift-prone in the 3' (rightward) direction at pyrimidine 3' cont exts, Here, several approaches are used to analyze frameshifting at su ch sites, The four permutations of the UUU/C (phenylalanine) and CGG/U (arginine) codon pairs were examined because they vary greatly in the ir expected frameshifting tendencies, Furthermore, these synonymous si tes allow direct tests of the idea that codon usage can control frames hifting. Frameshifting was measured for these dicodons embedded within each of two broader contexts: the Escherichia coli prfB (RF2 gene) pr ogrammed frameshift site and a 'normal' message site, The principal di fference between these contexts is that the programmed frameshift cont ains a purine-rich sequence upstream of the slippery site that can bas e pair with the 3' end of 16 S rRNA (the anti-Shine-Dalgarno) to enhan ce frameshifting, In both contexts frameshift frequencies are highest if the slippery tRNA(Phe) is capable of stable base pairing in the shi fted reading frame, This require ment is less stringent in the RF2 con text, as if the Shine-Dalgarno interaction can help stabilize a quasis table rephased tRNA:message complex, It was previously shown that fram eshifting in RF2 occurs more frequently if the codon 3' to the slipper y site is read by a rare tRNA, Consistent with that earlier work, in t he RF2 context frameshifting occurs substantially more frequently if t he arginine codon is CGG, which is read by a rare tRNA, In contrast, i n the 'normal' context frameshifting is only slightly greater at CGG t han at CGU. It is suggested that the Shine-Dalgarno-like interaction e levates frameshifting specifically during the pause prior to translati on of the second codon, which makes frameshifting exquisitely sensitiv e to the rate of translation of that codon, In both contexts frameshif ting increases in a mutant strain that fails to modify tRNA base A37, which is 3' of the anticodon, Thus, those base modifications may limit frameshifting at UUU codons, Finally, statistical analyses show that UUU Ynn dicodons are extremely rare in E.coli genes that have highly b iased codon usage.