THE LOW-LEVEL EXPRESSION OF CHLORAMPHENICOL ACETYLTRANSFERASE (CAT) MESSENGER-RNA IN ESCHERICHIA-COLI IS NOT DEPENDENT ON EITHER SHINE-DALGARNO OR THE DOWNSTREAM BOXES IN THE CAT GENE

Recent studies have shown that the canonical Shine-Dalgarno (SD)-anti- SD interaction is dispensable for the initiation of translation of cer tain mRNAs in Escherichia coli. Alternative non-SD sequences (located upstream from the initiation codon) and also downstream sequences (''d ownstream boxes'') complementary to 16S rRNA were found to be involved in the initiation of translation of mRNAs devoid of either SD or any leader sequences. In this study the chloramphenicol acetyltransferase (CAT) gene was modified to remove the 5' terminal non-translated regio n and/or the two potential downstream boxes in the CAT gene. Thus a se ries of ten CAT gene constructs was created and expressed in E. coli u nder a strong constitutive promoter. The results showed that CAT mRNAs devoid of both leader sequence nucleotides and the two downstream box es in the CAT gene remained active in vivo and produced CAT protein in sufficient amounts for survival of the transformed cells at chloramph enicol concentrations up to 20-30 mu g/ml.