Proteins encoded by a complex chloroplast transcription unit are each translated from both monocistronic and polycistronic mRNAs

Abstract

Chloroplast genes are typically organized into polycistronic transcription units that give rise to complex sets of overlapping RNAs through a series of processing steps. The functional significance of this complicated mode of expression is unknown. To determine whether processing of the primary transcript is required to create translatable mRNAs, the translational properties of the RNAs derived from the maize psbB gene cluster (containing the psbB, psbH, petB and petD genes) were examined. Almost all of the approximately 20 RNAs derived from this region co-sediment with polysomes in sucrose gradients, suggesting that at least one coding region on most transcripts is translated. To determine which sequences are translated on each polycistronic RNA, antibodies to psbB, petB or petD proteins were used to immunoselect polysomes engaged in the synthesis of each protein. Northern and S1 nuclease analyses of the immunoselected RNAs revealed that (i) potential start codons within the petB and petD introns are not functional in translation; (ii) all transcripts containing spliced petB or petD sequences are translated to give these proteins, regardless of upstream or downstream sequences; (iii) psbB is translated from all transcripts encoding it. It is concluded that intercistronic processing is not required for translation of these RNAs, although certain processing steps may enhance translational efficiency.