Identification of an rRNA operon promoter from Zea mays chloroplasts which excludes the proximal tRNA^Val _GAC from the primary transcript

Abstract

The transcriptional start site of the rRNA operon from Zea mays chloroplasts has been identified by a combination of S1 mapping and Southern hybridization with in vitro capped chloroplast RNA as radioactive probe. This is the first example in which the transcriptional start site of a choloroplast gene has been established by identification of the triphosphate‐bearing RNA terminus. The start site is located at position −117 proximal to the 16S rRNA gene and is preceded by −10 and −35 sequences homologous to prokaryotic promoters. The primary transcript directed by this promoter does not include tRNAValGAC sequences which are coded further upstream between positions −302 and −231. A major processing site of the primary rRNA transcript was identified at position −30 which is embedded in a secondary structure typical for prokaryotic RNase III processing sites. Several putative processing and start sites of the tRNAValGAC transcipts have been mapped by primer extension with reverse transcriptase.

Nuclear Mutants of Maize with Defects in Chloroplast Polysome Assembly Have Altered Chloroplast RNA Metabolism

“…Figure 5 shows the results of RNA gel blots in which clones derived from the maize Chloroplast rRNA genes were used as probes. Although a small fraction of the maize Chloroplast 23S rRNA is recovered intact, most of it is recovered as two smaller molecules: a 1.8-kb fragment is derived from the 5' portion of the 23S rRNA, and a 1.2-kb fragment is derived from the 3' portion ( Figure 5A; Kossel et al, 1985). Both of the "23S" fragments were unaltered in size and accumulated to normal levels in each mutant.…”

“…The probe for the 5' end of the pre-16s rRNA was an 880-bp Hincll fragment including the rRNA promoter and extending ~5 0 0 bp into sequences encoding the mature 16s rRNA (Schwarz and Kossel, 1980;Strittmatter et al, 1985). The 3' probe was a 1490-bp fragment with one end at the Hincll site in the middle of the 16s coding region and the other end at a Taql site in the frnl intron (Koch et al, 1981).…”

Alternative base pairing between 5′- and 3′-terminal sequences of small subunit RNA may provide the basis of a conformational switch of the small ribosomal subunit

“…On the other hand, precursors of bacterial 16S rRNAs are capable of forming long double stranded stems by base pairing of sequences immediately proximal and distal to the structural region (8), which is suggestive of, at least a temporary close proximity of the two mature ends during processing of 16S rRNAs. In an earlier proposal of such a processing stem for a 16S rRNA precursor from Zea mays chloroplasts, base pairing between the two terminal regions of mature 16S rRNA was also implied (9). Mapping of the two termini of E. coli 16S rRNA has been achieved by chemical labelling and immunoelectron microscopy of reconstituted 30S particles (for review see ref.…”

Organ-specific Transcription of the rrn Operon in Spinach Plastids

“…1A, lane C). The band named Pro2 corresponds in size to the processing intermediate, which was previously characterized in maize and tobacco chloroplasts (3,29), suggesting that this processing site is universal in plastids of different plant species. The band named PC corresponds to the previously characterized putative transcription start site of the spinach rrn operon (23) the presence of very long precursor transcripts in root tissues.…”

“…These E. coli-like consensus sequences serve as promoter structures (3)(4)(5) or as regulatory elements (6,7). However, the interpretation of results on studies of transcriptional regulation in plastids is complicated by the existence of different types of RNA polymerases.…”