Accumulation of pre-rRNAs is affected in brx1 mutants. (A) Shown is the primary pre-rRNA transcript of A. thaliana with the processing sites indicated by arrows on top. Probes (p) used for Northern blots are indicated below. Insertion: a 1083-nt sequence in the 5′ ETS specific for A. thaliana. (ETS) External transcribed spacer, (ITS) internal transcribed spacer. The A123B cluster is specific to Brassicaceae and was shown to be bound by the NF-B/D complex important for P-site cleavage (Sáez-Vasquez et al. 2004). (B) Total RNA from 18-day-old seedlings from wild-type (WT) and the brx1 mutants was separated on a 1.2% agarose gel, transferred onto nylon membrane and hybridized with the probes indicated in A. Mature chloroplast rRNAs visible with ethidium bromide (EtBr) staining (23S-1 and 16S) are shown in gray. The names of the detected pre-rRNAs are given on the right. (C) Individual Northern blots as shown in B were quantified and the band intensity was normalized to the 35S pre-rRNA that is unaltered in the mutants. Wild-type values were set to one. (D) Scheme of the pre-rRNAs that are detected in B. Light gray boxes mark the newly defined 33S(P′) and 27SA2 pre-rRNAs. Italic: 5′ extended pre-rRNAs seen in xrn2 mutants (Fig. 5). (E) Total RNA from 18-day-old seedlings from wild-type (WT) and the brx1 mutants was separated on an 8% acrylamide/8 M urea gel, transferred onto nylon membrane and hybridized with the probes indicated. Mature chloroplast rRNAs seen by ethidium bromide (EtBr) staining (23S-3, 5S, and 4.5S) are shown in gray. The names of the detected pre-rRNAs are given on the right.
