No abstract

We have determined the complete nucleotide sequence (4712 nucleotides) of the mouse 28S rRNA gene. Comparison with all other homologs indicates that the potential for major variations in size during the evolution has been restricted to a unique set of a few sites within a largely conserved secondary structure core. The D (divergent) domains, responsible for the large increase in size of the molecule from procaryotes to higher eukaryotes, represent half the mouse 28S rRNA length. They show a clear potential to form self-contained secondary structures. Their high GC content in vertebrates is correlated with the folding of very long stable stems. Their comparison with the two other vertebrates, xenopus and rat, reveals an history of repeated insertions and deletions. During the evolution of vertebrates, insertion or deletion of new sequence tracts preferentially takes place in the subareas of D domains where the more recently fixed insertions/deletions were located in the ancestor sequence. These D domains appear closely related to the transcribed spacers of rRNA precursor but a sizable fraction displays a much slower rate of sequence variation.

SummaryBy generating a specialized cDNA library from the archaeon Sulfolobus solfataricus , we have identified 57 novel small non-coding RNA (ncRNA) candidates and confirmed their expression by Northern blot analysis. The majority was found to belong to one of two classes, either antisense or antisense-box RNAs, where the latter only exhibit partial complementarity to RNA targets. The most prominent group of antisense RNAs is transcribed in the opposite orientation to the transposase genes, encoded by insertion elements (transposons). Thus, these antisense RNAs may regulate transposition of insertion elements by inhibiting expression of the transposase mRNA. Surprisingly, the class of antisense RNAs also contained RNAs complementary to tRNAs or sRNAs (smallnucleolar-like RNAs). For the antisense-box ncRNAs, the majority could be assigned to the class of C/D sRNAs, which specify 2 ¢ ¢ ¢ ¢ -O-methylation sites on rRNAs or tRNAs. Five C/D sRNAs of this group are predicted to target methylation at six sites in 13 different tRNAs, thus pointing to the widespread role of these sRNA species in tRNA modification in Archaea. Another group of antisense-box RNAs, lacking typical C/D sRNA motifs, was predicted to target the 3 ¢ ¢ ¢ ¢ -untranslated regions of certain mRNAs. Furthermore, one of the ncRNAs that does not show antisense elements is transcribed from a repeat unit of a cluster of small regularly spaced repeats in S. solfataricus which is potentially involved in replicon partitioning. In conclusion, this is the first report of stably expressed antisense RNAs in an archaeal species and it raises the prospect that antisense-based mechanisms are also used widely in Archaea to regulate gene expression.

RNA polymerase I terminates transcription of mouse rDNA 565 bp downstream of the 3' end of mature 28S rRNA. This specific termination event can be duplicated in a nuclear extract system. RNA molecules with authentic 3' ends are transcribed from ribosomal minigene constructs provided the templates retain a minimal length of downstream spacer sequences. The nucleotide sequence of the region of transcription termination contains a set of repetitive structural elements consisting of 18 bp conserved nucleotides surrounded by stretches of pyrimidines. Termination in vivo occurs within the first element. This site is preferentially used in vitro at low template concentrations. At increasing DNA concentrations a termination site within the second repetitive element is used. Competition experiments with defined 3'-terminal fragments suggest that transcription termination by RNA polymerase I requires interaction of some factor (or factors) with the repetitive structural elements in the 3' nontranscribed spacer.