RNA Processing:- Ribosomal RNAs and tRNAs Also Undergo Processing
Posttranscriptional processing is not limited to mRNA. Ribosomal RNAs of both prokaryotic and eukaryotic cells are made from longer precursors called preribosomal RNAs, or pre-rRNAs, synthesized by Pol I. In bacteria, 16S, 23S, and 5S rRNAs (and some tRNAs, although most tRNAs are encoded elsewhere) arise from a single 30S RNA precursor of about 6,500 nucleotides. RNA at both ends of the 30S precursor and segments between the rRNAs are removed during processing (Fig. 1).
FIGURE 1 Processing of pre-rRNA transcripts in bacteria. 1 Before cleavage, the 30S RNA precursor is methylated at specific bases. 2 Cleavage liberates precursors of rRNAs and tRNA(s). Cleavage at the points labeled 1, 2, and 3 is carried out by the enzyme's RNase III, RNase P, and RNase E, respectively. As discussed later in the text, RNase P is a ribozyme. 3 The final 16S, 23S, and 5S rRNA products result from the action of a variety of specific nucleases. The seven copies of the gene for pre-rRNA in the E. coli chromosome differ in the number, location, and identity of tRNAs included in the primary transcript. Some copies of the gene have additional tRNA gene segments between the 16S and 23S rRNA segments and at the far 3end of the primary transcript.
The genome of E. coli encodes seven pre-rRNA molecules. All these genes have essentially identical rRNA coding regions, but they differ in the segments between these regions. The segment between the 16S and 23S rRNA genes generally encodes one or two tRNAs, with different tRNAs arising from different pre-rRNA tran scripts. Coding sequences for tRNAs are also found on the 3 side of the 5S rRNA in some precursor transcripts. In eukaryotes, a 45S pre-rRNA transcript is processed in the nucleolus to form the 18S, 28S, and 5.8S rRNAs characteristic of eukaryotic ribosomes (Fig. 2). The 5S rRNA of most eukaryotes is made as a completely separate transcript by a different polymerase (Pol III instead of Pol I). Most cells have 40 to 50 distinct tRNAs, and eu karyotic cells have multiple copies of many of the tRNA genes. Transfer RNAs are derived from longer RNA pre cursors by enzymatic removal of nucleotides from the 5 and 3 ends (Fig. 3). In eukaryotes, introns are present in a few tRNA transcripts and must be excised. Where two or more different tRNAs are contained in a single primary transcript, they are separated by enzymatic cleavage. The endonuclease RNase P, found in all organisms, removes RNA at the 5end of tRNAs. This enzyme contains both protein and RNA. The RNA component is essential for activity, and in bacterial cells it can carry out its processing function with precision even without the protein component. RNase P is therefore another example of a catalytic RNA, as described in more detail below. The 3end of tRNAs is processed by one or more nucleases, including the exonuclease RNase D.
FIGURE 2 Processing of pre-rRNA transcripts in vertebrates. In step 1, the 45S precursor is methylated at more than 100 of its 14,000 nucleotides, mostly on the 2-OH groups of ribose units retained in the final products. 2A series of enzymatic cleavages produces the 18S, 5.8S, and 28S rRNAs. The cleavage reactions require RNAs found in the nucleolus, called small nucleolar RNAs (snoRNAs), within protein complexes reminiscent of spliceosomes. The 5S rRNA is produced separately.
FIGURE 3 Processing of tRNAs in bacteria and eukaryotes. The yeast tRNATyr (the tRNA specific for tyrosine binding; see Chapter 27) is used to illustrate the important steps. The nucleotide sequences shown in yellow are removed from the primary transcript. The ends are processed first, the 5 end before the 3 end. CCA is then added to the 3end, a necessary step in processing eukaryotic tRNAs and those bacterial tRNAs that lack this sequence in the primary transcript. While the ends are being processed, specific bases in the rest of the transcript are modified (see Fig. 4). For the eukaryotic tRNA shown here, the final step is splicing of the 14-nucleotide intron. Introns are found in some eukaryotic tRNAs but not in bacterial tRNAs.
Transfer RNA precursors may undergo further post transcriptional processing. The 3-terminal trinucleotide CCA(3) to which an amino acid will be attached during protein synthesis (Chapter 27) is absent from some bacterial and all eukaryotic tRNA precursors and is added during processing (Fig. 3). This addition is carried out by tRNA nucleotidyltransferase, an unusual enzyme that binds the three ribonucleoside triphosphate precursors in separate active sites and catalyzes formation of the phosphodiester bonds to produce the CCA (3) sequence. The creation of this defined sequence of nucleotides is therefore not dependent on a DNA or RNA template—the template is the binding site of the enzyme. The final type of tRNA processing is the modification of some of the bases by methylation, deamination, or reduction (Fig. 4). In the case of pseudo uridine (Ψ), the base (uracil) is removed and reattached to the sugar through C-5. Some of these modified bases occur at characteristic positions in all tRNAs (Fig.3).
FIGURE 4 Some modified bases of tRNAs, produced in posttranscriptional reactions. The standard symbols (used in Fig.3) are shown in parentheses. Note the unusual ribose attachment point in pseudo uridine.
