Rna Processing & Alternative Splicing

Functional RNAs are produced through modification of pre-RNA produced from transcription process except bacterial mRNAs which are as such used for protein synthesis without any modification. These series of modification involving removal of introns by splicing are known as processing of RNA.

Processing of rRNA & tRNA:

The processing of both rRNA & tRNA in prokaryotes and eukaryotes are similar. Eukaryotes – 4 species of rRNAs, three of which (28S, 18S & 5.8S) are derived by cleavage from single long precursor transcript(pre-rRNA). Whereas the fourth(5S) is transcribed from a separate gene. There are 3 rRNAs in prokaryotes(23S, 16S & 5S) which are also transcribed from a single pre-rRNA transcript. In prokaryotic cell initial cleavage yield separate precursor of 3 individual rRNAs which undergo secondary cleavage to produce functional forms. Whereas in eukaryotic cell(within nucleolus) initial cleavage near 5’ side of 5.8S rRNA results in 2 segments(18S & 28S + 5.8S). Further cleavage produce functional forms with 5.8S having hydrogen-bonded to 28S. Apart from this there will be also some addition of methyl groups to the bases & sugar moieties of specific nucleotides and conversion of some uridines to pseudouridines. Similarly from pre-tRNAs individual tRNA are synthesised in both prokaryotes & eukaryotes. In prokaryotes, some tRNA s are included in pre-rRNA transcripts.

RNase P enzyme(ribozyme) involved in processing at 5’ end of pre-tRNAs. It consists of RNA & protein molecules. Here RNA is responsible for catalytic activity whereas proteins are required for maximal activity. 3’ end processing involves action of conventional protein RNase and addition of a CCA terminus. As this CCA sequences are the site of protein attachment, it is present in all tRNAs for protein synthesis. In addition approximately 10% of bases of specific nucleotides are modified. Introns are spliced using endonuclease from pre-tRNAs.

Processing of mRNA in eukaryotes

In contrast to prokaryotes, pre-mRNA synthesised in nucleus of eukaryotes are modified before transported to cytoplasm. Processing involves modification of both ends of initial transcript and removal of introns. The C-terminal domain(CTD) of RNA polymerase II serve as binding site for enzyme complexes. Whereas polymerases I and III lack a CTD, so their transcripts are not processed by same enzyme complexes. Processing at 5’ end – addition of 7-methylguanosine cap. After transcription of first 20-30 nucleotides, enzymes for capping are recruited to pCTD. Capping initiated by addition of a GTP in reverse orientation to 5’ terminal nucleotide. Followed by addition of methyl group to this G residue & to ribose moieties of one or two 5’ nucleotides. This 5’ cap helps stabilising & aligning on ribosomes duting protein synthesis. At 3’ end – cleavage of primary transcript downstream to 10-30 nucleotides of highly conserved hexanucleotide(AAUAAA in mammalian cells) and addition of a poly-A tail known as polyadenylation. Thus G-U rich residue is degraded.

References:

1. The Cell by Cooper and Hausman, 5th edition.