Eukaryotes have three types of RNA polymerase. RNA polymerase I transcribes the genes that code for most of the ribosomal RNAs. All messenger RNAs are synthesized using RNA polymerase II. Transfer RNA genes are transcribed by RNA polymerase III. This last enzyme also catalyzes the synthesis of several small RNAs including the 5S ribosomal RNA. The chemical reaction catalyzed by these three RNA polymerases, the formation of phosphodiester links between nucleotides, is the same in eukaryotes and bacteria.
A newly synthesized eukaryotic mRNA undergoes several modifications before it leaves the nucleus (Fig. 6.13). The first is known as capping. Very early in transcription the 5'-terminal triphosphate group is modified by the addition of a guanosine via a 5'-5'-phosphodiester link. The guanosine is subsequently methylated to form the 7-methyl guanosine cap. The 3' ends of nearly all eukaryotic mRNAs are modified by the addition of a long stretch of adenosine residues, the poly-A tail (Fig. 6.13). A sequence AAUAAA is found in most eukaryotic mRNAs about 20 bases from where the poly-A tail is added and is probably a signal for the enzyme poly-A polymerase to bind and to begin the polyadenylation process. The length of the poly-A tail varies, it can be as long as 250 nucleotides. Unlike DNA, RNA is an unstable molecule, and the capping of eukaryotic mRNAs at their 5' ends and the addition of a poly-A tail to their 3' end increases the lifetime of mRNA molecules by protecting them from digestion by nucleases.
Many eukaryotic protein-coding genes are split into exon and intron sequences. Both the exons and introns are transcribed into mRNA. The introns have to be removed and the exons joined together by a process known as RNA splicing before the mRNA can be used to make protein. Removal of introns takes place within the nucleus. Splicing is complex and not yet fully understood. It has, however, certain rules. Within an mRNA the first two bases following an exon are always GU and the last two bases of the intron are AG. Several small nuclear RNAs (snRNAs) are involved in splicing. These are complexed with a number of proteins to form a structure known as the spliceosome. One of the snRNAs is complementary in sequence to either end of the intron sequence. It is thought that binding of this snRNA to the intron, by complementary base pairing, brings the two exon sequences together, which causes the intron to loop out (Fig. 6.13). The proteins in the spliceosome remove the intron and join the exons together. Splicing is the final modification made to the mRNA in the nucleus. The mRNA is now transported to the cytoplasm for protein synthesis.
As well as removing introns, splicing can sometimes remove exons in a process called alternative splicing. This allows the same gene to give rise to different proteins at different times or in different cells. For example, alternative splicing of the gene for the molecular motor dynein produces motors that transport different types of cargo (page 390).
precursor — mRNA
polyadenylation spliceosome snRNAs formation
7-methyl guanosine cap a a u a a a AAAA 3'
poly A tail o
|— intron loops out on polyadenylation "signal"
5' m7Gppp spliceosome
|— intron loops out spliceosome
Figure 6.13. mRNA processing in eukaryotes.
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