- Nascent RNA is a ribonucleotide chain that is still being synthesized, so that its 3 end is paired with DNA where RNA polymerase is elongating.
- Monocistronic mRNA codes for one protein.
- Polycistronic mRNA includes coding regions representing more than one gene.
- A coding region is a part of the gene that represents a protein sequence.
- The leader of a protein is a short N-terminal sequence responsible for initiating passage into or through a membrane.
- The leader (5 UTR) of an mRNA is the nontranslated sequence at the 5 end that precedes the initiation codon.
- A trailer (3 UTR) is a nontranslated sequence at the 3 end of an mRNA following the termination codon.
- The intercistronic region is the distance between the termination codon of one gene and the initiation codon of the next gene.
- Transcription and translation occur simultaneously in bacteria, as ribosomes begin translating an mRNA before its synthesis has been completed.
- Bacterial mRNA is unstable and has a half-life of only a few minutes.
- A bacterial mRNA may be polycistronic in having several coding regions that represent different genes.
Messenger RNA has the same function in all cells, but there are important differences in the details of the synthesis and structure of prokaryotic and eukaryotic mRNA.
A major difference in the production of mRNA depends on the locations where transcription and translation occur:
- In bacteria, mRNA is transcribed and translated in the single cellular compartment; and the two processes are so closely linked that they occur simultaneously. Since ribosomes attach to bacterial mRNA even before its transcription has been completed, the polysome is likely still to be attached to DNA. Bacterial mRNA usually is unstable, and is therefore translated into proteins for only a few minutes.
- In a eukaryotic cell, synthesis and maturation of mRNA occur exclusively in the nucleus. Only after these events are completed is the mRNA exported to the cytoplasm, where it is translated by ribosomes. Eukaryotic mRNA is relatively stable and continues to be translated for several hours.
Figure 5.13 shows that transcription and translation are intimately related in bacteria. Transcription begins when the enzyme RNA polymerase binds to DNA and then moves along making a copy of one strand. As soon as transcription begins, ribosomes attach to the 5 end of the mRNA and start translation, even before the rest of the message has been synthesized. A bunch of ribosomes moves along the mRNA while it is being synthesized. The 3 end of the mRNA is generated when transcription terminates. Ribosomes continue to translate the mRNA while it survives, but it is degraded in the overall 5→3 direction quite rapidly. The mRNA is synthesized, translated by the ribosomes, and degraded, all in rapid succession. An individual molecule of mRNA survives for only a matter of minutes or even less.
Bacterial transcription and translation take place at similar rates. At 37°C, transcription of mRNA occurs at ~40 nucleotides/second. This is very close to the rate of protein synthesis, roughly 15 amino acids/second. It therefore takes ~2 minutes to transcribe and translate an mRNA of 5000 bp, corresponding to 180 kD of protein. When expression of a new gene is initiated, its mRNA typically will appear in the cell within ~2.5 minutes. The corresponding protein will appear within perhaps another 0.5 minute.
Bacterial translation is very efficient, and most mRNAs are translated by a large number of tightly packed ribosomes. In one example (trp mRNA), about 15 initiations of transcription occur every minute, and each of the 15 mRNAs probably is translated by ~30 ribosomes in the interval between its transcription and degradation.
The instability of most bacterial mRNAs is striking. Degradation of mRNA closely follows its translation. Probably it begins within 1 minute of the start of transcription. The 5 end of the mRNA starts to decay before the 3 end has been synthesized or translated. Degradation seems to follow the last ribosome of the convoy along the mRNA. But degradation proceeds more slowly, probably at about half the speed of transcription or translation.The stability of mRNA has a major influence on the amount of protein that is produced. It is usually expressed in terms of the half-life. The mRNA representing any particular gene has a characteristic half-life, but the average is ~2 minutes in bacteria.
This series of events is only possible, of course, because transcription, translation, and degradation all occur in the same direction. The dynamics of gene expression have been caught in flagrante delicto in the electron micrograph of Figure 5.14. In these (unknown) transcription units, several mRNAs are under synthesis simultaneously; and each carries many ribosomes engaged in translation. (This corresponds to the stage shown in the second panel in Figure 5.13.) An RNA whose synthesis has not yet been completed is often called a nascent RNA (Brenner et al., 1961).
Bacterial mRNAs vary greatly in the number of proteins for which they code. Some mRNAs represent only a single gene: they are monocistronic. Others (the majority) carry sequences coding for several proteins: they are polycistronic. In these cases, a single mRNA is transcribed from a group of adjacent genes. (Such a cluster of genes constitutes an operon that is controlled as a single genetic unit; see 10 The operon.)
All mRNAs contain two types of region. The coding region consists of a series of codons representing the amino acid sequence of the protein, starting (usually) with AUG and ending with a termination codon. But the mRNA is always longer than the coding region, extra regions are present at both ends. An additional sequence at the 5 end, preceding the start of the coding region, is described as the leader or 5 UTR (untranslated region). An additional sequence following the termination signal, forming the 3 end, is called the trailer or 3 UTR. Although part of the transcription unit, these sequences are not used to code for protein.
A polycistronic mRNA also contains intercistronic regions, as illustrated in Figure 5.15. They vary greatly in size. They may be as long as 30 nucleotides in bacterial mRNAs (and even longer in phage RNAs), but they can also be very short, with as few as 1 or 2 nucleotides separating the termination codon for one protein from the initiation codon for the next. In an extreme case, two genes actually overlap, so that the last base of one coding region is also the first base of the next coding region.
The number of ribosomes engaged in translating a particular cistron depends on the efficiency of its initiation site. The initiation site for the first cistron becomes available as soon as the 5 end of the mRNA is synthesized. How are subsequent cistrons translated? Are the several coding regions in a polycistronic mRNA translated independently or is their expression connected? Is the mechanism of initiation the same for all cistrons, or is it different for the first cistron and the internal cistrons?
Translation of a bacterial mRNA proceeds sequentially through its cistrons. At the time when ribosomes attach to the first coding region, the subsequent coding regions have not yet even been transcribed. By the time the second ribosome site is available, translation is well under way through the first cistron. Usually ribosomes terminate translation at the end of the first cistron (and dissociate into subunits), and a new ribosome assembles independently at the start of the next coding region. (We discuss the processes of initiation and termination in 6 Protein synthesis.)