1Three RNAs, three jobs

An RNA molecule's job depends on two things: its sequence of bases and the shape it folds into. A single strand of RNA can fold back and pair with itself wherever its bases are complementary, giving loops and stems.

• Messenger RNA (mRNA) carries the information of a gene from the DNA (in the nucleus, in a eukaryote) to a ribosome in the cytoplasm. Its job is all sequence: its bases are read three at a time, as codons, in topic 6.4.
• Transfer RNA (tRNA) is a short RNA folded into a compact shape. Each kind of tRNA binds one specific amino acid at one end and has a three-base anticodon at the other. The anticodon pairs with a matching codon on the mRNA, so the tRNA delivers the right amino acid to the right place. That is how the sequence of codons becomes a sequence of amino acids, the primary structure of the polypeptide.
• Ribosomal RNA (rRNA) is the main building block of ribosomes. Folded rRNA, together with proteins, forms the ribosome's two subunits, and rRNA does the central work of translation (topic 6.4).

So "RNA" is not one thing. If an exam asks which RNA has an anticodon, it is tRNA; which carries the codons, mRNA; which forms ribosomes, rRNA.

2Transcription

The DNA of a gene has two strands, but only one is used as the pattern.

• The template strand is the one RNA polymerase reads. The RNA is complementary to it.
• The other strand, the coding strand (also called the non-template strand), has the same sequence as the RNA, except that it has T where the RNA has U.

RNA polymerase binds at the start of the gene (at a region called the promoter, topic 6.6), opens a short stretch of the double helix, and moves along the template strand. It reads the template 3′ to 5′ and builds RNA 5′ to 3′, adding each new nucleotide to the 3′ end of the growing chain. Behind it, the DNA strands close up again. At the end of the gene RNA polymerase releases the RNA. Unlike DNA polymerase, RNA polymerase needs no primer: it can start a chain on its own. Figure 1 shows the enzyme part-way along a gene.

Worked example. A stretch of template strand reads 3′-TACGGTCAA-5′. What is the mRNA?

• Pair each base: T → A, A → U, C → G, G → C.
• mRNA: 5′-AUGCCAGUU-3′. The mRNA runs antiparallel to the template, so its 5′ end sits opposite the template's 3′ end.
• Check against the coding strand, which would read 5′-ATGCCAGTT-3′: the same, with U for T. That is a fast way to check any answer.

3Processing the message in eukaryotes

In a eukaryote, transcription happens in the nucleus and the first transcript, the pre-mRNA, is changed by enzymes before it is translated. In prokaryotes there is no nucleus and the mRNA is used as it is made (topic 6.4). Figure 2 follows a pre-mRNA with four exons.

A cap on the 5′ end. A modified guanine nucleotide (the CED calls it the GTP cap) is added to the 5′ end. It helps the ribosome recognise the mRNA and attach to it, and it protects that end.

A poly-A tail on the 3′ end. A string of many adenine nucleotides is added to the 3′ end. It makes the mRNA more stable, so it lasts longer in the cytoplasm before enzymes break it down. A longer-lasting mRNA can be translated more times, so more protein is made from it.

Splicing. Most eukaryotic genes are split. Exons are the parts kept in the mature mRNA; introns are the parts cut out. A useful hook: ex*ons are *ex*pressed, *in*trons stay *in the nucleus. The introns are removed and the exons are spliced together in their original order.

Alternative splicing. The cell does not always keep the same exons. In one tissue it may keep exons 1, 2 and 4; in another, exons 1, 3 and 4. Each set gives a different mature mRNA, each mRNA a different polypeptide. In Figure 2 the one pre-mRNA becomes polypeptide X or polypeptide Y. This is how a few tens of thousands of human genes can make many more kinds of protein.

RNA is built 5′ to 3′ on a template read 3′ to 5′; in eukaryotes the pre-mRNA gets a cap, a tail, and its introns removed, and choosing which exons to keep lets one gene make several proteins.

4Working with processing data

Worked example (invented data). A gene's pre-mRNA has four exons of 240, 390, 180 and 510 nucleotides, separated by three introns. In liver cells all four exons are kept; in muscle cells exon 3 is skipped. Ignoring the cap and tail, how long is each mature mRNA?

• Liver: 240 + 390 + 180 + 510 = 1,320 nucleotides.
• Muscle: 240 + 390 + 510 = 1,140 nucleotides.
• The difference, 180 nucleotides, is exactly exon 3. Since 180 ÷ 3 = 60, the muscle protein lacks 60 amino acids in the middle and is otherwise identical (provided exon 3's length is a multiple of three, the reading frame of the later exons is unchanged).

Reading a stability experiment (invented). Two versions of one mRNA are injected into cells: one with a normal poly-A tail and one with no tail. The amount remaining is measured over time.

Claim: the tail makes mRNA more stable. Evidence: after 4 h, 77% of the tailed mRNA remains but only 17% of the untailed. Reasoning: the tail protects the 3′ end from being broken down, so the tailed mRNA survives longer and can be translated into more protein.