BBC2012 Molecular Genetics
John Bothwell (j.bothwell@qub.ac.uk)
Lecture 22: Principles of eukaryotic transcription
§ 1. Transcription: DNA to RNA
Transcription is the passing of information from DNA to RNA. Lots of RNA:
Lewin, Chapters 2.11
Two things will probably leap out at you:
I.
There’s a lot of RNA involved. Why should the cell bother with RNA as an intermediate?
Why, for instance, shouldn’t information go straight from DNA to protein?
II. Similarly, the whole transcriptional network is very complicated.
I.
A feature may confer some function which has been selected for during evolution - this is the
bog-standard evolutionary ‘adaptive’ explanation, e.g. ‘fish have fins because fins are good for
swimming with’ (see §3, below).
II. A feature may have no adaptive function, but may persist simply because its ancestors had it
and there was no easy way to remove it during evolution (see §4, below).
1
Lewin, Chapter 28
§ 2. Complexity may reflect function, or origin, or both
We’ll deal with the second point in §1 first - why is the transcriptional network so complicated?
Very generally speaking, biological features exist for one of two reasons (which aren’t always
mutually exclusive):
BBC2012 Molecular Genetics
John Bothwell (j.bothwell@qub.ac.uk)
§ 3. The eukaryotic transcriptional machinery is complex because complexity
ensures timely, sensitive, and stable gene expression
While there are many potential control points at which gene expression could be regulated, the
initiation of transcription is the major point of control. So, what functions must the transcriptional network
have if it’s to be trusted with a task of such importance?
I.
Transcription needs to be selective. The phenotypic differences between cells reflect tissuespecific patterns of gene expression, with only a relatively small subset of nuclear genes being
expressed in any one cell type at any one time. The transcriptome, therefore, doesn’t match the
genome, a fact most commonly shown today using ESTs, microarrays, or Northern blots 1,:
II. Transcription needs to amplify the DNA signal. This was, in fact, one of the first things which
made people realise what RNA did; Torbjörn Caspersson2 and Jean Brachet showed that RNA
was elevated in the cytoplasm of growing cells, such as roots and eggs.
III. This selective amplification needs to be timely; it needs to happen at, and for, the right time
(different RNAs have different half lives; see Figure, below3 ). Again, this timeliness was one of
the things which gave RNA away, following ‘Ken’ Volkin and Lazarus Astrachan’s finding that
RNA rose rapidly after phage infection.
1 This
example is from Darakhshan, F., Hajduch, E., Kristiansen, S., Richter, E. A. & Hundal, H. S. Biochemical and functional
characterization of the GLUT5 fructose transporter in rat skeletal muscle. Biochem J 336 ( Pt 2), 361-366 (1998).
Caspersson, T. & Schultz, J. Ribonucleic acids in both nucleus and cytoplasm, and the function of the nucleolus. Proceedings of the
National Academy of Sciences of the United States of America 26, 507-515 (1940)
2
3 This
example is from Coulon,V.,Veyrune, J. L., Tourkine, N.,Vié, A., et al. A novel calcium signaling pathway targets the c-fos
intragenic transcriptional pausing site. J Biol Chem 274, 30439-30446 (1999).
2
BBC2012 Molecular Genetics
John Bothwell (j.bothwell@qub.ac.uk)
IV. Timely selective amplification demands regulation - to ensure timeliness, confer selectivity and
guard against waste - and transcription is very tightly regulated (see §5, below). The
connexions between the components of the transcriptional machinery are, therefore, not
random, but highly structured and often fall into one of a limited number of motifs. By
combining these motifs, a regulatory network can give rise to stable and sensitive responses.
Three particular network architectures are worth noting:
1. Feed-forward loops, in which A activates B and C, and B also activates C (below).
Broadly speaking, feedforward loops improve the sensitivity of a network.
2. Feed-back loops, in which downstream elements regulate upstream ones. Broadly
speaking, again, feedback loops improve the stability of a network.
3. Noise, in which an intermediate (B, in the cartoon below) is continually made and
broken down. For various reasons, keeping an intermediate continuously turning over
improves the sensitivity of a network. An analogy would be keeping your car engine
running at a red light: the engine’s turning over (hence burning up fuel and money), but
the increased speed with which you can go once the light turns green makes up for that
extra cost in fuel.
So, the upshot of this section (§3) is that the complexity of the transcriptional machinery reflects its
function as a robust and sensitive gene expression system.
3
BBC2012 Molecular Genetics
John Bothwell (j.bothwell@qub.ac.uk)
§ 4. The complexity of the transcriptional network may also reflect its evolution
In §3 we assumed that the complexity of the transcriptional network is entirely functional and, in
general, it’s usually assumed that evolution produces optimal structures. This is not always the case.
Biological features are selected, but are also constrained - at least to some extent - by their starting
points, with one of the most famous examples being the mammalian recurrent laryngeal nerve1.
With this in mind, and having noted in §1 that there’s a lot of RNA about in the transcriptional
system, there’s a decent school of argument that there’s so much RNA about because, when life
began, RNA was the original genetic and catalytic material. This is the ‘RNA World’ hypothesis 2,
and we’ll come back to it in Lecture 29...
§ 5. Enough theory - how is the initiation of transcription regulated?
In eukaryotes, the initiation of transcription happens in a series of stages:
I.
1 This
2
is from the C4 series ‘Inside’s Nature’s Giants’ (sorry about the ads)
See here, as well...
3 These
are usually abbreviated to ‘TF’
4
Lewin, Chapter 20.1
We need to know where to start. Genes contain specific sequences which tell transcription
where to start, and these are located by sequence-specific factors.
II. We need to get at the DNA containing the relevant gene. This happens through chromatin
remodelling (Lecture 23).
III. Transcription factors 3 then bind to DNA promoter sequences in the gene (‘cis-acting’
sequences).
IV. Once a defined set of transcription factors - called the basal transcription factors - have bound to
the core promoter, the site is ‘primed’ for RNA polymerase to bind to the startpoint. RNA
polymerase on its own can’t recognise DNA sequences, so it needs the transcription factors to
direct its own binding.
V. Many genes have multiple promoters, which are activated by different combinations of
transcription factors. The exact promoter used will determine exactly where transcription
starts and this will, in turn, affect the exact RNA sequence produced which will, in its turn,
determine what happens to that RNA.
VI. Eukaryotes have three types of RNA polymerase - I, II and III - which transcribe different
types of RNA.
VII. The DNA strands are then separated and RNA is made.
VIII.We need to know where to stop. There are termination sequences at the end of the gene and,
once they’re reached, the RNA polymerase detached and RNA is released. Again, many genes
have multiple termination sites, allowing variations.
IX. The immediate products of transcription are the ‘primary transcripts’, or pre-RNAs. These
are then modified during or after transcription.
X. The modified RNA is then exported from the nucleus into the cytoplasm. This is, incidentally,
why attentuation doesn’t work in eukaryotes, as translation doesn’t occur at the same site as
transcription.
XI. RNA is then degraded.