Difficulty: Intermediate | Prerequisites: Part 1 of these notes (transcription phases, initiation sub-steps). You should understand closed complex, open complex, and promoter escape before reading this.
Part 1 covered what happens once RNA polymerase sits down on a promoter. This section steps back to ask: how does the polymerase find the right spot in the first place? In bacteria, the answer centres on the sigma factor, which converts a non-specific core enzyme into a promoter-recognising holoenzyme. The promoter itself is not a single sequence but a collection of short DNA elements (the -35 region, the -10 region, and several optional extras) whose exact sequences determine how strongly a gene is transcribed. Understanding promoter architecture is essential for gene regulation, because cells control gene expression partly by controlling how well RNA Pol recognises and binds each promoter.
Bacterial RNA polymerase needs a sigma factor (commonly sigma-70 in E. coli) to recognise promoters specifically. Promoters typically contain -10 and -35 sequence elements whose closeness to the consensus sequence determines promoter strength. Additional elements (UP-element, extended -10, discriminator) fine-tune polymerase binding and complex stability.
Core enzyme
The bacterial RNA polymerase without its sigma factor. Its subunit composition is α₂ββ'ω. The core enzyme can initiate transcription at random sites on naked DNA in vitro, but in living cells it cannot locate promoters on its own.
Think of it as a capable engine with no steering: it can synthesise RNA, but it cannot find the right starting point.
Sigma (σ) factor
A dissociable initiation factor that binds to the core enzyme and directs it to promoters. Different sigma factors recognise different promoter sequences, allowing the cell to redirect transcription under changing conditions.
In simple terms, sigma is the GPS unit that tells the engine where to go.
Holoenzyme
The complete, promoter-competent form of bacterial RNA polymerase: core enzyme + sigma factor. In E. coli, the predominant form is core + σ70.
Sigma-70 (σ70)
The primary (housekeeping) sigma factor in E. coli. It directs transcription of most genes under normal growth conditions by recognising the -10 and -35 promoter elements.
-10 region (Pribnow box)
A conserved 6-nucleotide sequence centred approximately 10 bp upstream of the +1 transcription start site. Part of the promoter recognised by σ70. The consensus is TATAAT.
In simple terms, this is the spot where the DNA most readily melts open to form the transcription bubble, because it is AT-rich.
-35 region
A conserved 6-nucleotide sequence centred approximately 35 bp upstream of the +1 site. The consensus is TTGACA. Together with the -10 region, it forms the core of σ70 promoter recognition.
Consensus sequence
The idealised, statistically derived sequence representing the most common nucleotide at each position across many aligned promoters. Very few promoters match it perfectly, but most differ by only a few bases.
Think of it as the "perfect" version that real promoters approximate to varying degrees.
Promoter strength
A measure of how many transcripts a promoter initiates per unit time. It depends on three things: how well the promoter binds polymerase initially, how efficiently it supports isomerisation (closed → open complex), and how readily the polymerase escapes.
UP-element
An AT-rich sequence found upstream of the -35 region in some strong promoters. It provides an additional contact surface for the α subunit of RNA polymerase, increasing binding affinity. Found, for example, at rRNA gene promoters.
Extended -10 element
A promoter variant in which a short additional sequence is appended to the upstream end of the -10 region, compensating for the absence of a -35 region. The extra contacts between this element and the polymerase replace those normally made by the -35 region.
The E. coli gal operon promoter is the classic example.
Discriminator
A short DNA element found just downstream of the -10 region in some promoters. The strength of the interaction between the discriminator and RNA polymerase influences how stable the enzyme–promoter complex is.
Purified core enzyme (α₂ββ'ω) can bind DNA and start RNA synthesis at essentially any point on a DNA molecule. This has been demonstrated in vitro.
Inside the cell, however, transcription initiates only at promoters. The difference is the sigma factor.
Adding sigma to the core enzyme produces the holoenzyme, which loses the ability to bind random DNA but gains the ability to recognise and bind specifically at promoter sequences.
After initiation, sigma typically dissociates, and the core enzyme carries out elongation on its own.
The standard σ70 promoter contains two conserved 6-nucleotide blocks:
The -10 region (consensus: TATAAT), centred ~10 bp upstream of +1
The -35 region (consensus: TTGACA), centred ~35 bp upstream of +1
These two blocks are separated by a non-conserved spacer of 17–19 bp. The optimum spacing is 17 bp.
The vast majority of σ70 promoters have recognisable -10 and -35 regions, but the exact sequences vary from gene to gene.
The consensus sequence is derived by aligning many promoters and taking the most frequent nucleotide at each position.
Hardly any real promoter matches the consensus perfectly, but most differ by only a few nucleotides.
Promoters whose sequences are closer to the consensus tend to be "stronger," meaning they initiate more transcripts per unit time.
Promoter strength depends on three factors:
Initial binding affinity (how well the holoenzyme recognises and binds the promoter)
Isomerisation efficiency (how readily the closed complex converts to the open complex)
Escape efficiency (how easily the polymerase transitions past the initial transcribing complex)
This heterogeneity is functional: genes that need high expression levels tend to have strong (consensus-like) promoters, while genes that need lower or more regulated expression have weaker ones.
Found upstream of the -35 region in certain strong promoters.
AT-rich; provides an extra binding surface for the α subunit (specifically the αCTD, the C-terminal domain of α).
Increases overall polymerase recruitment.
Classic example: promoters of rRNA genes, which need very high transcription rates.
Some σ70 promoters lack a recognisable -35 region entirely.
Instead, they carry an extended version of the -10 region with an additional short sequence at its upstream end.
The extra polymerase contacts provided by this extension compensate for the missing -35 element.
Example: the E. coli gal genes, whose products direct galactose metabolism.
A short element found just downstream of the -10 region.
The strength of the discriminator–polymerase interaction affects how stable the enzyme–promoter complex is once formed.
This can influence whether the polymerase proceeds efficiently to open-complex formation or tends to dissociate.
Promoter engineering is a cornerstone of synthetic biology and biotechnology. When researchers want a bacterium to produce a recombinant protein at high levels, they place the gene downstream of a strong, consensus-like promoter. Conversely, when tight regulation is needed (for example, producing a toxic intermediate), they use weaker or inducible promoters. Understanding the -10/-35 architecture and how UP-elements boost expression is directly relevant to designing expression vectors in the lab.
Students often assume the -10 and -35 sequences are identical across all promoters. They are not; the consensus is a statistical ideal. Real promoters vary, and that variation is what gives each promoter its characteristic strength.
It is common to think that a promoter missing the -35 region cannot function. It can, if it has an extended -10 element that compensates with additional polymerase contacts.
Students sometimes confuse sigma factors with transcription factors. Sigma factors are subunits of the RNA polymerase holoenzyme itself. Transcription factors (activators and repressors) are separate proteins that modulate polymerase activity at specific promoters.
Some students assume that "stronger promoter" simply means "tighter binding." Strength actually reflects three properties: binding, isomerisation, and escape.
⚠️ Know the subunit composition of core enzyme (α₂ββ'ω) vs. holoenzyme (core + σ).
⚠️ Be able to state the consensus sequences for the -10 (TATAAT) and -35 (TTGACA) regions, their positions relative to +1, and the optimal spacer length (17 bp).
⚠️ Understand what "promoter strength" means and the three factors that determine it (binding, isomerisation, escape).
⚠️ Be able to describe the UP-element, extended -10, and discriminator, including what each compensates for or enhances.
⚠️ Know the distinction between core enzyme behaviour in vitro (random initiation) vs. in vivo (promoter-only initiation with sigma).
True or False: The bacterial core enzyme can initiate transcription at promoters without a sigma factor.
A: False. Without sigma, core enzyme binds DNA non-specifically and cannot recognise promoters.
Fill in the blank: The consensus sequence for the -10 region is ______.
A: TATAAT
True or False: All σ70 promoters must have both a -35 and a -10 region to function.
A: False. Some promoters lack the -35 region and instead use an extended -10 element.
Fill in the blank: The optimal spacing between the -10 and -35 elements is ______ bp.
A: 17
True or False: A promoter closer to the consensus sequence is generally weaker than one that diverges from it.
A: False. Closer to consensus generally means stronger.
Q: What is the difference between the core enzyme and the holoenzyme?
A: The core enzyme (α₂ββ'ω) can synthesise RNA but binds DNA non-specifically. The holoenzyme is the core enzyme plus a sigma factor, which directs the complex to bind specifically at promoter sequences.
Q: Describe the structure of a typical σ70 promoter.
A: Two conserved 6-nucleotide sequences, the -10 region (consensus TATAAT) and the -35 region (consensus TTGACA), separated by a non-conserved spacer of 17–19 bp (optimally 17 bp), positioned upstream of the +1 transcription start site.
Q: What three factors determine promoter strength?
A: (1) How well the promoter binds polymerase initially, (2) how efficiently it supports isomerisation from closed to open complex, and (3) how readily the polymerase can escape the promoter.
Q: How does the extended -10 element compensate for the absence of a -35 region?
A: The extended -10 element includes additional sequence upstream of the standard -10 region, providing extra contacts between the polymerase and DNA that replace the contacts normally made by the -35 region.
Q: Name one biological example of a promoter that uses an UP-element, and explain why it needs one.
A: The rRNA gene promoters in E. coli. rRNA genes require very high transcription rates to supply the cell with ribosomes, so the UP-element boosts polymerase recruitment beyond what the -10 and -35 regions alone can achieve.
Q: What role does the discriminator play?
A: The discriminator is a short element just downstream of the -10 region whose interaction with RNA polymerase influences the stability of the enzyme–promoter complex. A stronger discriminator interaction makes the complex more stable.
Sigma factors connect to the broader topic of gene regulation: bacteria swap sigma factors (e.g. σ32 for heat shock, σS for stationary phase) to redirect the polymerase to different sets of promoters under stress. This links to Chapter 14+ material on regulatory networks. Promoter strength also ties into the quantitative side of gene expression, which matters when you reach topics like operon regulation (lac, trp) where promoter architecture is part of the control logic.
sigma factor, sigma-70, σ70, core enzyme, holoenzyme, RNA polymerase holoenzyme, -10 region, Pribnow box, -35 region, consensus sequence, promoter strength, UP-element, extended -10, discriminator, α₂ββ'ω, TATAAT, TTGACA, promoter architecture, gene expression, bacterial transcription, E. coli, gal operon, rRNA promoter, molecular biology chapter 13, UCF molecular biology II