# Transcription bubble

A transcription bubble is a molecular structure formed during DNA transcription when a limited portion of the DNA double helix is unwound, providing enough space for the RNA polymerase to bind to the template strand and begin RNA synthesis. The transcription bubble size is usually 12 to 14 base pairs, which allows the incorporation of complementary RNA nucleotides by the enzyme with ease. Dynamics and structure of the transcription bubble are variable, and play a role in the regulation of gene expression at the transcriptional level.

## Formation and structure

The transcription bubble is formed when RNA polymerase binds to a promoter site and unwinds a small portion of the DNA double helix. This exposes a single-stranded DNA segment, allowing RNA to be synthesized using it as a template. In both eukaryotes and prokaryotes, multiple transcription start sites have been observed within the same promoter, and transcription bubble dynamics—such as expansion ("scrunching") and contraction ("unscrunching")—have been shown to play a role in the positioning of these variable transcription start sites to the RNA polymerase active site. As such, the structure of the transcription bubble plays a role in regulating gene expression through mediating the creation of different transcripts.
Molecular dynamic simulations have found that the lifetime of the transcription bubbles are sequence-dependent, and longer bubble lifetimes are associated with A-T rich core promoter sequences. The formation and maintenance of the transcription bubble is also likely temperature dependent: temperature analyses on E.coli DNA suggest that the complex is formed at 37°C and collapses at lower temperatures. These temperatures may vary depending on species. In conjunction with temperature, the presence of magnesium ions (Mg2+) alongside an increase in temperature causes the unwinding of transcription bubbles further downstream up to a base position of +2, which correlates with the start of RNA synthesis.

## RNA polymerase

The bacterial RNA polymerase, a leading enzyme involved in formation of a transcription bubble, uses DNA template to guide RNA synthesis. It is present in two main forms: as a core enzyme, when it is inactive, and as a holoenzyme, when it is activated. A sigma (σ) factor is a subunit that assists the process of transcription and it stabilizes the transcription bubble when it binds to unpaired bases. These two components, RNA polymerase and sigma factor, when paired together, build RNA polymerase holoenzyme which is then in its active form and ready to bind to a promoter and initiate DNA transcription. Once it binds to the DNA, RNA polymerase turns from a closed to an open complex, forming the transcription bubble. RNA polymerase synthesizes the new RNA in the 5' to 3' direction by adding complementary bases to the 3' end of a new strand. The holoenzyme composition dissociates after transcription initiation, where the σ factor disengages the complex and the RNA polymerase, in its core form, slides along the DNA molecule.

## Transcription cycle of bacterial RNA polymerase

The Escherichia coli transcription bubble may be initiated at variable locations between the promoters and the transcription start site, this in turn may alter the length and sequence of the resulting transcript. During the formation of the transcription bubble in E.coli and most other bacteria, the RNA polymerase holoenzyme (RNAP) binds to a promoter of an exposed DNA strand in a process mediated by sigma (σ) initiation factors factors. The double helix DNA is unwound and a short nucleotide sequence is made accessible on each strand. The transcription bubble is formed as a region of unpaired bases on one of the exposed DNA strands. The DNA is unwound and single-stranded at the start site, the location of RNAP binding. The DNA promoter interaction is interrupted as the RNA polymerase moves down the template DNA strand and the σ factor is released. Once the σ factor dissociates from the RNA polymerase, the transcription continues. About 10 synthesized nucleotides of a new RNA strand are required for this to proceed to the elongation step. Elongation occurs quickly until the RNA polymerase comes across a termination signal (terminator) which arrests the process and causes the release of both the DNA template and the new RNA molecule. The DNA usually encodes the termination signal. In E.coli, the process of transcription termination via dissociation of the RNA polymerase have been found to depend on 3 possible mechanisms: an interaction between the polymerase and an intrinsic terminator sequence, the RNA-dependent termination factor Rho, and the ATP-dependent DNA translocase Mfd.
Factors affecting the transcription cycle in E. coli include the universally conserved transcription factor NusG, which slows RNA cleavage through the inhibition of backtracking.

## Eukaryotic transcription

The majority of eukaryotic genes are transcribed by RNA polymerase II, proceeding in the 5' to 3' direction. In eukaryotes, specific subunits within the RNA polymerase II complex allow it to carry out multiple functions. General transcription factors help binding RNA polymerase II to DNA. Promoters are sites where RNA polymerase II binds to start transcription and, in eukaryotes, transcription starting point is positioned at +1 nucleotide. Like all RNA polymerases, it travels along the template DNA, in the 3' to 5' direction and synthesizes a new RNA strand in the 5' to 3' direction, by adding new bases to the 3' end of the new RNA. A transcription bubble occurs as a result of the double stranded DNA unwinding. After about 25 base pairs of the DNA double strand are unwound, RNA synthesis takes place within the transcription bubble region. Supercoiling is also part of this process since DNA regions in front of the RNA polymerase II are unwinding, while DNA regions behind it are rewinding, forming a double helix again.
The RNA polymerase carries out the majority of the steps during the transcription cycle, especially in maintaining the transcription bubble open for the complementary base pairing. There are some steps of the transcription cycle that require more proteins, such as the Rpb4/7 complex and the RNA polymerase attached to the elongation factor transcription factor IIS (TFIIS).