In this post I’m going to show you how DNA transcription occurred in prokaryotic cell.
(step by step)
Before we dive into the transcription process you have to understand the basic meaning of transcription. Have you ever met any transcriber? Or maybe you transcribe something.
I’m sure, you all done transcription in your life. Let me explain with simple example. You borrow or took class notes and then rewrote them neatly with color pens to help you review right.
That’s it, this is the basic meaning of transcription, rewrite or copying the existing information.
cells do the same thing but with the help of several enzymes, in a more specialized and accurate way.
Let’s take a look on content which we are going to discussing further.
- RNA polymerase
- Stages of DNA transcription
As you know DNA is the genetic material of almost every living object. It stored genetic information in the form of nucleotide sequences.
This stored information only becomes useful when it is expressed in the production of RNA and proteins (you can say that transcription and translation).
You can consider that; the main function of DNA is to store genetic information for a long time. But RNA is the only molecule that acts as storage and the transition of information in catalytic activity. Because some RNA virus has RNA as genetic material.
what is transcription in molecular biology?
Transcription is the process in which it copying out the particular segment of DNA sequence in the similar alphabet of RNA (especially mRNA).
let’s take overview of an essential enzyme in RNA synthesis RNA polymerase enzyme
2) RNA polymerase.
RNA polymerase is the main enzyme which is used in the synthesis of RNA.
In prokaryotes (Bacteria), a single RNA polymerase transcribes all genes (More precisely RNA polymerase II).
the mechanism of RNA synthesis is closely related to DNA polymerase.
DNA polymerase add dNTPs to 3’- OH in 5’ – 3’ direction. RNA polymerase also adds the ribonucleotide in the same 5’ – 3’ direction.
3) Stapes of transcription:
As DNA replication, transcription also take place in three stages
Here we will see how this step are happene in prokaryotic cell step by step.
I) Initiation of transcription
To begin transcribing a gene, RNA should bind with a specific region of the DNA gene called the promoter region. The promoter gives the signal to polymerase were to “sit down” on the DNA and begin transcribing.
Binding of RNA polymerase to promoters.
The initial binding of the RNA polymerase with the transcriptional promoters is a nonspecific association with DNA. It is relatively weak compared with the binding of specific promoters.
The role of this nonspecific binding and the mechanism by which the enzyme moves towards specific promoter is not fully understood yet.
On the other hand, the transcription factor sigma is most important factor for initiation process. It helps RNA polymerase to find specific promoter.
In these 70 nucleotides there are two segments which are strongly conserved known as consensus sequence.
As described in figure promoter site have two of the particular note this is a sequence of six bases. These are the segment of the promoter where RNA polymerase binds. The TTGACA sequence is about 35 base pairs before (upstream) the transcription starting point. And TATAAT sequence called the Pribnow box, also known as the TATA box, usually about 10 base pairs upstream of the transcriptional start site.
These regions are called the -35 and -10 sites, respectively because these are their approximate distance in nucleotide upstream from the first nucleotide to be transcribed, it is known as +1 site.
That we discussed above is the sites and components which are present in the promoter site, now we are going to discuss how RNA polymerase bind right promoter
As you see in the above figure the promoter site has its -35 and -10 region. RNA polymerase moves along with DNA from non-specific binding sites to promoter sites.
Here, when it reaches the promoter site, the sigma factor first recognizes the -35 region and allowing holoenzyme to “settle down” on that region of the promoter. This binding complex is known as a closed complex, in which bound DNA is intact.
Now, it is bind with AT-rich -10 region, this complex known as open complex. In which DNA is intact but partially unwound. Here, sigma factor and proteins in the core enzyme undergo confidential changes that cause the DNA strand separate at -10 region. Because of the unwinding of the DNA bubble-like structure formation occur, this is known as the transcription bubble.
Transcription is initiated within the complex, leading the confidential change that converts the complex to the elongation form. Form here, transcription is entered in the elongation stage.
II) Elongation of transcription:
Once RNA polymerase is strongly positioned at promoter and form a transcription bubble, the next step of transcription “elongation” can begin.
Now, sigma factor dissociates polymerase enters in the elongation phase of transcription.
As you see in the figure, Elongation will start with the addition of the first ribonucleotide at the transcription start site.
RNA polymerase does not require primer like DNA polymerase to add nucleotide. But the reaction catalyzed by RNA polymerase is quite similar to that catalyzed by DNA polymerase. It uses all four nucleotides for the synthesis of RNA complementary to the DNA.
For each nucleotide in the DNA templet, RNA polymerase adds a matching (complementary) ribonucleotide to the 3’ end of the RNA strand. With the addition of a nucleotide, a pyrophosphate is produced as ribonucleotide monophosphate and is incorporated into the growing RNA strand.
Then released pyrophosphate is hydrolyzed to fuel the process. The newly transcript RNA is nearly identical to the coding strand of the DNA. As elongation of the mRNA continues, single-stranded mRNA is released, and two strands of DNA behind the transcription bubble resume their double-helical structure, as you can see in the diagram.
However, RNA strand has the base uracil(U) in the place of thymine(T), as well as a slightly different sugar in the nucleotide.
As you can see in the diagram below, each T of the coding strand is replaced with a U in the newly transcript RNA strand.
But if mistakenly wrong nucleotide was added by RNA polymerase, it holds the proses go back and cleave that nucleotide from the sequence and replace it with the right one.
III) Termination of transcription:
Termination of transcription occurs when the core RNA polymerase dissociates from the template DNA. Studies that occurs in prokaryotes have demonstrated that the termination event occurs by at least two mechanisms.
- Rho-independent termination.
- Rho-dependent termination.
1. Rho independent termination.
Rho-independent termination depends on specific sequences in the DNA template strand. As the RNA polymerase approaches the end of the gene being to transcribe, it hits a region rich in G and C nucleotides.
As you see in figure, these regions result formation of GC- rich region in the transcript, this region are able to base-pair into a ‘hairpin’ or “stem loop” like structure.
Such loop, which typically contains seven to ten G:C base pair, that causes RNA polymerase to pause at A-rich region of the DNA template. The A-U base pair holding the DNA and RNA together in the transcription bubble is too weak to hold RNA: DNA hybrid together and RNA polymerase falls off.
As a result, RNA strand is release from the DNA template.
2. Rho dependent termination:
The second kind of terminator is termed as Rho-dependent termination because it requires the aid of protein. The best-studied termination factor in Prokaryotes is the Rho factor (ρ). Rho factor can be involved in the transcription of all types of genes, but the action of this factor best studied for protein-coding genes.
As shown in the figure, the current model proposes that the Rho factor binds to mRNA at a site called rut-utilization site (labeled with yellow in figure). Once Rho factor bind with rut site in RNA then it moves toward the 3’ end, following the RNA polymerase.
But for this movement Rho factor requires energy, Rho factor uses energy supplied by ATP hydrolysis to move along the mRNA, as it tries to catch up with RNA polymerase. However, Rho’s rate of movement is slower than that of RNA polymerase.
Thus, Rho factor can only catch up with RNA polymerase, if the RNA polymerase pauses at a Rho-dependent pause site. When this pause occurs, the Rho factor catches up with RHA polymerase and causes RNA polymerase to dissociate from DNA.
Rho factor is known to have RNA: DNA hybrid helicase activity. This activity leads to the unwinding of the mRNA -DNA complex and release of RNA polymerase from the template DNA strand.
At the end of termination released RNA polymerase binds with other promoter sequence and start transcribing DNA strand. And newly synthesized mRNA continues protein synthesis (translation).
Now it’s turns on you.
Let us know if we miss something, leave comment below if any suggestion or you want next topic in your interest, we will defiantly look on it.
Now give us answer of this simple question in comment section.
What is gene expression? And in which site of DNA ‘transcription’ take place?