Molecular biology

DNA Transcription – RNA Synthesis

The process of DNA transcription also referred to as RNA synthesis , is the process where genetic information in DNA is transformed...

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This article writter by MN Editors on December 28, 2021

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DNA Transcription - RNA Synthesis
DNA Transcription - RNA Synthesis

Definition of DNA Transcription 

The process of DNA transcription also referred to as RNA synthesis , is the process where genetic information in DNA is transformed into messenger (mRNA) (mRNA) through an enzyme known as RNA polymerase.

The mRNA synthesized is then transported out of the cell’s nucleus to assist in the production of proteins via the mechanism of translation. The regulation of mRNA production within the nucleus, the cells automatically regulates the rate of expression of genes.

It is supported by the RNA polymerase enzyme that copies the correct sequences of DNA to create a complementary RNA replica that carries the genes. The fundamental mechanism for transcription is the same for both prokaryotes and eukaryotes, however, it can differ in several ways from them.

Definition of DNA Transcription 
Image Source: khanacademy.org

DNA Transcription Enzymes and Function

The most important enzyme for DNA transcription is the RNA polymerase. In prokaryotes there is a specific type of RNA-based polymerase employed, whereas in eukaryotes, three kinds of RNA polymerases are utilized i.e three types of RNA polymerase: I III, II and.

RNA Polymerase
RNA Polymerase | Image Source: Khan Academy.

The main roles of RNA polymerase are:

  • The initiator complex is formed that aids in unwinding DNA’s double-helical structure of DNA
  • Synthesis and elongation of transcript of the RNA is accomplished by adding nucleotide bases Adenine (A), Cytosine (C), Guanine (G) as well as Uracil (U).
  • It creates the termination sequences that stop and stop transcription.

DNA Transcription Steps

50 transcription factors from different proteins are bound to the promoter sites located at the five-foot end of the gene that is to be translated. The RNA polymerase bonds to this transcription factor complex permitting the double helix of DNA to expand. The RNA polymerase is then able to read one strand from the 3′-5 directions. This is the case in cells of eukaryotes the nucleosome is located in the growing of RNA polymerase (Pol II) is responsible for the protein-coding genes.

The transcription factors and RNA polymerase complex is a replacement for the nucleosome once DNA has been transscribed as well as Pol II has moved on. As the RNA polymerase travels along the DNA strand it is able to assemble ribonucleotides into an string of RNA by making use of triphosphate (ATP) Each ribonucleotide gets introduced into the RNA strand that is growing through an insertion process known as base-pairing i.e every Cytosine (C) is connected to the guanine (G) while the Uracil (U) Uracil(U) is connected with an Adenine (A).

The synthesis of RNA takes place in the 5′-3 direction. When each nucleoside triphosphate has been added to the 3′-end of the growing DNA strand both terminal phosphates get removed. Once transcription has come to its end, transcription gets taken away from the polymerase as well as the enzyme polymerase also gets removed from DNA.

DNA Transcription Process

In prokaryotic cells the whole process of transcription is described in three phases: Initiation Elongation, and Ending. When the process comes to closing process in prokaryotes the mRNA that is formed is now in a position to be translated. As opposed to eukaryotes termination, an immature version of mRNA is created, and as such it is necessary to go through additional processes to make an mRNA that can then be transformed into proteins.

In general, the process of transcription converts DNA into mRNA, the type of RNA which contains the information required for the production of proteins. In eukaryotes there are two main stages that happen during transcription:

  1. Pre-messenger RNA formation using an RNA polymerase enzyme. 
  2. Editing of pre-messenger RNA by splicing

Pre-messengerger RNA creation involves beginning, elongation and terminating phases, which are completed with the formation of an mRNA. The mRNA goes through various phases of splicing before forming mature mRNA.

Formation of pre-messenger RNA

DNA Transcription Initiation.
DNA Transcription Initiation. Image Source: Khan Academy.

As transcription proceeds DNA needs to unravel, assisted by RNA polymerase that catalyzes the process. When transcription is initiated, only one strand of DNA is transscribed, the one with the sequence that initiates transcription. This strand is called the sense strand. the other strand is referred to as the antisense strand.

The mRNA that’s transcribed usually a duplicate that of the sense-strand however, it’s the antisense strand which is being transcribed. The triphosphate ribonucleoside (NTPs) is aligned to the antisense DNA line by pairing bases, after which the RNA polymerase joins two ribonucleotides to form an RNA molecule that is pre-messenger, that is akin to a specific area on the antisense strand.

Transcription is complete when the enzyme RNA polymerase detects the triplet of bases as an end signal. At this point the DNA molecule reverses to create a double Helix. The creation of the messenger RNA (mRNA) takes place in three phases The three stages are: Initiation, Elongation and finally, termination.

Initiation

Promoter and initiation in prokaryotes

The transcription process is controlled by a specific location called the promoter. The promoter serves as the place where RNA polymerase binds and the promoter directs the polymerase to be placed on DNA to trigger transcription. The enzyme RNA polymerase which catalyzes the process of transcription.

Promoter and initiation in prokaryotes
Promoter and initiation in prokaryotes | Image Source: Khan Academy.

This enzyme features an Sigma (s) factor which is the dissociative unit which allows the enzyme to detect the sequence of promoters (starting the transcription) that is located between -35 to -10 regions. It is detected by the holoenzyme subunit of RNA polymerase that binds to and moving across the DNA template. It forms a closed promoter complex. The DNA molecules could contain multiple promoter sequences, and open promoter complexes.

The promoter that is bound to transcription factors and the polymerase, RNA, forms an intricate complex. They are proteins that regulate which regulate the rate of transcription. When the RNA polymerase becomes attached by the sequence of promoter it denatures the DNA duplex in a localized manner creating an the promoter complex that is open, which then becomes the unwound portion of the double-stranded DNA. This exposes the bases on the DNA’s two strands.

Promoter and initiation in Eukaryotes

In eukaryotes the RNA polymerase doesn’t directly join to the promoter’s sequence like in prokaryotes. A helper promoter, also known as the basic (general) transcription factor is able to bind to the promoter before, which assists the RNA polymerase in attaching with the DNA-based template. Eukaryotes possess a promoter sequence known as the TATA box. It is recognized by transcription factors that eventually allow binding to the polymerase. The TATA box contains a lot of As and Ts, making it simple to tear the DNA strands apart.

Promoter and initiation in Eukaryotes
Promoter and initiation in Eukaryotes

Elongation

When transcription is initiated after which it is then that the transcription begins and the sigma (s) factor separates from the polymerase. Template strands are read from the 3′-5 direction, which implies that RNA synthesis occurs in the 5′-3′ direction, with Nucleoside Triphosphate (NTPs) serving as substrates to the enzyme.

The second strand of DNA templates is referred to as the coding side due to it has the same base sequence. newly created mRNA is similar to that apart from the substitution of thiamine by the base uracil. The RNA polymerase is responsible for the formation of a phosphodiester bonds between adjacent Ribonucleotides.

Elongation
Elongation

The energy required by polymerase comes from breaking the triphosphate with high energy into monophosphate and releasing diphosphates that are inorganic (PPi). A transcription bubble forms and must be maintained since transcription occurs within the DNA double-strand template. The bubble moves across the DNA duplex as it elongates. Pausing or stalling are typical and later are essential to stop transcription.

Termination

It is the method that ends transcription. This is initiated when signaled by stop sequence, also known as the terminator sequence. The transcription of the terminator sequence. The RNA polymerase is then able to release the DNA temple that unravels and returns to the double-helical structure.

Termination in bacteria

There are two methods of termination for bacteria

Rho-dependent termination

This is the process of ending that occurs when the RNA molecule is home to the binding site of the protein called the Rho factor. This protein is able to bind to DNA sequences. It begins to ascend the transcript until it reaches the RNA polymerase, and then reaches an area called the transcription bubble. In the bubble, the Rho factor is able to pull the RNA transcript as well as the DNA template strands apart and releases the RNA molecule and completing this transcription. A stop-point sequence in transcription which is later discovered in the DNA triggers the RNA polymerase to cease and let to allow the Rho factor to catch up and end the process.

Rho-dependent termination
Rho-dependent termination

Rho-independent termination

The process is based on a particular sequence that is found in the DNA template. When transcription is taking place, as the RNA polymerase reaches the point at which the gene which is being transcribed and reaches an area abundant of Cytosine (C) along with Guanine(G). The RNA transcribed from this region folds back onto itself in a manner that the C and G bonds together creating a hairpin that is stable and causes the RNA polymerase to slow down. The hairpin is then followed by an Uracil (U) within the RNA terminator that is complements the template DNA Adenine (A). The U-A region is an insignificant interaction to the template DNA as well as with the RNA polymerase that is stalled creates an unstable environment that allows the enzyme to break down and disappear from the new transcript of RNA.

Rho-independent termination
Rho-independent termination

Pre-translational mRNA processing

In eukaryotes the mRNA which has been translated is known as pre-mRNA. Therefore it needs to undergo additional procedures to develop into mature MRNA. These are referred to as pre-translational mRNA process. They are:

5′ Capping

This is due to the adding of methylated guanine caps at the 5′ end of the mRNA. The cap 5′ aids to recognize the mRNA molecules by ribosomes, as well as to shield the immature mRNA from destruction of RNases.

Polyadenylation

This is the adding of an adenosine monophosphate (A) tail that is attached to the 3′-end of the mRNA. Its the poly (A) tail comprised of several molecules of adenosine monphosphate, which stabilizes RNA, as a result of its inherent instabilities.

Splicing

  • This is the coding of a single genetic sequence that is used for a variety of proteins, preserving this genetic information.
  • The procedure includes:
    • The removal of non-coding sequences referred to as introns through spliceosome removal.
    • Connecting the coding sequences known as exons through the process of ligation.
    • Splicing is a sequence-dependent process, which means it is a part of the transcript.
    • This allows a variety of proteins to be created by a single pre-mRNA
  • After the splicing process the mature mRNA has been created.
  • The mature mRNA is then the messenger that permits the synthesis of proteins to take place.
  • Mature mRNA contains opened Reading Frames (ORF), which is a region where it gets transformed into proteins. The translation process for the ORFs occurs in three blocks comprising three nucleotides, referred to as codons.
  • The 5 three- and five-digit ends , there are untranslated regions (UTRs) that aren’t transliterated during the process of protein synthesizing.

DNA Transcription in Eukaryotes (Difference from prokaryotes)

Transcription in eukaryotes as well as prokaryotes are similar, but with some differentiating features.

Similarities

Common similarities include;

  • DNA serves as the template for both organisms.
  • The RNA polymerase is a key enzyme that is responsible for the whole process within both organisms
  • The RNA molecule is the ultimate product of both organisms.
  • The chemical structure for the transcript identical in both the organisms.

Differences

The most significant differences in transcription of eukaryotes and prokaryotes are:

1. Promoter

Prokaryotes are composed of three promoters i.e promoters of -10, -35 and downstream elements. Eukaryotes are awash with a variety of promotors i.e The TATA Box, the initiator element, downstream core promoters, CAAT box, and CG box.

2. RNA polymerase

Prokaryotes have a specific kind of RNA polymerase that assists in the production of the DNA string. Eukaryotes contain three kinds of RNA polymerases I II and III that aid in the synthesis of the RNA strand.

3. Initiation

The initative complex of eukaryotes comprised of a variety of transcription factors, which dissociate upon the completion of the initiation process. Although prokaryotes are not part of an initiator complex, they do form an initiator.

4. Transcription and translation concurrence

Another significant difference is that in prokaryotes the processes of transcription and translation take place simultaneously, whereas in eukaryotes transcription must be completed before the translation process can be activated. The RNA found in eukaryotes undergoes post-transcriptional modifications such as capping, polyadenylation and splicing to produce an mature mRNA which proceeds to translation. These processes don’t occur in prokaryotes.

5. RNA genetics

Prokaryotes’ mRNA has multiple genes that are in a single mRNA. This is why they are called polycistronic. Eukaryotes have one gene that is located on a single molecule of mRNA therefore referred to as monocistronic.

6. Termination

Prokaryotes’ termination is assisted by Rho dependent or Rho independent factors, whereas in eukaryotes transcription is ended through the poly-adenylated (A) signal as well as an downstream terminator sequence.

Reverse Transcription

This is the process of converting DNA to RNA, through which RNA serves as a template during the creation of a particular type of DNA, referred to as complementary DNA (cDNA). The central dogma outlines the methods involved in DNA synthesizing (replication) as well as the synthesis of RNA (transcription) as well as protein production (translation) and cDNA production (reverse transcription). Thus DNA encodes for the RNA gene and RNA codes for proteins and RNA may encode DNA when it comes to reverse transcription.

RNA encoded viruses go through reverse transcription, that allows their genomic RNA to be converted into DNA with the help of a the reverse transcriptase enzyme. The reverse transcription process is also referred to by the name retrotranscription, or retrotras. It is a wildly incorrect procedure that could lead to mutations that may cause resistance. The transformation to RNA into DNA typically used in labs mostly as a diagnostic tool for the majority of RNA viruses, such as HIV and hepatitis, influenza coronaviruses and others.

Transcription Inhibitors

Transcription inhibitors are used to hinder the mechanism and action of the enzyme RNA polymerase that hinders this process. The use of transcription inhibitors is primarily to stop the transcriptional mechanisms of bacteria for pathogens that cause disease. The most frequently employed inhibitors are.

  • A-amanitin – This is a inhibitor made from yeast which is a specific inhibitor of the RNA polymerase II enzyme and RNA polymerase III.
  • Rifampicin blocks transcription in bacterial cells by inhibiting DNA-dependent polymerase through attaching itself to beta-subunit.
  • 8-hydroxyquinoline is also a antifungal transcription inhibitor
  • Other examples are actinomycin D CDK9 inhibitors like DRB and flavopiridol triptolide.
  • A mechanism that inhibits transcription is the methylation of histones that blocks the activity of transcription.
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Microbiology Notes is an educational niche blog related to microbiology (bacteriology, virology, parasitology, mycology, immunology, molecular biology, biochemistry, etc.) and different branches of biology.

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