Question

Describe the stages used to transcribe an mRNA molecule in great detail.

Answer 1

Transcription is a process of mRNA synthesis from DNA molecules, where RNA polymerase plays a key role. During the process RNA polymerase uses the template strand (antisense strand or non coding strand) of DNA and synthesizes a complementary RNA molecule using ribonucleoside triphosphates (ATP, CTP, GTP, UTP) as precursors. The RNA produced has the same sequence as the nontemplate strand, which is known to be coding strand or sense strand except that the RNA contains U instead of T. The ultimate prokaryotic transcription occurs in three phases: 1) initiation, 2) elongation and 3)termination.

1 Initiation stage has been subdevided into 4 steps

a. Promoter Recognition by RNA polymerase holoenzyme () through $\sigma$ factor

b. Binding of holoenzyme to the promoter upstream of the DNA containing two conserved promoter elements, the –10 sequence (TATA box/Pribnow box) with the consensus TATAAT and the –35 sequence with the consensus TTGACA. These promoter sequences which ar e highly conserved between species.

c. Formation of open promoter complexo

d. Formation of first phosphodiester bond of nasent RNA by RNA polymerase holoenzyme. The first nucleotide in the RNA transcript is usually pppG or pppA.

2. Elongation stage has been subdevided into 2 steps

a. Release of $\sigma$ factor: After transcription initiation, the $\sigma$ factor is released from the transcriptional
complex to leave the core enzyme ( $\alpha 2\beta \beta '$ ) which continues elongation of the mRNA transcript.

b. Direction of transcription: The core enzyme contains the catalytic site for polymerization, probably within the $\beta$ subunit. The RNA polymerase then synthesizes RNA in the 5’ →3’ direction, involving the four ribonucleoside triphosphates as precursors. The nucleophilic attack at 3-OH at the end of the growing RNA chain binds to the phosphate group of the incoming ribonucleoside triphosphate to form a phosphodiester bond. A ternary complex is formed by DNA template, RNA polymerase, and new RNA transcript. The portion of unwound DNA is undergoing transcription is called the transcription bubble. The RNA transcript forms a RNA–DNA hybrid (10 - 12 nucleotides long) with DNA template strand. As transcription proceeds the hybrid then moves away from the DNA . Further DNA is unwound ahead of the transcription bubble and the DNA rewinds after the transcription complex has passed. This process continues until a termination sequence is reached.

3. Termination stage: The most common termination signal sequence is a GC-rich with a inverted repeat sequence, followed by an AT-rich region in the DNA. The RNA makes self complementary stem loop or hairpin structure from the DNA inverted repeat sequence followed by few or more U residues. The ultimate termination process has been subdevided into 2 parts.

a. Rho dependent termination: In this process, Rho (hexameric protein) help to recognize the termination site and detach the nascent transcript from DNA template, thereby stop transcription.

b. Rho independent termination: In absence of Rho protein, transcript can be detach from the DNA template due to weak A-U bonds between DNA and RNA ahead of hairpin structure.