Transcription definition steps of transcription general structure of gene RNA polymerase structure Transcription in prokaryotes in detail (initiation, elongation and termination)

1. TRANSCRIPTION
By : Hassam Zafar

2. TRANSCRIPTION DEFINITION
It may be defined as the process in which RNA is synthesized using DNA as a template is known as
TRASNCRIPTION. The DNA is said to be transcribed into RNA, and the RNA is called a transcript.

3. Fig 1. DNA strands used as templates for transcription.
The direction of transcription is always the same for any gene and starts
from the 3 end of the template and the 5 end of the RNA transcript.
Hence genes transcribed in different directions use opposite strands of
the DNA as templates.

4. Fig. 2: Overview of transcription. (a) Transcription of two genes in opposite directions. Genes 1 and 2 from Figure 1 are
shown. Gene 1 is transcribed from the bottom strand. The RNA polymerase migrates to the left, reading the template strand
in a 3-to-5 direction and synthesizing RNA in a 5-to-3 direction. Gene 2 is transcribed in the opposite direction, to the right,
because the top strand is the template. As transcription proceeds, the 5 end of the RNA is displaced from the template as the
transcription bubble closes behind the polymerase. (b) As gene 1 is transcribed, the phosphate group on the 5 end of the
entering ribonucleotide (U) attaches to the 3 end of the growing RNA chain.

5. Stages of Transcription and what is the need of its stages
• The protein-coding sequence in a gene is a relatively small segment of DNA embedded in a much longer DNA
molecule (the chromosome).
Q. How is the appropriate segment transcribed into a single-stranded RNA molecule of correct length and nucleotide
sequence?
• Because the DNA of a chromosome is a continuous unit, the transcriptional machinery must be directed to the start of a
gene to begin transcribing at the right place, continue transcribing the length of the gene, and finally stop transcribing
at the other end. These three distinct stages of transcription are called initiation, elongation, and termination.
• Although the overall process of transcription is remarkably similar in prokaryotes and eukaryotes, there are important
differences. For this reason, we will follow the three stages first in prokaryotes and then in eukaryotes.

6. Fig: 4 Components of RNA polymeraseFig: 3 Simple structure of gene

7. Steps of transcription in prokaryotes
1. Initiation: Transcription begins with the binding of the RNA pol holoenzyme to a region of the DNA
known as the promoter, which is not transcribed. The prokaryotic promoter contains characteristic
consensus sequences (Fig. 5). [Note: Consensus sequences are idealized sequences in which the base
shown at each position is the base most frequently (but not necessarily always) encountered at that
position.] Those that are recognized by prokaryotic RNA pol σ factors include the following.
Fig. 5: Transcription initiation in prokaryotes

8. a. –35 Sequence:
A consensus sequence (5′-TTGACA-3′), centered about 35 bases to the left of the transcription start site (see
Fig. 6), is the initial point of contact for the holoenzyme, and a closed complex is formed. The first base at the
transcription start site is assigned a position of +1. There is no base designated “0”.
b. Pribnow box:
The holoenzyme moves and covers a second consensus sequence (5′-TATAAT-3′), centered at about -10,
which is the site of melting (unwinding) of a short stretch (~14 base pairs) of DNA.
Fig: 6. Structure of the prokaryotic promoter region. T = thymine; G = guanine; A = adenine; C = cytosine.

9. 2. Elongation: Once the promoter has been recognized and bound by the holoenzyme, local unwinding of the DNA
helix continues (Fig: 7), mediated by the polymerase.
Note: Unwinding generates supercoils in the DNA that can be relieved by DNA topoisomerases.
The elongation phase begins when the transcript (typically starting with a purine) exceeds 10 nucleotides in length.
Sigma is then released, and the core enzyme is able to leave (clear) the promoter and move along the template strand
in a processive manner, serving as its own sliding clamp.
Fig: 7. Local unwinding of DNA by RNA polymerase and formation of an open initiation
complex (transcription bubble).

11. 3. Termination:
The elongation of the single-stranded RNA chain continues until a termination signal is reached. Termination
can be intrinsic (occur without additional proteins) or dependent upon the participation of a protein known as the ρ
(rho) factor.
a. ρ-Independent:
Seen with most prokaryotic genes, this requires that a sequence in the DNA template generates a sequence in
the nascent (newly made) RNA that is self-complementary (Fig 7 & 8). This allows the RNA to fold back on itself,
forming a GC-rich stem (stabilized by hydrogen bonds) plus a loop. This structure is known as a “hairpin.”
Additionally, just beyond the hairpin, the RNA transcript contains a string of Us at the 3′-end. The bonding of these Us
to the complementary As of the DNA template is weak. This facilitates the separation of the newly synthesized RNA
from its DNA template, as the double helix “zips up” behind the RNA pol.

12. Figure 7 & 8: Rho-independent termination of prokaryotic transcription. A. DNA template sequence generates a
self-complementary sequence in the nascent RNA. B. Hairpin structure formed by the RNA. N represents a Non
complementary base; A = adenine, T = thymine; G = guanine; C = cytosine; U = uracil.

13. b. ρ-Dependent:
This requires the participation of the additional protein rho, which is a hexameric ATPase with helicase
activity. Rho binds a C-rich rho utilization (rut) site near the 5′-end of the nascent RNA and, using its ATPase activity,
moves along the RNA until it reaches the RNA pol paused at the termination site. The ATP-dependent helicase activity of
rho separates the RNA–DNA hybrid helix, causing the release of the RNA.
Fig 9: Rho dependent termination.