Download RNA Synthesis (Transcription)

RNA Synthesis
(Transcription)
Dr. Shumaila Asim
Lecture # 4
1
RNA Synthesis (Transcription)
Introduction
 First step in gene expression
 RNA is involved in the transfer of genetic information stored in DNA to
proteins
 Process of DNA directed RNA synthesis (copying process) (genes in DNA
make RNA copies)
 Synthesis of an RNA strand with a base sequence complementary to one
strand of DNA (template) and identical in sequence to the other strand
2
RNA Synthesis (Transcription)
 Selective process; one particular gene or groups of genes are
transcribed
 Specific sequences indicate the beginning and end of DNA
segment to be transcribed
 Some proteins are also involved in this “selectivity of the process”
3
RNA Synthesis (Transcription)
 Occurs in 5 − 3 direction
 Catalyzed by RNA polymerase I, II & III
(DNA dependent RNA polymerases (RNAPs)
 Each RNAP consists of
large subunits and many small subunits
4
two
Comparison of Replication & Transcription
S.No
1
2
3
4
5
5
Replication
Transcription
DNA template required
DNA template required
Whole genome is copied
Initiation & termination
signals not required
RNA primer is required
Selective parts of DNA
are copied
Initiation & termination
signals required
Primer is not required
Synthesized DNA strand
remains attached to
template strand
RNA does not remain
attached to template
DNA strand
Comparison of Replication & Transcription
S.No
6
Replication
Transcription
6
Deoxy-ribonucleotides are Ribonucleotides are
required
required
7
DNA polymerases
8
Very much accurate (exact) Less accurate
9
No changes required after Post-transcriptional
replication
changes required
RNA polymerases
Numbering
-5 -4 -3 -2 -1 +1 +2 +3 +4 +5 +6
DNA
RNA
A –T – A – C – G – T – T – G – A – C – G ----A – A – C – U – G – C -----
 Position +1 of a gene is the base that is complementary to the
first base of the 5 end of the RNA transcript of that gene
(initiation point)
 Sequences preceding first base are numbered negatively and are
said to be upstream of the initiation point
 Sequences following the first base are numbered positively –
7
downstream
Specific Sequences
a. Promoters – Specific areas or sequences of DNA for initiation
(different for different RNAPs) ,these are
8

Pribnow box (TATA box) (Prokaryotes)

Goldberg-Hogness box or TATA box or consensus sequence (Eukaryotes)
Specific Sequences
b. Terminators – Sequences for termination
c. Enhansers & silencers – Sequences for increasing or decreasing rate of
transcription
d. Transcription Unit – Region between promoter and terminator
(includes sequences for initiation, elongation & termination)
9
Requirements
 Template
=
DNA single strand
 Substrates
=
Ribonucleotides (ATP, GTP, UTP, CTP)
 Initiation factors
 Transcription factors [Tf – II or GTF (General Transcription factors)] (Tf-II
include Tf-IIA, Tf-IIB, Tf-IIC, Tf-IID, Tf-IIE, Tf-IIF, Tf-IIH)
 Tf-IID contains TATA binding protein (TBP) and 14 TAFs
(Transcription activating factors)
 Activators, Co-activators and components of PIC
(pre-initiation complex)
10
Requirements
 Enzymes
=
DNA-dependent RNA polymerases
[RNA polymerase (RNAP) I, II, III]
 RNAP–I synthesizes rRNA
 RNAP–II synthesizes mRNA
 RNAP–III synthesizes tRNA, 5S rRNA
 Each RNAP holoenzyme consists of
Core-enzyme + one or more sigma ()factors
 Sigma () factors
11
the core enzyme to recognize promoter region
assist
Requirements
 RNAP is zinc containing metallo-enzyme, requires
Mg++ ions
for its activity
 Core enzyme consists of two identical -subunits,
,  and  subunits
 -subunit is thought to be the
subunit
(the growing chain of RNA remains
12
attached to this subunit)
one each
catalytic
Generalized Functions of RNAPs
 Unwinding the DNA in front,
 Synthesis of RNA,
 Re-winding it behind,
 Dissociating the growing RNA chain from the template and
 Perform proofreading.
13
RNA Synthesis (Transcription)
14

A cyclic process, involving RNA chain initiation, elongation &
termination

RNA polymerase attaches to and initiates transcription at a
distinct site, the promoter (a special DNA sequence)

GTFs (Tf-II) facilitate promoter-specific binding of RNAPs to
form pre-initiation complex (PIC)

Other proteins (co-activator proteins) regulate the rate of
transcription initiation
RNA Synthesis (Transcription)
15
RNA Synthesis (Transcription)
16