2. The synthesis of RNA molecules using DNA strands as the
templates so that the genetic information can be transferred
from DNA to RNA.
TRANSCRIPTION
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3. TRANSCRIPTION Gene – functional unit of DNA that can be transcribed.
Template/ Non- coding strand/ antisense strand – DNA strand that
produces working copies of RNA.
Non- Template/Coding strand/ Sense strand – DNA strand that does
not participate in transcription.
Ribonucleotides used
Uracil instead of thymine used
Transcription is selective – due to some inbuilt signals in on the DNA
Primary Transcript (inactive) – product formed in Transcription
Post Transcriptional Modifications- primary transcripts undergo
modifications like splicing, terminal additions, base modifications etc
Reading is 3’ to 5’ direction, Synthesis is from 5’ to 3’ direction
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4. TRANSCRIPTION There are four major types of RNA molecules:
Messenger RNA (mRNA) encodes the amino acid sequence of a
polypeptide
Transfer RNA (tRNA) brings amino acids to ribosomes during
translation.
Ribosomal RNA (rRNA) combines with proteins to form a ribosome,
the catalyst for translation.
Small nuclear RNA (snRNA) combines with proteins to form
complexes used in eukaryotic RNA processing.
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5. TRANSCRIPTION The Transcription Process - RNA Synthesis
1. Transcription, or gene expression, is regulated by gene
regulatory elements associated with each gene.
2. DNA unwinds in the region next to the gene.
3. RNA is transcribed 5’-to-3’. The template DNA strand is read 3’-to-
5’. Its complementary DNA, the nontemplate strand, has the same
polarity as the RNA.
4. RNA polymerization is similar to DNA synthesis, except:
a.The precursors are NTPs (not dNTPs).
b. No primer is needed to initiate synthesis.
d. Uracil is inserted instead of thymine.
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6. zTemplate
The template strand is the strand from which the RNA is
actually transcribed. It is also termed as antisense strand.
The coding strand is the strand whose base sequence
specifies the amino acid sequence of the encoded protein.
Therefore, it is also called as sense strand.
5 '
3'
G C A G T A C A T G T C
C G T C A T G T A C A G
3 '
5 '
c o d i n g
s t r a n d
t e m p l a t e
s t r a n d
transcription
R N AG C A G U A C A U G U C5' 3'
TRANSCRIPTION
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7. TRANSCRIPTION
Both processes use DNA as the template.
Phosphodiester bonds are formed in both cases.
Both synthesis directions are from 5´ to 3´.
Similarity between replication and transcription
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8. TRANSCRIPTION
replication transcription
template double strands single strand
substrate dNTP NTP
primer yes no
Enzyme DNA polymerase RNA polymerase
product dsDNA ssRNA
base pair A-T, G-C A-U, T-A, G-C
Differences between replication and transcription
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9. TRANSCRIPTION
PROKARYOTES
RNA POLYMERASE
A single enzyme DNA dependent RNA
polymerase / RNA polymerase synthesizes
all the RNA in prokaryotes.
Complex Holoenzyme
mol. Wt. 465 kDa.
Five subunits - 2α, 1β , 1β’ with 1sigma
(σ)factor.
Enzyme without sigma factor is referred to as
core enzyme (α2ββ’)
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12. TRANSCRIPTION
PROKARYOTES
The specific region on the DNA where the enzyme binds is known as
promoter region.
There are two base sequences on the coding DNA strand which the
sigma factor of RNA polymerase can recognize for initiation of
transcription
Pribnow box (TATA box) : This consists of 6 nucleotide bases (TATAAT), located
on the left side about 10 bases away (upstream) from the starting point of
transcription.
The ‘–35’ sequence : This is the second recognition site in the promoter region
of DNA. It contains a base sequence TTGACA, which is located about 35 bases
(upstream, hence –35) away on the left side from the site of transcription start.
INITIATION
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16. TRANSCRIPTION
PROKARYOTES
RNA polymerase recognizes the promoter region.
the sigma factor is released & transcription proceeds.
RNA is synthesized from 5’ end to 3’ end antiparallel to the DNA
template using (ATP, GTP, CTP and UTP).
For the addition of each nucleotide to the growing chain, a
pyrophosphate moiety is released.
The sequence of nucleotide bases in the mRNA is complementary to
the template DNA strand.
this enzyme does not possess endo- or exonuclease activity.
No proof-reading activity
The problem of supercoils is overcome by topoisomerases
ELONGATION16
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17. TRANSCRIPTION
PROKARYOTES
1. Rho (ρ) dependent termination :
A specific protein, named ρ factor, binds to the growing RNA
(and not to RNA polymerase)
or weakly to DNA, and in the bound state it acts as ATPase and
terminates transcription and releases RNA.
The ρ factor is also responsible for the dissociation of RNA
polymerase from DNA.
TERMINATION17
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19. TRANSCRIPTION
PROKARYOTES
2. Rho (ρ) independent termination :
The termination in this case is brought about by the formation of
hairpins of newly synthesized RNA.
This occurs due to the presence of palindromes.
The presence of palindromes in the base sequence of DNA
template (same when read in opposite direction) in the
termination region is known.
As a result of this, the newly synthesized RNA folds to form
hairpins (due to complementary base pairing) that cause
termination of transcription.
TERMINATION19
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21. TRANSCRIPTION
EUKARYOTES
RNA POLYMERASES
1. RNA polymerase I is responsible for the synthesis of precursors
for the large ribosomal RNAs.
2. RNA polymerase II synthesizes the precursors for mRNAs and
small nuclear RNAs.
3. RNA polymerase III participates in the formation of tRNAs and
small ribosomal RNAs.
4. Mitochondrial RNA polymerase in eukaryotes resembles
prokaryotic RNA polymerase in structure and function.
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22. TRANSCRIPTION
EUKARYOTES
PROMOTER SITES
Hogness box (or TATA box)- is located on the left about 25
nucleotides away (upstream) from the starting site of mRNA
synthesis.
CAAT box - another site of recognition between 70 and 80
nucleotides upstream from the start of transcription.
One of these two sites (or sometimes both) helps RNA polymerase II
to recognize the requisite sequence on DNA for transcription.
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23. TRANSCRIPTION
EUKARYOTES
INITIATION
The molecular events are complex - involve three stages.
1. Chromatin containing the promoter sequence made accessible to the
transcription machinery.
2. Binding of transcription factors (TFs -(TFIID, TFIIA, TFIIB, TFIIF, TFIIE, TFIIH))
to DNA sequences in the promoter region. The TFs bind to each other, and in turn
to the enzyme RNA polymerase.
3. Stimulation of transcription by enhancers.
Enhancer can increase gene expression by about 100 fold. Enhancers bind to
transcription factors to form activators.
It is believed that the chromatin forms a loop that allows the promoter and
enhancer to be close together in space to facilitate transcription.
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24. TRANSCRIPTION
EUKARYOTES
Heterogeneous nuclear RNA (hnRNA)
The primary mRNA transcript produced by RNA
polymerase II in eukaryotes is often referred to as
heterogeneous nuclear RNA (hnRNA).
This is then processed to produce mRNA needed for
protein synthesis.
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25. TRANSCRIPTION
POSTTRANSCRIPTIONALMODIFICATIONS
POST TRANSCRIPTIONAL MODIFICATIONS
The RNAs produced during transcription are called primary transcripts.
They undergo many alterations—terminal base additions, base
modifications, splicing etc.,
This process is required to convert the RNAs into the active forms.
Ribonucleases, are responsible for the processing of tRNAs and rRNAs
of both prokaryotes and eukaryotes.
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27. TRANSCRIPTION
POSTTRANSCRIPTIONALMODIFICATIONS
MESSENGER RNA (mRNA)
1. The 5’ capping : capped with 7-methylguanosine by an unusual 5’→5’ triphosphate
linkage. S-Adenosylmethionine is the donor of methyl group. This cap is required for
translation, besides stabilizing the structure of mRNA.
2. Poly-A tail : A large number of eukaryotic mRNAs possess an adenine nucleotide
chain at the 3’-end. This poly-A tail, as such, is not produced during transcription. It is
later added to stabilize mRNA. However, poly-A chain gets reduced as the mRNA enters
cytosol.
3. Introns and their removal : Introns are the intervening nucleotide sequences in
mRNA which do not code for proteins. The removal of introns is promoted by small
nuclear ribonucleoprotein particles (snRNPs). snRNPs in turn, are formed by the
association of small nuclear RNA (snRNA) with proteins.
On the other hand, exons of mRNA possess genetic code and are responsible for
protein synthesis.
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29. TRANSCRIPTION
POSTTRANSCRIPTIONALMODIFICATIONS
Splicing
Splicing is done by spliceosomes.
It consists of the primary mRNA transcript , five snRNAs (U1, U2,
U4, U5, U6 and many proteins.
This complex -Small nuclear ribonucleoprotein complex (snurps).
The splicing starts from 5’- end of exon-intron junction.
5’ end of intron undergoes nucleophilic attack.
Intron forms a loop or lariat. Second cut is made at 3’ and of intron.
Ligation of 3’ end of exon-1 with 5’ end of exon-2 is done.
Intron is digested.
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31. TRANSCRIPTION
POSTTRANSCRIPTIONALMODIFICATIONS
Alternative splicing
The processing of mRNA is also a site for regulation of gene
expression.
By selective splicing and altering donor site, alternative splicing is
done.
Different mRNAs from the same primary transcript formed.
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32. TRANSCRIPTION
POSTTRANSCRIPTIONALMODIFICATIONS
Transfer RNA
Transfer RNA
All the tRNAs of prokaryotes and eukaryotes undergo post-transcriptional
modification.
Trimming,
converting the existing bases into unusual ones- Modification of
bases A,U,G and C - methylation, reduction, deamination and
rearranged glycosidic bonds.
addition of CCA nucleotides to 3’ terminal end of tRNAs by
nucleotidyl transferase .
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34. TRANSCRIPTION Inhibitors of RNA synthesis
Actinomycin D and Mitomycin intercalate with two GpC bp of DNA and
inhibits RNAsynthesis.
Rifampicin – TB drug binds to β-subunit of RNA polymerase which is
inactivated.
α-amanitin is a toxin from mushroom which inactivates RNAP II.
3-deoxy adenosine is a synthetic analog that causes chain termination.
Thiolutin, a sulfur based microbial antibiotic is an RNA polymerase
inhibitor.
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35. TRANSCRIPTION Reverse transcriptase
Retrovirus is a group of RNA viruses. e.g AIDS virus.
RNA dependent DNA polymerase (reverse transcriptase) synthesize a
new DNA strand.
RNA is degraded by RNAase H.
Another strand of DNA- using the DNA strand –to form dsDNA
Reverse transcriptase inhibitors as drugs in the treatment of AIDS.
Such as zidovudine , lamivudine and tenofovir.
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