RNA is synthesized from DNA in a process called transcription. In prokaryotes, a single type of RNA polymerase synthesizes all types of RNA. In eukaryotes, multiple RNA polymerases are involved. Transcription involves initiation, elongation, and termination phases. The initial RNA transcript undergoes post-transcriptional processing including 5' capping, polyadenylation, and splicing in eukaryotes to form mature RNA. Alternative splicing allows generation of multiple mRNA isoforms from a single gene.

1. RNA Synthesis
(Transcription)
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2. RNA
- The genetic master plan of an orgm is contained in DNA.
- RNA “working copy” of DNA(where the plan is expressed).
- Transcription is the copying process, in w/c DNA strand serves
as a template for the synthesis of RNA.
 Transcription produces:mRNAs = translated to AAs,&
- rRNAs,tRNAs & other small RNA molecules[aren’t transl
ated= non coding RNAs (ncRNAs)].
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3. Cont…
- Final product of gene expression is protein or RNA.
- transcription is highly selective as compared to repl.
(all regions of DNA won’t be transcribed).
- This selectivity is due to a guide that instruct RNA Pol.where to
start,how often to start & where to stop.
- Regulatory proteins also involve in selection process.
- transcription is in contrast to the “all or none” of replication.
- Post transcriptional modification is its typical feature.
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4. Cont…
- inactive 1o transcriptfunctional form by modification.
- RNA Polymerase: catalayze transcription.
a) In bacteria,only 1-type of RNA Pol(multimeric protein)that
synthesise mRNA, tRNA & rRNA.
b) In Eukaryotes:Several types:-RNA Pol.I,II & III.
- Coding/sense strand & non-coding/anti-sense strand.
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5. Transcription Of Prokaryotic Genes
A) Properties of prok.RNA polymerase.
 Synthesizes all RNAs except primer(by Primase).
 A multi-subunit enzyme
- recognizes promoter region to bind,
- make RNA complimentary copy of DNA &
- recognize the end of transcription.
 RNA is synthesized in 5’-3’ direction.
 It adds G on C, U specifies A instead of T.
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6. Cont…
- The formed RNA is complimentary to the template (antisense)
strand & identical to the coding(sense)strand with U replacing T.
- In DNA both strands can be a template for transcription but for
a given gene only one of the 2 strands has a promoter region.
- Prokaryotic RNA polymerase is a holoenzyme with.
- Core enzyme:4 subunits(a2BB’) & Sigma factor (σ polypeptide)
- it requires dsDNA & sometimes ssDNA as template and
5’-ribonucleoside triphosphate(UTP,ATP,GTP &CTP).
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7. Cont…
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1) Core enzyme:
- Enzyme assembly(2α)
- β’ for template binding
- β has 5’-3’ RNA polymerase activity
- Ω its function is unclear
2) Sigma factor(σ-poly peptide)
- enables RNA pol.to recognize
promoter regions on DNA.
- Primary transcript ? Initial
product of transcription.

8. Steps of transcription
 3 phases of transcription process:
1. Initiation
2. Elongation &
3. Termination.
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Antiparallel,complimentary base pairs b/n DNA& RNA.

9. 1.Initiation
- After the σ-factor recognizes the promoter region the holo-
enzyme binds to it trans.complex initiate transcription.
- Prok’s promoter region has consensus sequence(20-200 bases).
Chxs of a promoter sequence:
a. –35 sequence:A 5'-TTGACA-3',centered 35 bases to the left of
the transcription start site.
- the initial point of contact for the enzymeclosed complex.
- Regulatory sequences:control trans.& found on non template strand.
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10. - -Ve number before a base of a promoter region indicates,it is
found to the left/upstream/(closer to 5’ end)of trans.start site.
- Therefore,the TTGACA sequence is centered at –35.
- The 1st base at the trans.start site is assigned as +1.
b. Pribnow box:-Named after David Pribnow/TATA box
- the 2nd consensus sequence(5'-TATAAT-3')spanned by the
holoenzyme,centered at –10,w/c is the site of initial DNA
melting (unwinding).the T at position 6 is present in any promoter
region; so it is conserved.
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11. Cont…
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Local unwinding of DNA by RNA polymerase and formation of an open initiation complex.
Structure of the prokaryotic promoter region.
Melting of a short stretch (~14 bases) converts the closed complex to
an open onea transcription bubble.

12. 2. Elongation:
- When the holoenzyme unwinds DNAsupercoils(DNA-topoisome-
rase overcome this problem).
- RNA pol.begins to synthesize a transcript of the DNA sequence.
- The elongation phase begins when the transcript (typically
starting with a purine)exceeds 10 ntds in length.
- σ-is then released, & the core enzyme proceeds processively.
- RNA pol:Substrates(ATP,GTP,CTP,UTP) & releases PPi .
- As replication, transcription is always in the 5'→3' direction.
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13. Cont…
- Unlike DNA pol, RNA pol doesn’t require a primer & have no
proofreading activity.
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3. Termination:
- ssRNA elongates until a termination signal isreached.
- 2 types of terminations

14. Cont…
1) ρ-independent/intrinsic/spontaneous:seen prokaryotic genes
- The nascent RNA-generates a sequence i.e. self complimentary.
- RNA folds back on itselfGC-rich stem & loop “hairpin”.
- beyond the hairpin,the RNA transcript contains a string of ‘U’s
at the 3'-endbonding with DNA’s ‘A’sweak linkage
facilitate easy separration of RNA from its template as the
double helix “zips up”behind the RNA pol.
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15. Cont…
2) ρ-dependent:-help of rho(p) protein/requires ATP
- binds a C-rich “rho recognition site” near the 3'-end of the nascent RNA &
chases RNA pol.along RNA force it to pause.
- The ATP dependent helicase activity of ρ-separates the RNA-DNA hybrid
helixthe release of the RNA.
NOTE:some antibiotics work by inhibiting RNA synthesis.
- e.g.,Rifampin(anti-TB):inhibit initiation by binding with β-subunit of prok RNA
polyconformational change.
- but rifampin doesn’t bind to Eukaryotic RNA polymerase.
2. Actinomycine D: used in tumor chemotherapy & works by binding with DNA
template and affect mov’t of RNA pol.
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16. II.Transcription of Eukaryotic Genes
- More complicated than in prokaryotes.
- Involves separate RNA pols.to synthesize rRNA,tRNA& mRNA.
- Also, a large number of proteins called TFs are involved.
- TFs bind on the DNA—in promoter region, close to it, or distal.
- TFs are needed for the assembly of a trans.complex at the
promoter & determine w/c genes are to be transcribed.
- Each eukaryotic RNA pol.has its own promoters & TFs.
- for TFs to recognize & bind to promoter,the chromosome must
be remodeled to increase access.
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17. Cont…
A. Chromatin structure & gene expression
- Euchromatin:relaxed and active form to be transcribed
- Inactive:in condensed form= Heterochromatin
- The interconversion b/n the 2 forms =Chromatin remodeling.
 Formation of Nucleosome affects transcription.
- Acetylation & deacetylation of Lys residue of histone proteins at
N-terminus mediate remodeling.
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18. Cont…
1. Acetylation of Lys residue by Histone acetyl transferase (HATs)
eliminates the +Ve charge on Lys &↓es interaction of Histone with
–Vely charged DNAincrease accessibility.
2. Histone deacetylases(HDACs) remove the acetyl group-favor
strong interaction b/n histone & DNA.
B. RNA polymerases of Eukaryotic Cells:3 Types.
-1.RNA Pol-I: Synthesizes the precursor of 28S,18S& 5.8S rRNA.
- 2. RNA Pol-II:Synthesizes precursor of mRNA & certain small
ncRNAs;like,snRNA.
- 3. RNA Pol-III: synthesizes tRNA,5SrRNA,&some snRNA.
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19. Cont…
a) Promoters & TFs for RNA Pol-II
- ntds are identical with that of Pribnow box but at -25 bases.
- this promoter consensus sequence=TATA or Hogness box.
- CAAT(2nd consensus sequence):at-70to-80ntds from start site.
- In constitutively expressed genes,no TATA box is present;
instead,a GC-rich region (GC box) may be found.
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20. Cont…
- Consensus sequences are on the same DNA strand to be trans-
cribed,hence they are called cis-acting elements.
- Such sequences serve as binding sites for TFs,w/c in turn
interact with each other & with RNA pol.II.
- TFs are encoded by d/t genes & synthesized in cytosol,and act
in nucleus;hence,they are called Trans-acting factors.
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21. Cont…
 Function of TFs:Promoter recognition
- Recruitment of RNA pol.to the promoter&initiate transcription
 RNA Pol-II doesn’t recognize & bind the promoter by itself.
a) TF-IID- recognizes & bind to the TATA box.
b) TF-II F-brings the polymeraseto the promoter.
c) TF-II H:has helicase activity melts the DNA.
- TFs bind outside & inside the promoter sequence to modulate
transcription in response to cell signals like hormones.
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22. Cont…
- Some TFs also bind to proteins (“coactivators”),recruiting them
to the transcription complex.Coactivators:like HAT enzymes.
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Eukaryotic gene promoter
consensus sequences.

23. b. Role of enhancers
 Enhancers are special Cis-acting DNA sequences that ↑es the
transcription initiation rate by RNA pol-II.
- 1) located upstream/5'-side or downstream/3'-side of start site.
- 2)~ thousands of base pairs away from the promoter &
- 3) occur on either strand of the DNA.
 Enhancers contain DNA sequences called “response elements” that
bind specific TFs w/c bound to promoteractivate RNA-pol-II.
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24. Cont…
 Silencers like enhancers act over long distances;but reduce gene
expression.
- α-Amanitin(toxin from mush rooms)/death cap inhibits RNA-pol-
II tight complex with polymeraseinhibit mRNA synthesis.
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Possible locations of enhancers
B. mtRNA polymerase.
- Mitochondria contain a single
RNA pol. closely resembles
bacterial RNA pol.

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26. V. Posttranscriptional Modification Of RNA
- 1o transcript is the initial,linear,RNA copy of a transcription
unit(precursor).
- The precursors of both prokaryotic & eukaryotic tRNA & rRNA
are posttranscriptionally modified by ribonucleases’ cleavage.
- tRNAs further modified to give each species its unique identity.
- In contrast,prokaryotic mRNA is identical to its 1o transcript,but
eukaryotic is extensively modified bothco-&post transcriptionally.
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27. A.rRNA
 Formed from a single precursor pre-rRNA in both Euk & prokaryotes.
- In prokaryotes:23S,16S & 5S rRNA.
- In Eukaryotes:28S,18S & 5.8S rRNA.
- In Eukaryotes 5S is synthesized by RNA Pol-III & modified separately.
- Enzymes for modification:ribonucleases(RNases)& exonucleases.
- RNA synthesis & processing occur in the nucleolus,with base and sugar
modifications facilitated by small nucleolar RNAs (snoRNA).
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Posttranscriptional processing of
eukaryotic rRNA by
ribonucleases

28. B. tRNA
 formed from longer precursor in both Euk&prokmodified
(post transcriptionally).
- Sequences at both ends of the molecule are removed;
- if intron is present,cleaved by nucleases from anticodon loop.
- a–CCA sequence is added by Nucleotidyltransferase at its 3'-
end,and modification of bases at specific positions to produce the
“Unusual bases” chxs of tRNA.
- Modified bases include D(dihydrouracil),ψ(pseudouracil).
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29. Cont…
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Posttranscriptional modification of tRNA-transcript.

30. C. Eukaryotic mRNA
- 1o transcript synthesized in nucleus by RNA Pol-II as hnRNA.
- hnRNApre-mRNA undergooes extensive co-& posttranscriptional
modification in the nucleus w/c includes:
1. 5' “Capping”:the 1st processing rxn on pre-mRNA.
-The cap=7-methylguanosine is attached at 5'-endforms
5'→5'triphosphate linkage.
- Methylation of this terminal guanine occurs in the cytosol by
Guanine-7-methyltransferase by using SAM as methyl donor.
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31. Cont…
- the cap helps stabilize the mRNA & permits initiation of translation.
- Eukaryotic mRNAs lacking the cap aren’t efficiently translated.
2. Addition of a Poly-A tail:40-200 adenine ntds at 3’-end.
- by polyadenylate polymerase & use ATP as a substrate.
- Before adenylation,mRNA is cleaved downstream of a consensus
sequence,polyadenylation signal sequence(AAUAAA) near the 3’-end.
- The tail helps to stabilize mRNA&facilitate its exit from nucleus.
- The tail is shortened gradually in cytosol.
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32. 3. Removal of Introns by splicing process.
- Introns are removed by spliceosome & exons are joined together to
form mature mRNA translation
- A few eukaryotic 1o transcripts contain no introns;histone genes – but
1o transcripts of α-chains of collagen,contain >50 introns.
a) role of snRNAs:are Uracil-rich.
- snRNAs associate with proteinssmall nuclear ribonucleo protein
particles(snRNPs/snurps”designated as U1,U2..mediate splicing.
- introns are removed & exons joined togethermRNA leaves the
nucleus to cytosol through nuclear pore.
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33. Splicing
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34. 4. Alternative splicing of mRNA molecules
- d/t splicing of pre-mRNA in d/t tissues of the same gene d/t
isoforms of one protein(diverse protein formation).
- e.g.,mRNA of tropomyosin protein undergoes extensive tissue-specific
alternative splicing with production of multiple isoforms of it.
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35. 35
 Mutations at splice sitesimproper splicingaberrant proteins.
- Accounts ~15% of all genetic diseases.E.g.,β−thalassemia.

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