Transcription (contd)

• DNA template strand = Antisense , negative sense

• DNA coding strand = Sense , positive sense
• Consensus Sequence = A consensus sequence is a sequence
of DNA, RNA, or protein that represents aligned, related
sequences. These sequences are recognized by sigma factor .
Egs are pribnow box and -35 sequence
• The Pribnow box is a sequence of TATAAT of six nucleotides
that is an essential part of a promoter site on DNA for
transcription to occur in bacteria.

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Pribnow box
• Located at -10 from the transcription start site (+1).
• Site of melting (unwinding) of a short stretch ( ¬14 bp) of DNA
• Melting forms transcription bubble
-35 Sequence
• Consensus sequence 5′-TTGACA-3′
• 35 bases to the left of transcription start site
• Initial point of contact for holoenzyme
• A base is assigned negative number if it occurs prior to
transcription start site
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Rho Factor(p)
• The rho factor is a protein which recognizes and binds to termination sequences
in the 3′ portion of the RNA. This binding destabilizes the polymerase-template
interaction leading to dissociation of the polymerase and termination of
transcription.
• It is a hexameric ATPase with helicase activity. Separates RNA-DNA hybrid helix
causing release of RNA
• P independent termination
• Some sequences of nascent RNA are self complementary & RNA folds back on
itself forming GC rich stem and a loop ( hairpin ). Beyond hair pin RNA transcript
contains string of U’s at 3′ end. The formation of the stem-loop in the RNA
destabilizes the association between polymerase and the DNA template. This is
further destabilized by the weaker nature of the AU base pairs that are formed,
between the template and the RNA, following the stem-loop. This leads to
dissociation of polymerase and termination of transcription.

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Antibiotics which inhibit RNA synthesis
• Rifampin ( treats tuberculosis)
• Binds with beta subunit of prokaryotic RNA pol and prevent
chain growth beyond 3 nucleotides
• Dactinomycin (actinomycin D)
• Treats tumors
• Inserts (intercalate) between DNA bases inhibit transcription
• Initiation and elongation both inhibit

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Post-transcriptional Processing of RNAs
• Eukaryotic RNAs (all three classes) undergo significant processing, some of which occurs co-
transcriptionally and some post-transcriptionally
1) mRNA 5′-End Capping
• The capping process occurs after the newly synthesizing mRNA is around 20–30 bases long, at
which point RNA pol II pauses on the template.The 5′ ends of nearly all eukaryotic mRNAs are
capped with a unique 5′ → 5′ triphosphate linkage to a 7-methylguanosine residue. Synthesis of
the mRNA cap structure is catalyzed by RNA guanylyltransferase and 5′-phosphatase
• The capped end of the mRNA is protected from exonucleases & is recognized by specific
proteins of the translational machinery.
2) mRNA 3′-End Polyadenylation
• Almost all mammalian mRNAs are polyadenylated at the 3′-end.
• A specific sequence, AAUAAA, is the primary poyadenylation signal sequence . Polyadenylate
polymerase is the enzyme involved. 40-250 adenosine monophosphates are attached .Tailing
helps to stabilize mRNA and aid in translation.
3) Splicing
• Maturation of eukaryotic mRNA requires removal of RNA sequences that do not code for proteins (
introns or intervening sequences)
• The remaining coding sequences ( exons ) are joined together
• Process of removing introns and joining exons is called splicing
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