Topic 6-L1- Transcription Flashcards
DNA encodes genes, but itself is inert. For DNA sequences to carry
out functions, it must be converted to
RNA via transcription
Transcription uses an enzyme called
RNA polymerase
RNA polymerase works by
- Binds to DNA template and makes an RNA copy of one of two strands
- Copied strand = coding strand, other strand = template strand
- Only builds RNAs 5’ - 3’ – Because transcription goes in only one
direction and in opposite directions on each strand
During transcription, Coding strand matches RNA sequence…
except T is Template strand replaced with U
RNA polymerase core enzyme made up of 4 subunits:
⍺ (2 copies) β β’ ω. Holoenzyme also includes additional subunit, (sigma factor)
Sigma factor in holoenzyme binds
promoter region, then dissociates from core enzyme
Core enzyme unwinds DNA to expose template —>
forms transcription bubble
Using NTPs (ATP, CTP, GTP, UTP) as substrates and the template strand as guide, the
RNA chain is built one nucleotide at a time
Ultimately, RNA polymerase will encounter a transcriptional
terminator and will dissociate from the template & release the RNA
Transcription will (generally) continue until RNA polymerase (RNAP) encounters a
transcriptional terminator. RNAP then dissociates from DNA, stops making RNA & releases transcript.
Intrinsic (rho-independent) terminators form when
RNA hairpin structures form, followed by a string of “U” residues. U residues act as pause signal for RNAP – formation of hairpin forces RNAP off template.
Rho-dependent terminators: A
protein called
Rho binds RNA as it is being transcribed and causes RNA polymerase to dissociate after it encounters certain sequences
Transcriptional initiation is guided by DNA sequences called
promoters
Whether or not a sequence acts as a promotor
promoter is active dictated by binding of sigma factors & the activity of regulatory proteins
The housekeeping (most commonly used/most important) sigma factor is called
SIGMA70 (or RpoD) – it recognizes two sequences upstream of the
transcriptional start site.
Bacterial promoters
- Transcriptional start site: sequence ~35 bp upstream of the +1 site
- Pribnow box : ~10 bp upstream of the +1 site. This sequence (TATAAT)
There are 3 major classes of RNAs (and other non-coding RNAs with
a range of functions – often regulatory):
- Messenger RNA (mRNA): converted to protein via translation
- Transfer RNA (tRNA): functional RNAs, used in translation process
- Ribosomal RNA (rRNA): functional RNAs, used in translation process
Open reading frames (ORFs) are sequences that are
translated into proteins
mRNAs contain both ORFs and untranslated regions (UTRs) - which are?
parts of the mRNA transcript that are not translated into protein
mRNAs that encodes multiple ORFs are _________ and are called _______
polycistronic operons
Genes in an operon are
cotranscribed
5’UTR –
everything from first transcribed residue (+1) through the start codon of gene. Contains ribosome binding site (RBS)
ORF
Start codon (e.g. ATG) through stop codon (e.g. TAA)
3’ UTR
everything from the stop codon of the gene through the final transcribed residue. Often contains transcriptional terminator sequences
transcription is different in eukaryotes since they
- Have 3 RNA polymerases. RNA polymerase II produces mRNA (encode proteins). Others encode RNA transcripts that are not translated (e.g. rRNA, tRNA, and other RNAs)
- Are more complex = 12+ subunits
- Require transcription factors to recognize promoters – they bind specific DNA sequences & recruit RNA polymerase (e.g TATA box)
Do eukaryotes use operons?
NO, each gene has its own promoter
Eukaryotic primary mRNA transcripts contain
protein-coding exons and non-
coding introns. mRNAs are processed (spliced) to remove introns
Eukaryotic mRNAs are
polyadenylated at the end (3’) of transcript and capped at 5’ end using a modified G residues attached using an unusual linkage
Where does transcription and translation occur in eukaryotes
transcription in nucleus
translation in cytoplasm
The archaeal RNA polymerase resembles
RNA polymerase II from eukaryotes and has 11-13 subunits.
Archaea RNA polymerase is recruited to
promoters using transcription factors (related to eukaryotic proteins)
Do archaea use TATA boxes and transcriptional factors that bind these elements and recruit RNA polymerase?
Yes
Are archaea more complicated then eukaryotes
NO, eukaryotes more complex
Archaea do not have :
- 5’ cap,
- poly A tail, mRNAs
- introns & are not spliced
- nucleus – transcription and translation often coupled (like in bacteria)
Do archaea not use operons like eukaryotes?
No they do use operons like bacteria
multiple genes encoded by one RNA (single promoter controls expression of several genes)
