Lec 20 Flashcards
Prokaryotic vs. Eukaryotic Transcription
Prokaryotic:
- Polycistronic mRNA
- no introns (usually)
- One RNA polymerase
Eukaryotes:
- Monocistronic (usually)
- introns
- 3 RNA polymerase
Transcription: Synthesis of RNA
- DNA used as template
- No primer needed
- rNTPs (no thymine, uracial pairs with adenine)
- Nucleotides are added 5’-3’
- Two phosphate groups are cleaved from the incoming rNTP as the nucleotide is added
- RNA polymerase joins the nucleotides together
What RNAs are transcribed in both prokaryotes and eukaryotes
mRNA
rRNA
tRNA
What RNAs are only produced in eukaryotes
pre-mRNA snRNA snoRNA scRNA miRNA siRNA piRNA
What are other names for contemplate DNA strand
sense strand or coding strand
TorF:Different genes can be transcribed from different strands of the same DNA segments
True
Transcription unit
is segment of DNA that codes for an RNA molecule and the sequences necessary for its transcription
Transcription
process of making RNA from DNA (5’-3’)
what joins nucleotides together in transcription?
phosphodiester bonds
RNA polymerase
enzyme that joins nucleotide together
Template strand
strand of DNA that is transcribed
coding strand
other DNA strand: sequence is the same as the RNA transcribed from the template strand except for T/U difference
Upstream
sequences after the start of transcription
Downstream
sequences after the start of transcription
where does transcription start at?
+1 nucleotide
What are the four stages of transcription? and which stages require holoenzyme or core enzyme
- template bind (holoenzyme)
- chain initiation (holoenzyme)
- chain elongation (core enzyme)
- chain termination (core enzyme)
holoenzyme vs core enzyme
holoenzyme contains the sigma factor
What are the control regions located in the Promotor?
Pribnow Box- 5’-TATAAT-3’
-35 Control Region 5’- TTGACA-3’
Consensus Sequence?
a sequence that describes the nucleotides most often present in a segment of interest
Y=pyrimidine
R=Purine
N=any
Which strand are consensus sequences located on?
coding strand
Template binding and initiation
holoenzyme bind to promoter forming closed complex(has to happen in the proper place)
-the DNA strands separate forming open complex
After template binding and initiation what happens?
Sigma dissociates (forming core enzyme) after about 9-12 nucleotides are joined to form RNA -Elongation occurs 5'-3' using only coreenzyme
What is transcription reaction catalyzed by
RNA polymerase
Direct vs. Inverted repeats
Direct repeats- sequences are the same on either side
inverted repeats- same sequences but are the same but backwards at the diagonal location
-leads to hairpin formation on the RNA during transcription
READ 5’-3’
Termination
May or May not require RHO
- inverted repeats are present on the DNA sequence near termination and are transcribed
- these form hairpin structures in the RNA that can signal termination of transcription
- The hairpins are followed by a STRING OF ADENINE RESIDUE ON THE TEMPLATE STRAND OF THE DNA. The A-U bonds are relatively weak and allow dissociation of the DNA-RNA pairing (allows two DNA strands to come back together)
-Hairpin causes RNA polymerases to pause
Termination using RHO
The inverted repeats and adenine residue sequence are still present on DNA and still transcribed
- RHO binds to the RNA and moves toward the 3’ ned and RNA polymerase
- RNA polymerase pauses at termination hairpin
- Rho has helicase activity that causes the DNA-RNA hybrid to unwind and transcription ends
Prokaryotic mRNA:Shine-Dingrano sequence
- found in the leader region (not translated)
- is important for ribosome binding
- the sequence is about 7 nuts upstream from the start of translation
Polycistronic mRNA
1 RNA molecule is produced, but it contains the information for more than one gene and is translated into more than 1 gene product
Transcription in Eukaryotes: 3 RNA polymerases
RNA polymerase I- large rRNA (28s, 18s, 5.8s)
RNA polymerase II- mRNA, most snRNA, snoRNA, miRNA
RNA polymerase III- small rRNA (5s), tRNA, some nRNA
-recognize different promoters, different transcription factors and different termination factors
hnRNA
heterogeneous RNA=pre-mRNA=precursor for mRNA= primary transcript for mRNA
Where does transcription occur in eukaryotes
nucleus
Does mRNA processing occur in eukaryotes or in prokaryotes?
eukaryotes
What transcribes the transcription unit of rRNA genes in Eukaryotes?
RNA polymerase I
NTS
non-transcribed spacer- contains control region, promoter, and terminators
ETS
external transcribed space- at ends of transcription unit
ITS
internal transcribed spacer- within transcription unit
rDNA transcription unit
transcribed into 1 pre-rRNA molecule then undergoes processing
cis Elements
- nucleotide sequence that are close to the coding region of a gene and help the cell to determine when the gene should be transcribed
- binding spots for proteins (or RNAs)
trans-acting factors
- proteins of RNAs from other genes
- attach to cis-elements or other transcription factors
- Recruit RNA polymerase
What does RNA polymerase II transcribe?
pre-mRNA, pre-mRNA, and some snRNA
TATA Box
- promoter element
- near -25
- consensus sequence read off non-template strand is 5’-TATAAA-3’
- Part of the core promoter-important in initiation of tx
CAAT Box
- promoter element
- near -80
- important of initiation of TX
- consensus 5’-GGCCAATCT-3’
GC box
Promoter element
- may have more than 1 and location varies
- help RNA polymerase bind near sit of TX (transcription)
- consensus sequence 5’-GGGCGG-3’
Octamer BOx
promoter element
- consensus 5’-ATTTGCAT-3’
- helps RNA polymerase bind properly in initiation
- location varies
Enhancers
more distant element NOT a promoter element
-required for maximum transcription
-can function in either direction and from a “distance”, but on same piece of DNA transcriptional activators bind enhancers to help them interact with transcription factors at the start site for TX
(CIS ACTING)
Core promoter
immediately upstream of gene and where the basal transcription apparatus binds. Has TATA box
Regulatory promoter
immediately upstream of core promoter. Variety of consensus sequences (Octamer box, GC box, CAAT box)
Elongation in Eukaryotes
after -30 BP are synthesized, RNA polymerase leaves the promoter and begins elongation
- Many transcription factors stay bound at the promoter to reinitiate transcription
- transcription bubble contains about 8bp of DNA-RNA hybrid
- The structure of the RNA polymerase causes seperation of the newly formed RNA strand from the DNA template Strand
Initiation Eukaryotes
?
RatI exonuclease
required for termination of transcription
-once RNA is cleaved the RatI binds and degrades the trailing RNA
What are the 3 processes of pre-mRNA to form mRNA
- removal of introns
- polyA tail added to 3’ end
- capping of 5’ end
RNA polymerase III in eukaryotes
transcribes tRNA, 5s rRNA, and most snRNAs
- promoter is located within the gene for the RNA
- 2 conserved “boxes” that are important for proper transcription