RNA and transcription Flashcards
What is a UTR on a gene?
What is the order leading to a protein?
The 5′ untranslated region (UTR) is a regulatory region of DNA situated at the 5′ end of all protein-coding genes that is transcribed into mRNA but not translated into protein.
start and end of transcription
They are mRNA domains that control critical post-transcriptional gene regulation processes.
Genome DNA
unspliced mRNA
Mature mRNA
Protein (tends to be a polyA tail at the end)
translated needs to be multiple of 3
The protein part is the only part without the UTR as they are not translated.
What is a gene
a gene is a region of DNA that codes for an active molecule whether that becomes a protein or not.
it’s not single units, there are genes between genes and within genes and between exons.
You can also have many different transcripts due to alternative splicing
What did whole-genome analysis reveal about genes
whole-genome analyses revealed significant findings:
- Transcription is pervasive, with more than 85% of the human genome being transcribed.
- The distinction between genes and intergenic space is less apparent, and about 70% of human genes are transcribed from both strands.
- Coding DNA accounts for less than one-quarter of the highly conserved fraction of the genome. This indicates the presence of many more functionally important noncoding DNA sequences than previously expected.
- Intensive analyses of mammalian transcriptomes uncovered many transcripts of unknown function, including conserved regulatory sequences and noncoding RNAs (ncRNAs) within or spanning introns of known genes.
- The blurring of gene boundaries at the transcript level is illustrated by complex patterns of transcription, where both ncRNA and protein-coding transcripts overlap.
The passage emphasizes that recent discoveries challenge the traditional, simple idea of a gene and highlight the complex and dynamic nature of gene organization and regulation.
Transcription by DNA-dependent RNA polymerase
DNA-dependent RNA polymerases
synthesises mRNA 5’→3’
using riboNT, does not
need RNA primer
transcription is about which genes are active at which times in certain tissues at certain conditions.
many cells may have same code and structure but transcription is different and the genes expressed.
It creates an RNA strand but it depends on DNA
Describe and explain transcription in Bacteria?
Initiation:
RNA polymerase, the enzyme responsible for transcription, recognizes and binds to the promoter region of DNA.
The promoter sequence typically includes a specific region known as the -10 box (TATAAT) and the -35 box.
Formation of the Transcription Bubble:
RNA polymerase unwinds a short stretch of DNA, forming a transcription bubble.
The enzyme starts synthesizing an RNA strand complementary to the template DNA strand.
Elongation:
RNA polymerase moves along the DNA template in the 3’ to 5’ direction, synthesizing an RNA strand in the 5’ to 3’ direction.
The growing RNA chain contains nucleotides complementary to the template DNA.
Termination:
Intrinsic termination: Transcription stops when a terminator sequence is transcribed. This forms a hairpin structure in the RNA, causing RNA polymerase to dissociate.
Rho-dependent termination: Requires the Rho protein to recognize a specific sequence and cause RNA polymerase to detach.
Describe the structure of an RNA polymerase
Describe it within the process of transcription
RNA polymerase :
two α, β, β’ (and ⍵) + σ factor = holoenzyme
(α2ββ’ = core enzyme)
In bacteria, there is a single type of RNA polymerase responsible for transcribing all types of RNA.
Core Enzyme:
The core enzyme of bacterial RNA polymerase consists of multiple subunits. The primary subunits include α (alpha), β (beta), β’ (beta prime), and ω (omega).
The α subunits are involved in enzyme assembly and regulatory interactions.
The β subunit is part of the DNA clamp and is a target for certain antibiotics.
The β’ subunit forms part of the DNA clamp and provides a conserved domain essential for catalysis.
The ω subunit facilitates the folding of the β’ subunit and helps in its recruitment to the core enzyme.
Holoenzyme Formation:
The core enzyme can associate with a σ (sigma) factor to form the holoenzyme, which is involved in the recognition of promoter sequences on DNA. σ factor provides DNA-binding specificity.
Different sigma factors can associate with the core enzyme, allowing the bacterial RNA polymerase to recognize different promoter sequences under various conditions.e.g σ32 is generally for “heat-shock (number is molecular weight)
Initiation:
The holoenzyme binds to the (closed) promoter region on DNA, recognizing specific promoter sequences.
The sigma factor helps in the formation of the transcription bubble, where the DNA strands separate to allow RNA synthesis.
Once transcription begins the sigma factor leaves as you don’t need recognition factor anymore
Elongation:
The RNA polymerase holoenzyme moves along the DNA template in the 3’ to 5’ direction, synthesizing an RNA strand in the 5’ to 3’ direction.
The enzyme reads the DNA template and incorporates complementary ribonucleotides into the growing RNA chain.
Termination:
Transcription can terminate at specific terminator sequences on the DNA.
There are different termination mechanisms, including intrinsic termination, where a hairpin structure in the RNA causes dissociation, and Rho-dependent termination, which involves the Rho protein.
Transcription: how does the RNA polymerase recognise where to start transcribing in bacteria?
Why is the distance for each promotor vital?
Start of transcription is different to the start of translation
Promoter in bacteria: -35 and -10 sequences (start of transcription: +1)
+1 start of transcription (usually a purine, A or G) (you need experimental data to find this)
-10 sequences (consensus=most common sequence) TATAAT) minus means going left (it needs to be -10 and similar to TATAAT but not necessarily)
-35 sequence (consensus TTGACA)
RNA synthesizes 5’ to 3’
RNA polymerase only uses coding strand not a template strand so RNA sequence will be the complement of the promoters
-10 from from the start of transcription leads to you to roughly the middle of the consensus and then between the -10 prometer and the -35 consensus there is roughly 15-20 bases between them
why is the distance so important
- Recognition by Sigma Factor:
Sigma factor recognizes and binds to promoter elements.
Positions RNA polymerase for transcription initiation. - Melting of DNA Strands:
Promoter elements facilitate DNA strand unwinding.
Allows RNA polymerase access to the template strand.
3.Determination of Transcription Start Site:
Promoter elements, especially the -10 box, define the start site.
Ensures accurate RNA synthesis with the correct sequence.
4.Efficiency of Transcription Initiation:
Conserved promoter sequences ensure efficient initiation.
Promoters closely adhering to consensus sequences are more efficient in RNA polymerase binding and initiation.
Where is the start of translation
It needs to be to the right (downstream)
Translation starts downstream of the transcription start site, and it is initiated at the Shine-Dalgarno sequence (ribosome-binding site) and the start codon. The Shine-Dalgarno sequence is typically located a -8 bases before the start codon, which is usually AU(T)G.
You need this because the ribosome is complementary to the sequence in the ribosome
You are looking for the start of transcription to see what sequence corresponds going from
UTR becomes the sequence before the translation promoter
on the downstream right there needs to be a stop coding to tell us where there is the end of transcription and translation and UTR
What are the differences in transcription in bacteria and eukaryotes:
explain each polymerase
Bacteria have one RNA polymerase.
Eukaryotes have three RNA polymerases:
* RNA pol I to transcribe rRNA
* RNA pol II to transcribe mRNA
* RNA pol III to transcribe tRNA
and much more complex promoters
Eukaryotic promoters
promoter are
-20/-30
-50/-70
-90
They also have enhancers which can be upstream and downstream which shows complex regulation
activator protein help activate transcription by folding DNA and (connecting to something)
TF =transcription factor e.g TFIIa (eukaryotic TF) helps activate transcription
There are huge protein complexes for transcription
Post-transcriptional modifications in bacteria (mRNA)
mRNA in bacteria
they often have polycistronic MRNA means they have an mRNA can make more than one protein
often 3 proteins are needed at the same time this could be useful, as proteins needed at the same time to make the process more efficient
There can also be the addition of a polyA tail=signal of degradation of mRNA opposite in eukarya
Post-transcriptional modifications in eukarya
3 main types of post transcriptional modification
*addition of poly(A) tail and these stabilize the mRNA (opposite than bacterial) Polyadenylation
removal of non-coding introns by splicing
addition of 5’ capping structure
What is capping
5’ capping is the addition of 7-methylguanosine. This structure is added near the beginning of transcription. It pauses to stick this to mRNA as its coming out to make it more stable so it prevents degradation by 5’ to 3’ exonucleases
a number of viruses (poliovirus) targets cap structure to take over host and steals the cap from the mRNA so it can protect and stabilise its own mRNA.
Polyadenylation in bacteria and eukarya
*addition of poly(A) tail and these stabilise the mRNA (opposite than bacterial) Polyadenylation
Splicing
spliceosome =nuclear multiprotein complex
small nuclear ribonucleoproteins contain (snRNP’s) to help recognise specific NT sequences at the exon/intron boundaries
it cuts in two positions, removes the introns and binds the two exons (need ligase for this)
