RR2: Eukaryotic Transcription Flashcards
What mostly controls gene expression?
73%: rates of transcription.
11%: rates of mRNA degradation
8%: rates of protein degradation
8%: rates of mRNA translation
Do an overview of transcription.
- RNA polymerase recognizes dsDNA.
- DNA double helix locally denatures.
- One strand acts as a template.
- Incoming rNTPs base-pair with bases on the template strand.
- RNA polymerase joins the rNTPs from 5’ to 3’on the new RNA.
DNA is read from 3’ to 5’
Why is polymerization energetically favoured in transcription?
Because the high energy bond between alpha and beta phosphates on the rNTPs are replaced by a lower-energy phosphodiester bond.
Where does transcription start?
At +1.
Is the promoter upstream or downstream of the start site?
It can be either upstream (negative numbers) or downstream (positive numbers)
Is transcription going upstream or downstream?
It’s going downstream of the start sign (in the positive)
What’s a coding sequence?
A region of DNA or RNA whose sequence determines the sequence of amino acids in a protein.
At what rate does RNA Pol 2 go?
1000 to 3000 nucleotides per minute. which is pretty fast but not super fast
What’s the longest human gene and how long does it take to transcribe it once?
Dystrophin (DMD) and it takes a day to transcribe.
What are the 3 stages of transcription?
Initiation
Elongation
Termination
What’s the first stage of the 3 transcription stages and explain.
Initiation
Polymerase binds to the promoter sequence,
locally denatures the DNA,
catalyzes the first phosphodiester linkage.
What’s the second stage of the 3 transcription stages and explain.
Elongation
Polymerase advances 3’ to 5’ down the template strand,
denaturing the DNA and
polymerizing the RNA
What’s the last stage of the 3 transcription stages and explain.
Termination
Polymerase recognizes a stop site, releases the completed RNA and dissociates from DNA.
What are the 3 aspects that are the same in both prokaryotic and eukaryotic transcription?
𝜎 Factors
- 𝜎 Factors confer specificity to RNA polymerase (so it binds to a specific sequence that needs to be transcribed)
regulation of the rate of RNA synthesis
- DNA binding proteins regulate the rate of RNA synthesis by enhancing or impeding RNA polymerase binding to promoter regions
DNA proximal
- Sequences in the DNA proximal to the transcribed gene are critical for efficient transcription
lacZ is a reporter gene, what would need to bind to the promoter to stop lacZ from being trasncribed?
A lac repressor.
What if we wanted to increase the rate of transcription, what would need to happen?
CAP would bind to the CAP site, and then cAMP would bind to CAP.
What are some differences between prokaryotic and eukaryotic transcription?
Prokaryotic transcription has multiple transcription start site, while eukaryotic has only one.
Prokaryotic transcription changes very rapidly because environments control what genes have to be expressed. Those fast changes require allosteric regulation, while eukaryotic transcription isn’t changing rapidly because of the environment and doesn’t use as much allosteric regulation.
Prokaryotes also use polycistronic mechanisms to activate fast transcription. From one mRNA, you can get multiple different proteins.
Eukaryotes are monocistronic.
In Eukaryotes, transcription is used to differentiate cell types. The changes in transcription efficiency and the activation of specific genes that allow us to distinguish the various cells that make up an embryo.
Eukaryotes have 3 types of RNA polymerases.
What do polycistronic and monocistronic mean?
Polycistronic: Describes an mRNA corresponding to multiple genes, so this mRNA can give rise to multiple different proteins.
Monocistronic: Describes an mRNA that has one cistron in a transcription unit, so only one structural gene that codes for one protein.
What is allosteric regulation?
Allosteric: binding to an enzyme and have the ability to modify it.
Allosteric regulation would mean binding to the site and activating or deactivating the enzyme or increasing or decreasing the enzyme activity.
What are the 3 RNA polymerases in eukaryotic transcription?
RNA polymerase 1
RNA polymerase 2
RNA polymerase 3
What are the roles of RNA polymerase 1?
Transcription of pre-rRNA (28S, 18S, 5.8S rRNAs)
Ribosome components.
Protein Synthesis.
What are the roles of RNA polymerase 2?
Transcription of mRNA - snRNA - siRNA - miRNA.
Encodes proteins.
RNA splicing.
Chromatin-mediated repression.
Translation control.
What are the roles of RNA polymerase 3?
Transcription of tRNAs - 5S rRNA - snRNA U6 - 7S RNA - other small stable RNAs.
Protein Synthesis
Ribosome component.
RNA splicing.
Signal recognition particle for insertion of polypeptides into the endoplasmic reticulum.
More unknown functions.
Explain the sensitivity of each RNA polymerase to the mushroom toxin α-amnitin.
RNA Pol 1: High sensitivity, very high RNA synthesis.
RNA Pol 2: Lower sensitivity, but still high RNA synthesis.
RNA Pol 3: Lowest sensitivity, low RNA synthesis.
