module 6 :( Flashcards
why is gene expression simpler in prokaryotes?
- in prokaryotes: transcription –> translation
- in eukaryotes : transcription –> RNA processing (mRNA) –> translation
Which forms of RNA are found in both prokaryotes and eukaryotes?
- mRNA, tRNA, rRNA
Why is RNAs structure different from DNA?
- has uracil instead of thymine
- usually single stranded but has a complex secondary structure: hair pin loops and stem loops
- RNA is more reactive since the 2’ end has an OH (DNA has an H)
Is all DNA synthesized into RNA?
no! Transcription is selective synthesis of RNA
Does initiation require a primer? What are the next steps after initiator?
- Does not require a primer
- ribonucleotides are added to the 3’ OH group of the growing RNA strand
- DNA unwinds at the head of the transcription bubble and then rewinds!
What are the 3 requirements for transcription?
- DNA template
- Ribonucleotides
- RNA polymerase and other proteins
What is the substrate for RNA synthesis called?
- ribonucleoside 5’ triphosphate
- triphosphate - sugar - base (A,G, U,C)
Does RNA have a specific template it uses for synthesis?
Nope! It can use either DNA strand as a template strand
- synthesizes in the 5’ to 3’ direction, and the DNA template is read in the 3’ to 5’ direction
What is the transcription unit?
- the unit that is transcribed as well as the sequences necessary for transcription
- includes the promoter, the RNA coding region, and the termination site (RNA coding region is the only part that is transcribed)
What is the promoter (for transcription)?
- region of DNA that is recognized and bound by transcription apparatus (RNA polymerase and other proteins)
- RNA polymerase binding to promoter region orients the enzyme towards the start site
What are the 3 steps to transcription in prokaryotes?
- initiation: RNA polymerase and transcription apparatus binds to the promoter region and begins synthesizing RNA
- elongation: DNA is winded through RNA polymerase, unwinded and RNA nucleotides are added to the 3’ end of RNA strand
- termination: stop sequence reached
Describe prokaryotic RNA polymerase
- holoenzyme is the entire enzyme complex composed of core polymerase and the sigma factor
- sigma factor: many types found in bacteria which aid in recognizing many classes of promoters
- core RNA polymerase: initiates transcription
What are sigma factors?
- many types of sigma factors found in prokaryotes, aid in recognizing many classes of promoter regions
- without the sigma factor, the core enzyme would initiate transcription randomly
What is a consensus sequence?
- promoters contain short sequences that are conserved among most promoters known as consensus sequences (most common nucleotides)
- most commonly at -10 (pribnow box) and -35 nucleotides from the start site
- binding of transcription apparatus to these sites orients it towards the start site
are -35 and -10 the only consensus sequences for all promoters?
no! it’s common but not identical in all promoters: each is a variation in a theme
- variation affects strength of the promoter {stronger = frequency of transcription)
What are down mutations and up mutations?
- up mutations: mutations that make the sequence more similar to consensus sequence = greater rate of transcription
- down mutations: mutations that make the sequence less similar to consensus sequence : decreased rate of transcription
When does RNA transcription begin?
When the core RNA polymerase binds to the promoter region with the help of sigma factor (holoenzyme)
Describe termination in prokaryotes
- termination occurs AFTER a termination sequence is transcribed
- two major types of terminators in bacteria: Rho dependent (requires rho protein) and rho independent (intrinsic terminator)
Describe Rho dependant termination?
- in bacteria
- rho binds to RNA upstream of terminator
- RNA polymerase reaches termination sequence and pauses
- Rho catches up and unwinds DNA RNA hybrid using helicase activity
Describe Rho Indépendant termination
- intrinsic terminator
- RNA polymerase países at Us
- hairpin formation destabilizes RNA DNA hybrid (AU base pairs are relatively weak)
- transcription terminates when inverted repeats forms a hairpin followed by a string of uracils : RNA transcript dissociates from polymerase and DNA reanneals
What makes up the promoter of eukaryotes?
- core promoter and regulatory promoter form RNA polymerase II promoter
Describe the parts of the eukaryotic transcription apparatus
- enhancer sequence : transcriptional activator protein
- regulatory promoter: transcriptional activator protein
-core promoter : basal transcriptional apparatus
What is the core promoter?
- In eukaryotes
- extends upstream and downstream of start site
- has a number of consensus sequences: TATA, for transcription factor binding
what is the regulatory promoter?
- upstream of the core promoter, exact location can vary
- transcriptional activator proteins bind to consensus sequence and affect the rate of transcription
- can enhance or decrease transcription, varies a lot!
How does the basal transcription apparatus form?
- forms the core promoter of RNA polymerase II
- TATA Box + tata binding protein : combine with transcription factors, RNA polymerase, and a mediator to form the basal transcription apparatus
- basal transcription apparatus forms around the promoter
Describe the termination of transcription in eukaryotes
- does not end at a specific sequence
- requires cleavage of mRNA at a specific site
- at 5’ to 3’ exonuclease degrades reminding mRNA terminating transcription
