Transcription Flashcards
What is a gene?
basic unit of hereditary and a sequence of nucleotides in DNA that encodes for the synthesis of a gene -> protein product.
Process and role of transcription
role of transcription is to convert DNA to RNA to make a protein.
Initiation Transcription
Promotor region: TATA, CAAT, GC
RNA polymerase 2 and GTFs will bind at the promotor site to start transcription
RNA polymerase requires several general TFs to start transcription:
- TFIID binds the (promotor site) TATA box, through the TBP subunit
-> this enables binding of TFIIB - The rest of general TFs (TFIIE, TFIIH) and RNA polymerase II assemble at the promoter
- TFIIH contains a DNA helicase as one of its subunits and can use the energy from ATP hydrolysis to separate DNA double helix at the transcription start point, exposing the template strand.
- TFIIH also phosphorylates RNA pol II, changing its shape
-> in RNA pol II being released from the TFs and starting the elongation
In addition to the promotor needing the 5TFs and RNAP 2, transcription needs regulatory proteins:
- Mediator proteins: large protein complex
- Transcription activator proteins: bind to enhancers to attract RNA2, TFs, promotor, and mediator. -> they also attract ATP dependant chromatin modelling complexes and histone modifying enzymes.
Elongation
RNA Polymerase reads the template:
- Reads the DNA in the 3’ to 5’ end. (Template strand)
- Synthesizes RNA in the 5’ to 3’ end.
The RNA transcript is nearly identical to the non-template, or coding, strand of DNA. However, RNA strands have the base uracil (U) in place of thymine (T), as well as a slightly different sugar in the nucleotide.
Amastin
toxin component of mushrooms which can inhibit RNA polymerases.
Rifampin
antibiotic that can inhibit the RNA in prokaryotic cells
Termination:
Sequence dependant termination of the RNA growth chain
hits a terminator sequence AA U AAA
a polyadenylation signal
activates enzymes cleaving RNA
RNA polymerase will keep transcribing until it gets signals to stop. The process of ending transcription is called termination, and it happens once the polymerase transcribes a sequence of DNA known as a terminator.
How does RNA polymerase differ to DNA polymerase?
RNA polymerases catalyse the linkage of ribonucleotides
RNA polymerases can start an RNA chain without a primer
RNA polymerases are absolutely processive – the same RNA polymerase starts and finishes an RNA mole
Post Transcriptional Processes that transform pre-RNA to Mature mRNA
Capping
Tailing
Splicing
The processing of mRNA involves modification of the 5’ terminus by capping with 7-methlyguanosine (m7G), modification of the 3’ terminus by polyadenylation, and removal of introns by splicing.
CAPPING
When RNA polymerase 2 has made a nucleotide sequence of RNA, the 5’ end is modified by the addition of a cap consisting of m7G
Capping is initiated by the addition of guanosine triphate (GTP) in reverse linkage of the terminal nucleotide of the RNA
Then methyl groups are added to the G residue and the ribose moieties of one or two 5’nucelosides of the RNA chain.
The 5’ cap protects the new mRNA from degrading and assists ribosome binding during translation
POLYDEYLATION
The 3’ end is not defined by termination of transcription but by the cleavage of the primary transcript and addition of a POLY-A tail -> polyadenylation
The POLYA tail:
protects from degrading
aids export o mature mRNA to cytoplasm
involved in binding proteins to initiate translation
SPLICING
Introns removed and exons are spliced together -> this takes place at complexes called spliceosomes.
Cleavage at the 5’ splice site: formation of a lariat like intermediate
Cleavage at the 3’ splice site: ligation of exons
Why splice?
New transcripts are spliced in more than 1 way (alternative splicing)
Increases diversity of proteins that can be encoded by the 20,000 genes.
RNA splicing enables eukaryotes to increase the coding potential of their genomes
mRNA Transport:
mRNA is synthesised in the nucleus and needs to go to the cytosol for translation.
Only can be transported once the proteins present on the mRNA molecule collectively signify that processing was successfully completed.
The mRNA is then guided via nuclear pore complexes in the nuclear membrane which connects the nucleoplasm and cytosol
β thalassemia and transcription
Abnormal processing of the β-globin
primary RNA transcript in humans with the β thalassemia
The disease (severe anaemia due to aberrant haemoglobin synthesis) is caused by splice-site mutations found in the genomes of affected patients
Phenylketonuria (PKU)
A single base mutation
156bp deletion in the mRNA, corresponding precisely to the length of Exon 12 that leads to a truncated PAH (phenylalanine hydroxylase) protein that is unstable with almost zero PAH activity
Tamoxifen
selective oestrogen receptor modulator (SERM) medication used to treat breast cancer in men and women and as a prophylactic agent against breast cancer in women.
Reduces the risk of breast cancer coming back by 40% to 50% in postmenopausal women and by 30% to 50% in premenopausal women
