Exam 2- Transcription Flashcards
What are the different types of rRNA present in prokaryotes and eukaryotes?
Prokaryotes: 28S, 16S, 5S rRNA’s
Eukaryotes: 28S, 18S, 5.8S, 5S
Describe the RNA Polymerase subunits involved in prokaryotic mRNA synthesis
Prokaryotic RNA Polymerase has a core enzyme that is made up of 2alpha, 1beta, and 1beta’ subunits that posses 5’ —> 3’ polymerase activity and 1alpha subunit that functions as an initiator that recognizes the promoter sequence
What are the different RNA polymerase of eukaryotes?
RNA polymerase I- rRNA
RNA Polymerase II- hnRNA (precursor for mRNA)
RNA Polymerase III- tRNA
Promoter elements in eukaryotes
TATA Box
GC Box
CAAT Box
RNA polymerase II starts polymerization once the transcription is initiated by transcription factors that bind to promoter regions of the gene that consists of promoter elements
Describe the transcription factors involved in eukaryotic TATA binding
TFIID: TATA box always recognizes TFIID
- TBP: TATA binding protein
- TAF’s: TATA Box activating factors
- TFIIA’s & TFIIB’s: bind RNA Polymerase II
RNA polymerase II binds to TFIIF which interacts with TFIIE and TFIIH
- TFIIH (helicase) unwinds DNA
- RNA Polymerase II starts transcription
What is the protein factors that binds to GC box?
SP1
What are enhancers?
Enhancers bind to specific transcription factors far (>1000 bp’s) away from the general transcription initiation. During transcription, they bend from a far away distance to interact with the transcriptional machinery
- can be upstream (5’) or downstream (3’)
- can be sense or antisense orientation
Give 2 examples of enhancers
- NFKB1 (Nuclear Factor k B1): binds to enhancer region of immune response genes and regulates the inflammatory response
- PEP Carboxy Kinase (PEPCK): PEPCK contains an enhancer called cAMP response element (CRE) that binds to cAMP response element binding protein (CREBP). It also contains another enhancer called glucocorticoid response element (GRE) that binds to cortisol banding protein
Describe 2 enhancer elements of the PEPCK gene and their respective binding proteins
CRE (cAMP response element) that binds to CREBP (cAMP response element binding protein)
GRE (glucocorticoid response element) that binds to cortisol binding protein
What are euchromatins and heterochromatic should?
Euchromatins- heavily acetylated making it transcriptionally activated part of the chromosome
Heterochromatins- hypermethylated making it transcriptionally inactive part of the chromosome
How does methylation and acetylation affect gene transcription?
Acetylation: transcriptionally active
Methylation: transcriptionally inactive
What is primary (pri-RNA/hnRNA)?
mRNA precursor synthesized by RNA Polymerase II
Describe the 5’ and 3’ modifications that happens to primary RNA/hnRNA?
7meG-cap: 7-methyl guanine cap containing nucleotide is added to hnRNA at the 5’ end
- the cap itself would contain a 5’-5’ linkage (Me-G-P-P-P-5’-N) that protects the newly synthesized RNA from degradation
- In addition, the 7-meG-cap helps recognize the 5’ end of the processed mRNA by the ribosome complex during protein synthesis (ie translation initiation)
Poly-A tail: 3’ modification immediately after Tx of the complete hnRNA in the nucleus, a protein complex along with polyA polymerase (PAP) recognize the motif AAUAAA present upstream to the 3’ end of the hnRNA. PAP complex removes the hnRNA from RNA Polymerase II, before adding around 200 adenylyl residues at the 3’ end. The 3’ end with a bunch of A residues is called the polyA tail
What is the recognition motif needed in the hnRNA for it to be poly adenylated.
AAUAAA
What are some purported functions for the poly A tail
Facilitation of hnRNA transport from nucleus to cytosol
Stability of mRNA
Describe the process of splicing.
Exon1-Intron1 junction has the motif GpGU
In the middle of the intron, the prominent catalytic A called branch site A
At the intron1-exon2 junction there is a consensus motif AGp
First 2’-OH of the branch site A attacks the phosphodiester of G on the GpGU site
Now the cleaver exon 1 attaches the 5’ end of the exon 2 and form spliced Exon1-Exon2
Splicing requires small nuclear ribonucleoprotein particles (SnRNP’s)
What is mRNA copy number?
Transcripts per cell
Describe the processing of tRNA.
For tRNA, 5’ leader sequences removed first
Introns are removed and some bases are modified
At the 3’ end, UU are removed and the 3 bases CCA are added
The CCA at the 3’ end of the tRNA is crucial for amino acid addition (ie charging) during translation
Describe the processing of hnRNA.
Splicing - snRNA in the splicing of exons and introns on hnRNA to obtain mature mRNA
5’cap
3’PolyA tail
Describe the mode of actions of rifampicin, actinomycin D,k and alpha-amanitin
Rifampicin: specific to Beta-subunit of prokaryotic RNA polymerase
- Abx
Actinomycin D: binds to DNA template and interferes with the movement of RNA polymerase along the DNA
- Chemotheurapeutic
Alpha-amanitin: it is a peptide produced by the mushroom Amanita phalloides. It inhibits eukaryotic RNA polymerase II. Liver toxicity is the most immediate effect
- toxin
Describe the transcriptional regulation of p21.
p21: cyclin dependent kinase inhibitor
P53 (guardian of the genome) is an anti-oncogene protein product that binds to the promoter of the p21 gene and causes upregulation of p21 gene product which in turn stops cell cycle progression by inhibiting cell cycle kinases
Describe the transcriptional regulation of gamma-globulin
GATA-1 is a suppressor of gamma globulin gene. GATA-1 keeps the mRNA expression at an appropriate amount so that the proper stoichiometry of alpha and beta Hb chains can be maintained.
Upregulation of gamma-globulin leads to down-regulation of beta-globulin
Thalassemia
Autosomal disorder
G to A mutation of upstream element of gamma Hb gene results in loss of binding by transcription factor, GATA-1
Upregulation of gamma-globin, down regulation of beta-globin
