Transcription/RNA Processing/Gene Regulation Flashcards
In what polarity does DNA transcription occur?
5’->3’
What strand of DNA does the RNA product come from?
Template strand.
Don’t get confused with coding strand. Call it the non-template strand when you see it.
When the sigma unit of prokaryotic RNA polymerase binds with the core enzyme, what does that generate?
Holoenzyme
In what direction does RNA polymerase read DNA?
3’->5’
Generates 5’ -> 3’
What is prokaryotic RNA pol composed of?
sigma unit
2 alpha units
2 beta units
1 omega unit.
What are the three stages of transcription?
- Initiation
A. Holo-RNAP binds to promoter
B. Sigma-dependent, promoter-dependent
C. Principal site for regulation!!! - Elongation
A. RNAP leaves and starts reading 3’-5’
B. Sigma dissociated from RNAP and promoter - Termination
A. DNA-RNA-RNAP complex disassembles depending on signals (2 terminators in bacterial transcription)
What are the structures of bacterial promoters?
Made of of consensus seq. 2 sigmas can binds
- -35 sequence [TTGACA]
(19 BP later) - Pribnow box [TATAAT]
(7 BP)
Transcription starts
Promoter is always DS-DNA
What is the Rho-dependent termination sequence?
Hexamer protein. ATPase!!!
Binds with RNA, not DNA.
Will influence the dissociation of the complex.
What is the Rho-independent termination seq?
Stem-loop terminator.
Structure starts BP-ing with self and run of Us at 3’ end.
Clinical Correlation:
Where does Rifampin bind?
Bacterial RNA polymerase.
What is each of the eukaryotic RNA polymerases responsible for in Eukaryotic cells?
RNA Pol I -> rRNA
RNA Pol II -> mRNA
RNA Pol III -> tRNA
What kind of regulatory elements are present in Eukaryotic transcription? (Pol II)
1. Core promoter: A. Transcription start site (INR) +1 B. TATA box (-25) C. DPE (downstream promoter element) +30 D. General: binds site for Pol II and GTFs
- Proximal promoter
A. 250 BP from start
B. Can be b4,during, or downstream of gene
C. Also binds activators.
What is TFIID?
Nucleates assembly of transcription preinitiation complex.
Composed of TBP and 14-tbp associated factors (TAFs)
What is composition of RNA Pol II? How is it different/similiar to prokaryotic RNA pol?
Eukaroytic = 12 subunits Rbp1-12 (6 in prokaryotic)
Rpb1 - B’
Rpb2 - B
Rpb3/11 - a2
Rpb6 - omega
There is no sigma counter part in RNAP II
If there was a counterpart, it would just be all these general transcription factors.
What is the 7-methylguanosine “cap”? Where is it located?
5’ end of mRNAs.
Prevents degradation
Need to prevent breakdown of mRNA being produced b/c it isnt being translated right away.
Why is the polyadenylation og primary transcript important? How is it added?
MRNA is cleaved after a cleavage signal
Poly-A tail added by poly(A) polymerase
ATP dependent!!!
Why is TFIIH importaqnt in DNA repair?
Couples transcription with DNA repair.
What is in poisonous mushrooms that makes them?
Alpha-amatin. Binds with RNA pol II and inhibits elongation.
What are introns?
Intervening sequences
What are the consensus seq for splicing mRNA precursors?
5’ (end of the upstream exon) -> “GU”
3’ (start of downstream exon) -> “AG”
What kind of chemical reactions are involved in the mRNA splicing mechansim?
2 Transesterifications
What is a snRNP? When do we see them appear in the spliceosome?
Small nuclear ribonucleo proteins
U1 - binds with 5’ GU seq
U2 - binds with 3’ A / AG seq
U4/5/6 complex comes in and starts removal of intron
CLINICAL Correlation:
What happens in certains types of B-thalessemias?
SNP (single base mismatch) that makes the splice site change.
In B-thalessemia, this will make the exon site longer (b/c thinks it should be splicing sooner). Yields 18BP (6 codons extra) in functional exon area
What is alternative splicing?
Production of miltiple, related proteins form single gene.
Happens when exons are skipped over during splicing.
Genes producing different mRNAs
When are transcripted polycistronic? Monocistronic?
Poly - Prokaryotes
Mono - Eukaryotes
What binds to the promoter in eukaryotic cells? Prokaryotics?
TFIID(TBP) - Eukaryotic promoter binding
(TBP, TFIIB, TFIIF- bind RNA Pol II)
Sigma factor (RNA pol holozyme subunit) binds to promoter.
What are all steroid hormones derived from?
Cholesterol (c27)
Eventually get to :
Glucocorticoids (C21) - metabolism
Mineralocorticoids (C21) - Blood pressure
Androgens (C19 -male sex)
Estrogens (c18- female sex)
What is an example of a hormone antagonist?
Tamoxifen - competitive inhibitor.
What 4 domains are present on a steroid receptor?
- Activation Domain
- Dimerization domain
- DNA binding (contains Zn finger)
- Ligand binding
What steroid receptor domain contains a zinc finger motif?
DNA-binding domain
What is the difference between glucocorticoid response elements and estrogen response elements?
Both are 17 bp in length. Both are pallindromic. Both occur on dsDNA
Difference is 2 sets of BP in sequence.
Gluco: starts w/ AA(5’), ends w/ TT (3’)
Estrogen: startw/ GT (5’), ends w/ AC(3’)
Binding of a ligand to a SRE recruits what?
Co-activators
How do steroid hormones regulate intracellular transcription?
- Cortisol binds with receptor activating Zn finger DNA binding
- Steroid-receptor complex binds with specific regulatory seq.
- Hormone receptor complex with coactivators cause transcription modification
What is actinomycin used for, how does it work?
Binds DNA, inhibiting transcription elongation in prokaryotes and eukaryotes
Effective cancer therapeutic
What is the TATA binding protein?
TBP
Part of the TFIID complex.
What is the RNA Pol II preinitiation complex compased of?
TFIIA/B/D (TBP)/F/
E/H (Downstream)
