Transcription, RNA Processing, and Regulation of Gene Expression

Question 1

DNA Topoisomerases

Answer 1

Essential for replication; not necessarily essential for transcription

Question 2

DNA Helicases

Answer 2

the “zippers” that separate the two strands of DNA; ESSENTIAL for transcription
- RNAP; TFIIH

Question 3

RNA Polymerase (prokaryotic)

Answer 3

Transcribes all RNA

Question 4

RNA Pol I

Answer 4

• transcribes: rRNA
• location: nucleolus
• function: ribosome assembly

Question 5

RNA Pol II

Answer 5

• transcribes: hnRNA/mRNA
• location: nucleus
• function: protein synthesis

Question 6

RNA Pol III

Answer 6

• transcribes: tRNA, miRNA, siRNA, snRNA, snoRNA
• location: nucleus
• function: protein synthesis (cytoplasm); diverse nuclear functions

Question 7

RNA polymerase (mitochondrial)

Answer 7

• transcribes: all mitochondrial RNAs
• location: mitochondria
• function: transcription of mitochondrial genes

Question 8

Prokaryotic (bacterial) RNA Polymerase

Answer 8

Core: alpha x2, beta x2, omega
Holo: includes sigma
- sigma confers promoter recognition to RNA pol
- sigma is not required for catalytic activity of RNA pol

Question 9

3 Stages of Transcription

Answer 9

• Initiation: RNAP/Pol II binds promoter (sigma recognizes promoter in prok); no primer required; principal site for REGULATION of transcription
• Elongation: lengthens transcript; Holo-RNAP dissociates from sigma subunit (prok); RNAP II dissociates from GTF (euk)
• Termination: RNA synthesis stops; depends on DNA encoded RNA signals (DNA says when, RNA does the stopping)

Question 10

Genetic Elements of Bacterial Operon

• structural genes
• operon
• promoter
• activator binding site
• operator
• concept

Answer 10

• structural genes: encode protein; down stream of all
• operon: mRNA encoded info; transcribed as single mRNA chain
• promoter: binding site for RNA polymerase
• activator binding site: DNA sequence that binds a protein that enhances transcription
• operator: DNA sequence that binds proteins that inhibits transcription; blocks holoenzyme
• concept: compact arrangement of DNA sequence elements: often less than 150bp of DNA from start site of transcription to activator binding site

Question 11

Structure of Bacterial Promoter

Answer 11

-35 sequence — 19 base pairs — -10 Pribnow Box (TATAAT) — ~ 2 bp — Start of transcription

Question 12

Rho-dependent Transcription Termination

Answer 12

PROKARYOTES ONLY
- termination signal is encoded in the gene
- signal is manifest as RNA
- results in dissociation of the DNA-RNA-protein-complex
Rho = torpedo
RNA pol = ship
Termination = BOOM

Question 13

Rho-independent Transcription Termination

Answer 13

PROKARYOTES ONLY
Stem-Loop (S-L)
- RNA S-L structure is encoded in DNA and functional as RNA
- structural features: S-L and a run of UUUUUU immediately following
- terminates transcription within the catalytic core of RNAP (in CONTRAST with RHO-dep)

Question 14

Rifampicin Antibiotic

Answer 14

binds RNA polymerase and inhibits initiation of transcription

Question 15

RNA polymerase II

Answer 15

EUKARYOTES

• 12 subunits; large complex with more subunits than prok; some subunits look and act similar to prok
• NO SIGMA COUNTERPART IN RNAP II

Question 16

Eukaryotic Transcription (Pol II) Core Promoter

Answer 16

REGULATORY dsDNA sequence

• transcription start site (INR): +1
• TATA box: -25
• DPE (downstream promoter element): +30
• promoter function: binding site for RNA Pol II and “GTF”s

Question 17

Eukaryotic Transcription (Pol II) Proximal Promoter

Answer 17

REGULATORY ds DNA sequence; NOT A BINDING SITE FOR RNA POL II

• <120 bp from start
• binding site for activator proteins

Question 18

Eukaryotic Transcription (Pol II) Enhancer

Answer 18

REGULATORY dsDNA sequence; binds regulatory proteins

• > 250 bp from start (far from genes controlling)
• function in either orientation
• can act anywhere
• binding site for transcription activators

Question 19

General Transcription Factors

Answer 19

multi sub unit GTF that has responsibility to recognize promoter DNA sequences and tell RNA poly where to associate with DNA
*** TFIID ALONE NOT sufficient to create RNA pol binding site

Question 20

TFIID

Answer 20

Equivalent of sigma factor
* recognizes promoter*
ALONE NOT SUFFICIENT FOR BINDING

Question 21

GTF assembly

Answer 21

assemble on promoter in a stepwise manner

Question 22

Pol II Binding orientation

Answer 22

nothin on Pol II recognizes the DNA; doesnt have to because Pol II DOESNT touch DNA, instead touches TF

Question 23

TFIIH

Answer 23

• involved in transcription coupled repair (NER)

- required to melt promoter DNA

Question 24

5’ Cap

Answer 24

• not encoded in DNA
• protects 5’ end from exonucleases
• important in translation initiation: bound by eIF4E

Question 25

eIF4E

Answer 25

binds 5’ cap and initiates protein synthesis in cytoplasm

Question 26

PolyA tail

Answer 26

• encoded in DNA
• recognized and function in RNA
• endonuclease recognizes and cuts at the end; polyA tail added

Question 27

Transcription-coupled DNA repair

Answer 27

-NER pathway
- fast, preferential repair of the transcribed strand of an active gene
TCR factors include subunits of TFIIH, including two helicases, XPB and XPD, proteins whose genes products are known to be involved in the disease Xeroderma pigmenosum (XP), Tricothiodystropy (TTD) and Cockayne Syndrome (CS)

Question 28

Transcription-coupled repair

Answer 28

sub-pathway of NER that repairs the TRANSCRIBED DNA strand faster than the rest of the genome
***TCR requires TFIIH subunits XPB and XPD)

Question 29

Cockayne Syndrome

Answer 29

premature aging syndrome with developmental and neurological defects
a-amanitin?

Question 30

Alpha-amanitin

Answer 30

binds to RNAP II and inhibits the elongation stage of transcription (starts but can’t be completed)

Question 31

Define:
5’ splice site
3’ splice site

Answer 31

5’ splice site: splice donor site

3’ splice site: splice acceptor site

Question 32

Intron starts/ends

Answer 32

starts 5’ GU ends 3’ AG

INTRON TELLS WHERE TO SPLICE

Question 33

RNA Chain Phosphodiester Bonds

Answer 33

3’ (base n) to 5’ (base n+1)

ALL nucleotides in RNA contain a 2’-OH group

Question 34

SNURPs

Answer 34

small, nuclear, uracil rich, RNA-protein complexes

Question 35

Spliceosome

Answer 35

large complex that removes introns from pre-RNA

Question 36

U1

Answer 36

SNURP

guide sequence; recognizes invariant sequence at 5’ splice site

Question 37

U2

Answer 37

SNURP

recognizes branch site (special A; use it’s OH)

Question 38

U1&2

Answer 38

recognize each other and conformationally change to promote A-OH linkage and splicing

• conformational change requires ATP
• splicing does not require ATP

Question 39

B-thalassemia

Answer 39

results from intronic mutation

• mutation in DNA but effect seen on RNA splicing level
• splice sites changed by point mutations which can incorrectly start/stop intron splicing at incorrect locations

Question 40

Alternative Splicing

Answer 40

a gene produces multiple proteins from the same DNA sequence

Question 41

UTR and Introns

Answer 41

Introns can be found in UTR- REGULATION
Upstream- “leader region”; regulatory effects; change in protein expression
Downstream- change in stability; regulatory effects

Question 42

Transcriptional Activators

Answer 42

hormones BOUND to receptor proteins interacting with enhancer sequences

Question 43

Agonist

Answer 43

substance which initiates a physiological response when combined with a receptor
- bind hormone receptors
- stimulate receptor activity; stimulate gene expression
I.e. anabolic steroids; fentanyl

Question 44

Antagonist

Answer 44

substance that interferes with or inhibits the physicological action of another

• bind hormone receptors
• block receptor acrtivity; repress gene expression
  i. e. tamoxifen; naloxone

Question 45

How to answer test Qs on introns/exons

Answer 45

exon 1 | intron 1 | exon 2

• where’s mutation
• draw out new mRNA and then answer question