Transcription

Question 1

Coding strand

Answer 1

Strand of DNA that RNA strand ends up essentially being the same as
Also called sense strand

Question 2

Template

Answer 2

Strand of DNA that RNA is synthesized off of

Also called antisense strand

Question 3

Time and place of transcription and translation in prokaryotes versus eukaryotes

Answer 3

Proks: coupled transcription and translation
Euks: transcription and translation separated spatially and temporally

Question 4

Splicing in prokaryotes vs. eukaryotes

Answer 4

Proks: no splicing
Euks: splicing

Question 5

of RNA polymerases in eukaryotes versus prokaryotes

Answer 5

Proks: 1 polymerase
Euks: 3 polymerases

Question 6

of genes per RNA in prokaryotes versus eukaryotes

Answer 6

Proks: polycistronic RNA (many genes per RNA)
Euks: monocistronic RNA (1 gene per mRNA)

Question 7

What directs polymerase to the correct spot in prokaryotes versus eukaryotes?

Answer 7

Proks: sigma
Euks: general transcription factors

Question 8

Polymerase I

Answer 8

Class of promoter: I
Transcripts: rRNA precursor
Location: nucleolus

Question 9

Polymerase II

Answer 9

Class of promoter: II
Transcripts: mRNA, miRNA, snRNA
Location: nucleoplasm

Question 10

Polymerase III

Answer 10

Class of promoter: III
Transcripts: tRNA and specialized rRNA
Location: nucleoplasm

Question 11

DNA footprinting

Answer 11

Can be used to determine where on DNA polymerase binds
2 samples of DNA: treat 1 sample of DNA with protein of interest
Nuclease chops up DNA samples
Gel electrophoresis: no fragments shown where protein was bound to DNA sample (protein protects DNA from nuclease)

Question 12

Filter binding assay

Answer 12

Way to measure polymerase affinity for DNA
1. Label promoter region
2. Incubate DNA with polymerase
3. Apply DNA/polymerase sample to filter that binds proteins tightly
If polymerase is bound to DNA, radioactivity is seen on filter (free DNA flows through)
Increased radioactivity=increased binding

Question 13

Class I promoter

Answer 13

Made up of core promoter and UCE (upstream control element)

Question 14

Transcript of pol I to rRNA

Answer 14

Transcript contains rRNA linked by external and internal transcript spacers
Cleaved to make rRNA
Processing is done to maintain stoichiometry of transcripts (all under the same promoter)

Question 15

Class III promoter

Answer 15

Made up of Box A and Box B (separated by a short element)

Located downstream of +1 site

Question 16

Class II core promoter

Answer 16

Contains:
1. TATA box or DCE (downstream core element)
2. BRE (transcription factor II B response element)
3. Inr (initiator; contains +1 site)
Sequences are recognized by proteins that recruit polymerase

Question 17

Reporter assay

Answer 17

Way to determine which sequences in a promoter are necessary for transcription
Make plasmid clone and put promoter of interest upstream of reporter gene (ex- luciferase)
If promoter is being used, reporter gene is also transcribed
Luciferase: measure light production

Question 18

Linker scanning

Answer 18

Changing big chunks of DNA and seeing results of change via qPCR
Done by cutting piece of DNA and ligating in linker (doesn’t contain sequence of interest)
If linker is where key sequence in promoter used to be, then transcription is lessened
Not just removing sequences: also reversing sequences or moving them around

Question 19

TATA box

Answer 19

Contained in promoter regions of specialized genes
Bound by TATA binding protein
Responsible for recruitment of other general transcription factors

Question 20

TATA binding protein binding to TATA box

Answer 20

TBP (saddle-shaped protein) forces open the minor groove
Bent conformation (80 degree angle) is recognized by other transcription factors

Question 21

Transcription factor that can take over the role of TBP in TATA-less genes

Answer 21

TFIID

Question 22

Housekeeping gene

Answer 22

Always on (always transcribed)

Question 23

Conditionally expressed gene

Answer 23

Used (transcribed) only when needed

More likely to have TATA boxes

Question 24

Transcription initiation steps

Answer 24

1. TATA binding protein and transcription factor II D (TFIID) bind to and bend DNA
2. TFIIA and TFIIB recognize bent DNA and recruit RNA pol II and TFIIF
3. TFIIE and TFIIH come in and TFIIH opens promoter
4. TFIIH phosphorylates tail of pol II, causing tail to dissociate from pre-initiation complex (promoter escape)

Question 25

Transcription elongation factors

Answer 25

Add phosphates to RNA pol II when it stalls during transcription initiation

Question 26

Abortive transcripts

Answer 26

Transcripts that are 10 nucleotides long or less and are degraded during promoter escape

Question 27

Mediator

Answer 27

Mediates interaction between the activator and RNA pol II | Multi-subunit complex that is required for transcription

Question 28

Components of preinitiation complex

Answer 28

Activators (bound to enhancer sequences as well as mediator) Mediator complex (bound to some activators and RNA pol II) RNA pol II Chromatin remodelers and HATs (bound to other activators)

Question 29

Cis regulatory elements

Answer 29

Sequences that are on the same strand as the encoded gene of interest

Question 30

Trans acting factors

Answer 30

Come from elsewhere (not near or on the gene) to bind to regulatory elements

Question 31

Places where regulatory promoters can exist relative to the gene of interst

Answer 31

Can be either upstream or downstream of gene of interest

Question 32

2 domains of activators

Answer 32

Activation domain | DNA-binding domain

Question 33

Leucine zipper

Answer 33

Form of DNA binding domain of activators Dimer (like pair of tongs picking up DNA) Stretches of leucines interacting with each other (tong handles; coiled coil domain)

Question 34

Helix-loop-helix

Answer 34

Form of DNA binding domain of activators Dimer Basic amino acids interact with DNA Flexibility of loop allows for tight packing

Question 35

Homeodomain

Answer 35

Form of DNA binding domain of activators 3 alpha helices Amino acids in 3rd helix interact with DNA Helix-turn-helix

Question 36

Zinc finger

Answer 36

Form of DNA binding domain of activators "Finger": alpha helix Zn stabilizes structure Inserts into minor groove

Question 37

Combinatorial control advantages

Answer 37

``` Energy conservation (can activate multiple genes simultaneously with the same transcription factors) Tighter gene control (specificity of response to cell signals) ```

Question 38

2 roles of activators

Answer 38

Bind to pre-initiation complex | Enhance chromatin remodeling

Question 39

Insulators

Answer 39

Big, bulky proteins that can be used to selectively block activators

Question 40

4 ways that repressors decrease gene expression

Answer 40

1. Physically block activators 2. Block activation domain 3. Block RNA pol escape 4. Recruit HDAC

Question 41

Phosphorylation of RNA pol II tail

Answer 41

Recruits RNA processing enzymes Which ones depend on stage of transcription (promoter escape- capping factors recruited; elongation- splicing factors recruited)

Question 42

Type of linkage that m7G cap undergoes

Answer 42

5'-5' linkage (5' end of RNA to 5' end of cap)

Question 43

How m7G cap is added onto RNA

Answer 43

Beta phosphate of transcript (gamma phosphate is cleaved by RNA pol II) attacks alpha phosphate of m7G

Question 44

3 purposes of 5' cap

Answer 44

1. Protect RNA from degradation 2. Ribosome can recognize cap 3. Encourage splicing of 1st intron

Question 45

Transcription termination and polyadenylation

Answer 45

1. Poly A signal sequence in RNA is recognized by splicing factors 2. RNA is cleaved 3. Poly A polymerase adds adenine nucleotides to RNA 4. Poly A binding proteins bind to adenines

Question 46

4 purposes of polyadenylation

Answer 46

1. Protect 3' end of RNA 2. Extend RNA half life 3. Encourage splicing of last intron 4. Enabling translation

Question 47

Torpedo model of transcription termination

Answer 47

Torpedo protein chews up excess transcript | When torpedo catches up to RNA pol (chews up all of excess transcript), RNA pol is released