Ch. 23: RNA Synthesis-DNA Transcription

Question 1

What 3 identical but functionally different RNA molecules are transcribed from anti-sense (-) DNA (template strand)?

Which is the most abundant in the cell?

Answer 1

messenger RNA (mRNA)
transfer RNA (tRNA)
ribosomal RNA (rRNA) ➡️ most abundant

Question 2

Describe the sequence of rRNA compare to that of mRNA.

What are the subunits & nucleotides found in prokaryote ribosomes? In eukaryote ribosomes?

Answer 2

Has sequence complementarity to regions of mRNA

Prokaryote: 5S (120), 16S (1541), 23S (2904)
Eukaryote: 5S (120), 5.8 (150), 18S (2100), 28S (5050)

Question 3

What is the function of tRNA?

Answer 3

Binds an a.a. at one end, mRNA at other end

Acts as adaptor to carry a.a. of a protein to correct place as coded for by mRNA

Question 4

Describe the 5’ terminus and 3’ terminus of tRNA.

What part of tRNA interacts with mRNA codon?

Answer 4

5’ terminus: base-paired

3’ terminus: 4 base single-stranded region, XCCA-3’-OH
(CCA = acceptor stem ➡️ adenine is a.a. attachment site)

Anticodon loop

Question 5

What are the modified bases for tRNA?

Answer 5

dihydrouridine (DHU)
ribosylthymine (rT)
pseudouridine
inosine (I)
wyosine (Y)

Question 6

What is the function of mRNA in DNA transcription?

Answer 6

photocopy of a gene (has sequence that is complimentary to 1 DNA strand and identical to other strand) ➡️ carries the info stored in DNA in nucleus to cytoplasm (for translation)

Question 7

What is the start codon in mRNA to initiate translation?

What are the stop codons in mRNA to terminate translation?

Answer 7

Start codon: AUG

Stop codons: UAG, UGA, UAA

Question 8

What makes prokaryotic mRNA different from eukaryote mRNA?

Answer 8

Polycistronic (polygenic): carry info for synthesis of several polypeptides from 1 mRNA

Question 9

How is the timing of transcription and translation different in prokaryotes compared to eukaryotes?

Answer 9

Prokaryotes: transcription and translation are coupled (happens at the same time, since mRNA does not have to be stable)

Eukaryotes: transcription and translation happen separately

Question 10

What makes mRNA in eukaryotes different from mRNA in prokaryotes?

Answer 10

1) Has info for only 1 polypeptide
2) it is heterogeneous nuclear RNA (hnRNA) = has introns & exons
3) goes through post-transcriptional modifications
4) transported to cytoplasm for translation

Question 11

What are the post-transcriptional modifications eukaryote mRNA goes through?

What is the main purpose of these modifications?

Answer 11

5’ Capping
Poly A tailing
Splicing (alternative splicing, RNA editing)

Give mRNA stability, protection, ⬆️ lifespan

Question 12

What is the function of small nuclear RNA (snRNA)?

Answer 12

Mediate the processing of primary transcripts (large precursor molecules formed from DNA transcription of genes for mRNA, tRNA, and rRNA) in nucleus ➡️ produces functional molecules for export to cytosol

Question 13

In what direction does an RNA chain grow?

What enzyme initiates RNA synthesis? Is a primer needed?

Answer 13

‘5 to 3’ direction

RNA polymerase
No primer needed

Question 14

What are 4 features specific to transcription (differ from DNA replication)?

Answer 14

1) Transcription has more initiation sites
2) More RNA polymerase molecules per cell than DNA polymerase
3) Rate of RNA polymerase (50-100 bases/s) slower than DNA polymerase (1000 bases/s)
4) RNA polymerization has ⬇️ fidelity than DNA

Question 15

Prokaryote transcription:

Describe the structure of E. coli RNA polymerase and the function of its subunits.

Answer 15

Has 5 subunits ➡️ alpha2 beta beta’ sigma = holoenzyme

Beta’: binds to DNA
Beta: binds to rNTPs, interacts with sigma
Sigma: recognizes promoter sequences on DNA

Question 16

What are the 3 promoter regions in prokaryotic transcription?

Answer 16

Initiation site (+1)
Pribnow box (-10)
-35 region

Question 17

What are the 3 main steps of transcription & translation?

Answer 17

1) Initiation
2) Elongation
3) Termination

Question 18

What occurs to transition from initiation ➡️ elongation in prokaryote transcription?

Answer 18

After initiation: RNA polymerase holoenzyme loses sigma subunit ➡️ becomes core enzyme ➡️ conducts elongation

Question 19

Inhibitors of prokaryotic transcription initiation:

Rifamycin B: how does it ➖ transcription initiation?

Answer 19

Binds to beta subunit of RNA polymerase ➡️ blocks binding of incoming rNTPs at initiation site

Question 20

Inhibitors of prokaryotic transcription initiation:

Rifampin (rifampicin): how does it ➖ transcription initiation?

Answer 20

Prevents translocation of RNA polymerase along DNA template

Question 21

Inhibitors of prokaryotic & eukaryotic transcription initiation:

Actinomycin D: how does it ➖ transcription initiation?
What diseases can it be used to treat?

Answer 21

Binds to DNA

Useful to treat childhood neoplasms (Wilms’ tumor) & choriocarcinoma

Question 22

What 3 characteristics are required for termination of transcription to occur at a specific base sequence?

Answer 22

Factor independent transcription termination:

1) inverted repeat base sequence with a central non-repeating segment
2) ⬆️ G + C content
3) ⬆️ A + T content resulting in 6-8 U’s in RNA

Question 23

What factor is required in factor dependent transcription termination?

Answer 23

rho factor

Question 24

What are the specific features of transcription in eukaryotes?

Answer 24

1) transcription & translation separated in space & time ➡️ better regulation of gene expression ➡️ variety of eukaryotic forms & functions
2) RNA stabilized via post-transcriptional modification
3) 3 types of RNA polymerases (I, II, III)
4) Promoters have a TATA box near start site

Question 25

What types of RNA are each of the 3 classes of eukaryotic RNA polymerases used for?

Answer 25

I: rRNA (nucleolus)
II: mRNA, snRNA
III: tRNA, 5S rRNA

Question 26

What are the sensitivities to amanitin (of specific 🍄) of each of the 3 classes of RNA polymerases?

Answer 26

I: insensitive to amanitin
II: strongly ➖ by amanitin
III: ➖ by ⬆️ [amanitin]

Question 27

What makes promoters of eukaryotic transcription more complex than that of prokaryotes?

Answer 27

TATA box & CAAT box

Many non-polymerase factors needed for RNA polymerase binding to DNA

Question 28

What are the structural features eukaryotic RNA polymerases I, II, and III have in common?

Answer 28

1) Big, multimeric proteijs (500-700 kD)
2) Have 2 large subunits (sequences similar to beta and beta’ in E. coli RNA polymerase)
3) have subunit homologs of alpha and omega in E. coli polymerase

Question 29

What makes up the RNA polymerase II transcription initiation complex? What does each part do?

Answer 29

1) Activators: binds to genes at enhancer sites ➡️ help to decide which genres to switch on, ⬆️ transcription rate
2) Coactivators: “adapter” molecule integrate & relay signals from Activators ➡️ basal transcription factors
3) Basal transcription factors: position RNA polymerase at start of protein coding region

Also, repressors: binds to genes at silencer sites ➡️ interfere with activator function ➡️ slows transcription

Question 30

Transcription factors for RNA polymerase II:

Step 1: Formation of DAB complex ➡️ what is the function of each TF?

Answer 30

TFIID: Binds to TATA box (TBP), bends DNA

TFIIA: stabilizes promoter binding by TBP

TFIIB: stabilizes TBP-TATA complex, recruits RNA polymerase II (essential)

Question 31

Transcription factors for RNA polymerase II:

Step 2: Binding of RNA polymerase II to DNA ➡️ which TF is the polymerase complexed with?

Answer 31

TFIIF: Binds RNA polymerase II, positions template during initiation

Question 32

Transcription factors for RNA polymerase II:

Step 3: Which TFs bind after RNA polymerase II binds to DNA? Functions of each TF?

Answer 32

TFIIE: Assists recruitment & activity of TFIIH, promoter melting (denaturation or promoter region)

TFIIH: ATPase, CTD kinase, helicase, promoter melting & escape

Question 33

Transcription factors for RNA polymerase II:

Step 4: Elongation ➡️ what TF stimulates elongation for RNA polymerase II?

Answer 33

TFIIS (TFIIF may also help)