07a: RNA Processing

Question 1

Actinomycin D acts as a (X) in (prok/euk).

Answer 1

X = inhibitor of transcription initiation and elongation

Both prok/euk

Question 2

What’s the mechanism of Actinimycin D?

Answer 2

Slips into major groove (intercalates) and prevents unwinding of DNA template

Question 3

In bacteria, mRNA would have which structure at 5’ end? This is (similar/different) in euk.

Answer 3

5’-ppp

Different in euk due to mRNA processing

Question 4

(Prok/euk) active mRNA requires post-(X) modification of precursor, called (Y).

Answer 4

Euk

X = transcriptional

Y = hnRNA (heterogenous nuclear RNA)

Question 5

What are the post-transcriptional modifications?

Answer 5

1. Add 5’ cap
2. 3’ polyadenylation
3. Splicing (remove introns)

Question 6

In which compartment do post-transcriptional modifications take place?

Answer 6

Nucleus

Question 7

Through which pores does hnRNA leave the nucleus after transcription?

Answer 7

Doesn’t! Must be post-transcriptionally modified into active mRNA before it can be transported to cytoplasm

Question 8

What molecule is the mRNA cap?

Answer 8

GTP (added backwards)

Question 9

What type of linkage is found in the mRNA cap?

Answer 9

5’-5’ triphosphate linkage

Question 10

The enzyme (X) is responsible for which events, leading to addition of the mRNA cap?

Answer 10

X = guanylyl transferase

1. Cleaves gamma phosphate off 5’ end of mRNA
2. Cleaves beta and gamma phosphates off GTP
3. Adds GTP backwards to 5’ end of mRNA

Question 11

What’s the function of the mRNA cap?

Answer 11

1. Protect from nuclear degradation
2. Form scaffold for protein binding
3. Needed for efficient translation

Question 12

For polyadenylation, a(n) (X) signal is needed. What’s the sequence?

Answer 12

X = cleavage

AAUAAA

Question 13

What’s the function of the poly-A tail?

Answer 13

1. Protects 3’ end

2. Stabilizes mRNA

Question 14

Which enzymes participate in polyadenylation?

Answer 14

1. Endonucleases (cleave 11-30 NT downstream of cleavage signal)
2. Poly(A) polymerase (adds tail)

Question 15

Do any steps in cap/tail addition require ATP?

Answer 15

Yes - polyadenylation by poly(A) polymerase requires ATP

Question 16

T/F: there are proteins that directly associate with the polyA tail.

Answer 16

True. PolyA binding proteins

Question 17

What are snRNPs composed of?

Answer 17

One small snRNA and several proteins

Question 18

What bond is found at the branch point after slicing?

Answer 18

2’-5’ phosphodiester bond

Question 19

The primary RNA transcript is about (X)% introns and (Y)% exons.

Answer 19

X = 90

Y = 10

Question 20

All (exons/introns) begin with GU and end with:

Answer 20

Introns;

AG

Question 21

Where is the branch point in (intron/exon)? What surrounds it?

Answer 21

Intron; located 50 bases from 3’ end of intron, in a pyrimidine-rich sequence

Question 22

Role of U1

Answer 22

Binds 5’ splice site

Question 23

Role of U2

Answer 23

Binds branch site (and part of catalytic center)

Question 24

Role of U5

Answer 24

Binds 3’ splice site and loops over 5’ site

Question 25

Role of U4

Answer 25

Masks U6 catalytic activity

Question 26

Role of U6

Answer 26

Catalyzes splicing (ribozyme)

Question 27

How does U(X) interact with the 5’ splice site?

Answer 27

X = 1

Anti-parallel base pairing

Question 28

Which two snRNPs have complementary sequences?

Answer 28

U2 and U6 have complementary RNA sequences (form catalytic center)

Question 29

Spliceosome catalyzes two (X):

Answer 29

X = transesterifications

1. 2’ OH of A attacks 5’ splice site
2. 3’ OH of Exon 1 attacks 3’ splice site

Question 30

Introns are removed in what order, if any?

Answer 30

Based on accessibility to splicing apparatus

Question 31

Different protein (X) usually encoded in separate (introns/exons). That’s why one gene can produce multiple mRNAs via:

Answer 31

X = domains

Exons;

Alternative splicing

Question 32

Where is fibronectin found?

Answer 32

On cell surface and in plasma

Question 33

Including exon 5 in mRNA for fibronectin will have what effect?

Answer 33

Will include transmembrane domain, producing cell surface protein (plasma fibronectin can’t have this domain)

Question 34

What allows alternative splicing to occur?

Answer 34

Different tissues have different sets of RNA binding proteins

Question 35

Function of RNA binding proteins

Answer 35

Facilitate alternative splicing by binding (shielding) exons and preventing the binding of splicing apparatus

Question 36

SR, aka (X), protein (favors/inhibits) splicing.

Answer 36

X = serine/arginine

Favors

Question 37

HnRNP, aka (X), protein (favors/inhibits) splicing.

Answer 37

X = heterogenous nuclear ribonucleoprotein

Inhibits

Question 38

(Exons/introns) have splicing (enhancers/silencers) that bind:

Answer 38

Both have both! Splicing enhancers bind SRs and silencers bind hnRNPs

Question 39

Beta Thalassemia is a result of (X) mutation in (exon/intron) of (Y) gene. Why is this problematic?

Answer 39

X = Single point mutation (G to A)

intron

Y = beta-globin

This results in new 3’ splice site (mis-splicing) and truncated beta globin protein

Question 40

High Ras levels results in (low/high) (X), which affects (SR/hnRNP) binding to (Y) in CD44 Gene. What is the result?

Answer 40

High
X = SAM

Prevents hnRNP binding

So V5 exon included in gene (cancerous)