22: W2 The mRNA and the tRNA transcript and its processing

Question 1

What are the three main parts of the mature mRNA?

Answer 1

-Protein coding region
-Two untranslated region
(5’ = left, 3’ right)

mRNA vary significantly in size since proteins vary significantly in size

Each untranslated region contains elements which regulate the expression of information in the coding region

Question 2

What are the key features of the typical eukaryotic mRNA

Answer 2

Draw this out (look through notes)

But label these key features
-5' cap
-5' UTR
-Coding region
-3' UTR
-3' Poly(A) tail 
  important stabilising component

Question 3

What are the key features of a typical prokaryotic mRNA?

Answer 3

Draw this out (notes)

Key features:

• Majority is protein coding
• Only has short UTR on either side
• No 5’ cap
• No 3’ Poly(A) tail
• Prokaryotic RNA has a half life of 2-4 minutes

Question 4

How are prokaryotic mRNA translated?

Answer 4

Prokaryotic mRNA is ready to be translated into protein -no further processing is required.

In prokaryotes translation can start before transcription is finished -this is called coupled transcription and translation

Question 5

Describe how eukaryotic mRNA is translated

Answer 5

Requires three things to occur before translation can begin:

1. Addition of the 5’ cap
2. Addition of the 3’ Poly(A) tail
3. Splicing out of introns

These events all occur in the nucleus, prior to export of the transcript to the cytoplasm for translation

Question 6

What types of introns can splice on their own?

Answer 6

Group I and group II introns can splice on their own.

Question 7

Describe the splicing mechanism for group I introns

Answer 7

Look on slides for images –> draw this out:

• The pre-mRNA contains introns
• At either end of the intron there are splice sites (called the 5’ and 3’ splice site)
• The mRNA folds creating a pseudo-active site
• Guanine nucleotide with a free 3’OH comes into the active site and acts as an attacking nucleophile and cleaves at the exon-intron junction
• The guanine ends up attached to the intron
• The 3’OH ends up attached to the exon then acts as a nucleophile and cleaves the other exon-intron junction
• This results in the removal of the intron

Question 8

Describe the splicing mechanisms for group II introns

Answer 8

Look through slides for images –Draw this out

Step 1: Intron bends over, forming a ‘lariat’ structure with the help of 2’OH (acts as a nucleophile) of one of the bases in the intronic region

Step 2: Similar to group I, the 3’OH from the end of the exon post-cleavage can then act as a nucleophile and attack the other exon-intron junction, removing the intron

Question 9

What is the signal which marks the intron exon boundary

Answer 9

AG| cut | GU ———AG| cut |GU

“this is where the cleavage must occur”

Question 10

What does the spliceosome do?

Answer 10

Help the introns splice

Question 11

What are the five subunits of the spliceosome?

Answer 11

U1, U2, U3, U4, U5 and U6

Question 12

Describe U1 of the spliceosome (where it binds and its composition)

Answer 12

Binds to 5’ of splice site

RNA base pairs to the nucleotides at the end of the intron, forms hair pins, 7 core proteins, 3 additional proteins

Question 13

Describe U2 of the spliceosome (where it binds and its composition)

Answer 13

Binds to pyrimidine-rich group (where 2’OH comes from)

There is a little bit of base pairing, a lot more protein than A1

Question 14

Where does U4, U5 and U6 assemble on the spliceosome?

Answer 14

On top of U1 and U2

Question 15

Describe the composition of U4

Answer 15

Inhibitor, base pairs with U6 to prevent unwanted cleavage

Question 16

Describe the composition of U5

Answer 16

base pairs to both of the introns