19.01.07 Transcription, mRNA processing (excluding splicing) and translation

Question 1

Central dogma

Answer 1

The expression of genetic information in a cell is almost exclusively unidirectional. DNA>RNA>protein

Question 2

Where does transcription occur

Answer 2

Nucleus (limited extent in mitochondrial)

Question 3

What direction is RNA synthesised?

Answer 3

5’ to 3’, complimentary to template (anti sense strand) with same bases except U/T.

Question 4

how many classes of RNA polymerase molcules

Answer 4

3 (I, II, III)

Question 5

how does transcription start

Answer 5

-Transcription factors bind to the promoter region and position RNA polymerase to initiate RNA synthesis.

Question 6

Are transcription factors trans- acting?

Answer 6

Yes, migrate to sites of action following synthesis by remotely located genes

Question 7

Are promoters trans-acting

Answer 7

No, they are cis-acting. Function is limited to the duplex on which they reside.

Question 8

What other elements are cis-acting

Answer 8

Other than promoters, enhancers and silencers are also cis-acting

Question 9

Elements of promoters

Answer 9

TATA box, GC box, CAAT box

Question 10

TATA box

Answer 10

TATAAA sequence. 25-35 bp upstream of transcriptional start site. Able to define direction of transcription, indicates the DNA strand to be read. Mutations can cause transcription to begin at an incorrect location

Question 11

GC box

Answer 11

Variant of GGGCGG. 110bp upstream of transcriptional site. Can function in both directions.

Question 12

CAAT box

Answer 12

GGCCAATCT. 80bps upstream. Strongest determination of promoter efficiency. Functions in both directions

Question 13

Mutations in promoters

Answer 13

1% of single base pair substitutions causing genetic disease occur in promoter regions. Disrupt transcription initiation (alter/abolish cis-activing DNA sequence motids for trans acting TFs), leading to altered amounts of mRNA and thus protein.

Question 14

enhancers

Answer 14

Enhance transcriptional activity. Variable distrance from start site. Function independent of orientation. Cause DNA between promotor and enhancer to loop out.

Question 15

silencer

Answer 15

inhibit transcriptional activity

Question 16

Main steps of RNA processing

Answer 16

5’ capping, 3’ polyadenylation

Question 17

5’ capping, when does it occur

Answer 17

During elongation

Question 18

functions of 5’ capping

Answer 18

protect from 5’ - 3’ exonuclease activity. Facilitate transport from nucleus to cytoplasm, facilitate RNA splicing. Attaches 40s subunity of ribosome to mRNA

Question 19

What happens during 5’ capping

Answer 19

methylated nucleoside, m7g (7-methylguanosine), is linked to the 5’ end of RNA by a phosphodiester bond.

Question 20

3’ polyadenylation

Answer 20

AAUAAA (or AUUAAA) signal sequence is sequentially added by pol(A)polymerase, around 200 added, to form a poly(A) tail. Acts as a signal for 3’ cleavage for RNA pol II transcripts

Question 21

Functions of 3’ polyadenylation

Answer 21

Facilitates mRNA transport to cytoplasm, stabilises mRNA molecules, facilitates translation

Question 22

Mutations affecting 3’polyadenylation

Answer 22

rare. Causes read0-through and reduced accumulation of alpha-haemoglobin in haemoglobin H (HbH) disease.

Question 23

rRNA processing

Answer 23

RNA polymerase I makes a large pre-rRNA molecule. Then cut into 3 pieces: 18S, 5.8S and 28S

Question 24

What makes 5S (smallest eukaryote rRNA

Answer 24

RNA polymerase III

Question 25

Translation

Answer 25

mRNA transcribed from genes in nuclear DNA, migrates to cytoplasm and engages with ribosomes, tRNA and other components to direct polypeptide synthesis.

Question 26

Functions of untranslated regions (UTRs)

Answer 26

Stabilise mRNA on ribosomes for translation of the central segment

Question 27

Mutation in 5'UTR

Answer 27

BRCA1 has an alpha and beta promoter which encode transcripts with different 5'UTR lengths. The longer is translated less efficiently. Cancerous breast tissue has the longer 5'UTR transcript so BRCA1 protein expression is inhibited in cancer tissue.

Question 28

Ribosomes

Answer 28

RNA-protein complexes

Question 29

Structure of a ribosome

Answer 29

2 subunits: 60S and 40S subunits.

Question 30

Ribosome 60s composed of

Answer 30

28S, 5.8S and 5S rRNAs, with various ribosomal proteins (50)

Question 31

Ribosome 40s composed of

Answer 31

18S rRNA and >30 ribosomal proteins

Question 32

How many ribosome biding sites

Answer 32

3. A site (aminoacyl-tRNA), P site (peptidyl- tRNA), E site (for tRNA released following peptidyl transferase reaction)

Question 33

What is tRNA

Answer 33

75-95 nt ribonucleotides in length

Question 34

What do tRNAs do

Answer 34

Mediate decoding of mRNA sequence. Have a specific amino acid covalently bound to acceptor arm by amino acyl tRNA synthetases

Question 35

How many types of tRNA synthetases

Answer 35

20 (1 specific to each amino acid).

Question 36

What does the tRNA acceptor arm contain

Answer 36

Sequence to allow discrimitation of tRNAs by tRNA synthetases.

Question 37

tRNA anti codon sequence

Answer 37

This recognises the complimentary mRNA codon.

Question 38

Steps of translation

Answer 38

Initiation, elongation, termination

Question 39

What initiates mRNA translation

Answer 39

Binding of tRNAfmet to P (peptidyl) site.

Question 40

What is then delivered to A (aminoacyl site)

Answer 40

tRNA in complex with elongation factor= EF-Tu-GTP

Question 41

What happens when codon-anticodon pairing occurs

Answer 41

Activates GTPase centre of ribosome, causing hydrolysis of GTP and the release of aminoacyl end of tRNA from EF-TU.

Question 42

What also happens when tRNA binds

Answer 42

COnformation changes in rRNA, optimally orientates the peptidyl-tRNA and aminoacyl-tRNA for the peptidyl-transferase reaction to occur.

Question 43

What happens during the peptidyl-transferase reaction

Answer 43

transfer of the peptide chain onto the A-site tRNA

Question 44

How does the ribosome shift in the 3' mRNA direction

Answer 44

GTPase EF-G casuses the deacylated tRNA at the P site to move to the E (exit) site and the peptidyl-tRNA at the A site to move to the P site upon GTP hydrolysis.

Question 45

When is the deacylated tRNA in the E site released

Answer 45

Upon binding of the next aminoacyl-tRNA to the A site.

Question 46

When does elongation end

Answer 46

when stop codon is reached. No tRNAs complimentary to the stop codon. Leads to hydrolysis of the bond between tRNA and polypeptide at the P site. They are then released and ribosomal subunits and template dissociate

Question 47

How many combinations of codons for amino acids

Answer 47

64 possible codons (4 bases at 3 positions, 4^3) with 20 amino acids

Question 48

Wobble hypothesis

Answer 48

allows interpretation of all 64 codons. 5' end of anticodon can bind any of the several bases at 3' end of codon.

Question 49

When does nuclear-encoded mRNA translation stop

Answer 49

When a termination codon is encountered (UAA, UAG, UGA)