Lecture 6: Gene expression and human disease

Question 1

What is the central dogma in bioligy?

Answer 1

DNA is transcribed to mRNA which is translated to make proteins

Question 2

How is transcription initiated?

Answer 2

Transcription can often only start when the correct complex of transcription factors and other components come together. DNA may need to loop around for some regulatory transcription factors to bind.

Question 3

Describe the process of transcription

Answer 3

1. Transcription factors bind first at the 5’ end
2. RNA polymerase binds alongside other factors
3. C-terminal domain of RNA polymerase II is initially un-phosphorylated but it becomes phosphorylated when RNA polymerase needs to transcribe DNA
4. RNA polymerase moves along the DNA unwinding the strand
5. DNA exposed to complimentary base-pairing by RNA nucleotides to form mRNA.

Question 4

What are the 3 modification that take place place in transcription to the mRNA strand?

Answer 4

1. Capping (addition of guanine triphosphate (GTP) molecule to 5’ UTR)
2. Splicing (removal of introns)
3. Polyadenylation (addition of a poly-A tail to the 3’ UTR).

Question 5

What are introns?

Answer 5

Non-coding regions of DNA

Question 6

How is splicing initiated?

Answer 6

AG at the 3’ end of one exon is spliced to G at the 5’ end of the next exon. Introns are removed by a complex of RNAs and proteins called the spliceosome.

Question 7

What confers the accuracy of splicing?

Answer 7

Accuracy of splicing is conferred by small nucleolar RNAs (snRNAs) within the spliceosome.

Question 8

What is alternative splicing?

Answer 8

Removal of introns which means exons can be spliced together in different ways to produce different proteins.

Question 9

What are 3 diseases associated with incorrect splicing?

Answer 9

1. Beta- thalassemia
2. Cystic Fibrosis
3. Familial isolated growth hormone deficiency type II

Question 10

Beta-Thalassemia?

Answer 10

• Caused by aberrant processing which results in incorrect splicing leading to premature stop codon; removal of exon 3
• Leads to anaemia in young children which requires frequent blood transfusion

Question 11

Cystic Fibrosis?

Answer 11

• Caused by incorrect splicing that results in the removal of exon 5 coding for phenylalanine
• Leads to thick mucus build up in lungs which increases risk of infection

Question 12

Familial isolated growth hormone deficiency type II ?

Answer 12

• Caused by mutations in the growth hormone gene (GH-1) as a result of loss of exon 3; mRNA shortening.
• Leads to short stature.

Question 13

What happens to DNA after it is transcribed into mRNA?

Answer 13

mRNA is transported out of the nucleus and into the cytoplasm for translation.

Question 14

What is a reading frame?

Answer 14

The combination of 3 nucleotides in an mRNA sequence. The RNA code has 3 “reading frames” and each different reading frame encodes a different protein.

Question 15

What is the start codon found on every mRNA molecule that initiates translation?

Answer 15

AUG: Methionine

Question 16

Describe the process of translation

Answer 16

1. Ribosome scans along the RNA from the 5’ cap for the start codon (AUG).
2. Aminoacyl-tRNA binds to A site of ribosome and a peptide bond is formed between the amino acid and the growing peptide chain.
3. tRNA moves to P site and allows another aminoacyl-tRNA to enter A site and form peptide bond.
4. When initial tRNA reaches E site, the tRNA is ejected and allowed to migrate off to attach to another amino acid.

Question 17

How is translation initiated?

Answer 17

Small (40S) ribosomal subunit scans along the RNA from the 5’ cap for the start codon (AUG) and carries the initiator tRNA and initiation factors. Initiation factors dissociate allowing for binding of large (60S) ribosomal subunit.

Question 18

How are aminoacyl-tRNA molecules formed?

Answer 18

AAs and tRNA molecules are bound together by tRNA synthetase in a process which requires energy from ATP.

Question 19

What are the 6 areas in which gene expression is controlled?

Answer 19

1. Transcriptional control
2. RNA processing control
3. RNA transport and localisation control
4. mRNA degradation control
5. Translation control
6. Protein activity control

Question 20

What is an adaption of antibiotics that capitalises on the process of transcription and translation?

Answer 20

Some antibiotic are designed to bind to binding sites on bacterial ribosomes, exploit structural and functional differences between the eukaryotic and prokaryotic ribosome, and inhibiting gene expression of bacteria to ultimately treat infection.

Question 21

Describe the action of different antibiotics and how they capitalise on the processes of transcription and translation

Answer 21

1. Tetracycline- blocks binding of AA-tRNA to A site.
2. Streptomycin- prevents transition from initiation complex to chain elongating ribosome.
3. Chloramphenicol- blocks peptidyl transferase reaction on ribosome.
4. Cycloheximide- blocks translocation reaction on ribosome.
5. Rifamycin- binds to RNA polymerase.

Question 22

What are the 3 types of mutations in RNA that affect translation of a protein?

Answer 22

1. Frameshift mutations
2. Missense Mutations
3. Nonsense Mutations

Question 23

What are frameshift mutation?

Answer 23

Mutations that result in the insertion or deletion of any number of nucleotides that is not a multiple of 3 which alter the triplet reading frame
Example: Congenital deafness

Question 24

What are missense mutations?

Answer 24

Mutations which results in the wrong amino acid being incorporated into a protein.

Example: Sickle cell anaemia

Question 25

What are non-sense mutations?

Answer 25

Mutations that causes a protein to terminate or end its translation earlier than expected.

Example: Beta-thalassemia

Question 26

What are microRNAs (miRNAs)?

Answer 26

MicroRNA (miRNAs) are 21-22nt non-coding RNAs that control gene expression and translation by binding to target sequences in 3’ untranslated region of mRNAs.

Question 27

What is the importance of microRNAs?

Answer 27

Loss of microRNAs can cause disease such as cancer (eg. CLL) and the upregulation of some microRNAs can promote metastasis of cancerous cells. MicroRNAs can also be beneficial and prevent metastasis of cancers. MicroRNAs can also be used as biomarkers.