15. Adenovirus 2

Question 1

what is pTP and its function?

Answer 1

pre-terminal protein which primes replication

Question 2

describe the first phase of adenovirus replication (3 steps)

Answer 2

1. pTP binds C on the end of DNA opposite of TP
2. DNA pol starts replicating and displaces a ssDNA
3. produces 1 dsDNA and 1 displaced ssDNA associated with DNA binding protein

Question 3

describe the second phase of adenovirus replication (4 steps)

Answer 3

1. the displaced ssDNA folds on itself at the inverted terminal repeats to make pan-handle structure
2. a new pTP joins at a C opposite the TP
3. DNA pol replicates and displaces the DNA binding protein
4. produces another dsDNA

Question 4

what happens once the genome is replicated? why?

Answer 4

late stage proteins can be made bc the late proteins are made from this new DNA

Question 5

how many late proteins are there?

Answer 5

14

Question 6

how many promoters are there for the late proteins?

Answer 6

1 major late promoter

Question 7

how is it possible that there are 14 late proteins but 1 promoter? why does this let the virus be more efficient

Answer 7

ALTERNATIVE SPLICING!

can pack a lot of info into genome

Question 8

describe the 5 steps of alternative splicing

Answer 8

1. DNA transcribed to make 1 long RNA transcript
2. transcript is poly-adenylated by cellular machinery at 5 different sites
3. this gives different splice sites where transcript is spliced by cellular machinery
4. tripartite leader is assembled
5. many proteins can be made

Question 9

does the machinery for poly-adenylation and splicing come from the virus or the cell?

Answer 9

cell!

Question 10

where is the packaging sequence in the genome?

Answer 10

at the beginning

Question 11

which mRNA have the tripartite leader?

Answer 11

tripartite leader is spliced onto every late mRNA

Question 12

What do IVa2 and L2 52/55k do?

Answer 12

join to form ATPase driven motor

Question 13

what is the role of L4 33K/100K?

Answer 13

viral chaperones/scaffolds for capsid assembly

Question 14

can adenovirus capsid self-assemble?

Answer 14

no, requires chaperones and scaffold proteins

Question 15

what is the role of E4 Orf4 and E3 11.6K?

Answer 15

to induce apoptosis once the viral life cycle has completed to leave the cell

Question 16

how does E1A induce apoptosis?

Answer 16

1. E1A binds Rb to allow E2F promoter to be active
2. p14 ARF is produced
3. p14 ARF binds MDM2 to stabilize p53
4. stabilized p53 = apoptosis

Question 17

what is MDM2 function? how does p14 ARF affect it?

Answer 17

ubiquitinates p53 so it can be degraded

but p14 ARF binds MDM2 to prevent p53 from being degraded

Question 18

which oncogene inhibits p53?

Answer 18

E1B-55K

Question 19

how does E1B-55K inhibit p53? what does this result in?

Answer 19

sequesters p53 to inhibit it –> no apoptosis, cell can survive

Question 20

E1A and E1B vs E6 and E7

Answer 20

E1A = E7

E1B = E6

Question 21

what are oncolytic virus?

Answer 21

genetically engineered viruses that can induce immune system and kill tumour cells

Question 22

what is the other type of E1B?

Answer 22

E1B-19k

Question 23

what is the role of E1B-19K

Answer 23

inhibits Bax in mitochondria to inhibit apoptosis

Question 24

what does Bax do? how is it upregulated?

Answer 24

Bax forms pores at mitochondria to eventually induce apoptosis

upregulated by p53

Question 25

do E1B-55K and E1B-19K work together or is only 1 used?

Answer 25

both work together to maximize inhibition of apoptosis

Question 26

what type of RNA is VA RNA?

Answer 26

non-coding RNA

Question 27

what are the 3 functions of VA RNA?

Answer 27

1. inhibit PKR activation 2. block nuclear export of pre-mRNAs 3. inhibit dicer

Question 28

what is PKR? how does VA RNA affect it?

Answer 28

detects virus infection by binding the dsRNA - Phosphorylates itself then phosphorylates eIF2a to inhibit translation VA RNA blocks this --> translation can continue

Question 29

why does PKR recognize a viral infection?

Answer 29

dsRNA is only present in a cell when a virus has infected it

Question 30

what is the role of E3?

Answer 30

immunosuppression to make infected cell bypass immune system

Question 31

can we see E3 function in cell culture?

Answer 31

no, it only works in an in vivo system where the immune system is present

Question 32

what are 3 functions of E3?

Answer 32

1. internalize TNF/fas receptors so cell can't recognize virus 2. inhibit MHC complex so cell can't present peptides 3. makes Adenovirus Death Protein so virus can easily be released from cell