Retrovirus Replication

Question 1

What are retroviruses?

Answer 1

Enveloped RNA viruses, linear, single stranded +sense RNA with 5’-caps and polyA tails.

Question 2

Give two examples of retroviruses

Answer 2

1. HIV - lentivirus

2. Murine leukaemia virus - simple virus

Question 3

What does the genome have?

Answer 3

5’cap - direct repeat sequence - 5’unique sequence - primer binding site - viral RNA - poly purine tract - 3’unique sequence, direct repeat sequence - polyA tail

Question 4

What initiates reverse transcription?

Answer 4

Annealing of a partially unwound transfer RNA to the primer binding site.

Question 5

Which tRNA does HIV utilise as a primer?

Answer 5

tRNAlys3

Question 6

What are the two enzymatic activities of HIV-1 reverse transcriptase?

Answer 6

1. RdDp

2. Ribonuclease H, degrades the RNA strand of the RNA:DNA hybrid

Question 7

How is the genome copied?

Answer 7

1. tRNA binds to PBS at the 5’end and copies the U5 and direct repeat.
2. RNA is degraded
3. Sequence is complementary to 3’-end so moves and synthesis occurs.
4. PPT is resistant to reverse transcriptase degradation but is eventually degraded
5. Replication occurs from U3 until positive strand strong stop
6. Circularises and then replicates

Question 8

Why is the copy longer than the original?

Answer 8

Because of the duplication of U3 and U5 on either side

Question 9

What is the preintegration complex?

Answer 9

Nucleoprotein complex containing proviral DNA, capsid and integrase.

Question 10

When can a retroviral preintegration complex enter the nucleus?

Answer 10

At cell division, nuclear membrane breakdown

Question 11

what does the HIV PIC contain?

Answer 11

Matrix, integrase, RT and Vpr. No capsid so can enter the nucleus whenever due to nuclear localisation signals on the matrix

Question 12

What does integrase to?

Answer 12

Catalyses a series of independent events that result in the cleavage of host DNA and splicing of provirus into the cleavage site

Question 13

Why does integrase have exonuclease activity?

Answer 13

Makes a staggered break with a 5’overhang. The proviral 5’overhang is then removed by host cell enzymes

Question 14

What does the 5’LTR act as in the integrated provirus?

Answer 14

Enhancer/promoter

Question 15

What does the 3’LTR act as in the integrated provirus?

Answer 15

Mediates polyadenylation of RNA transcripts

Question 16

What is removed in mRNA splicing?

Answer 16

Gag and Pol sequences

Question 17

What are endogenous retroviruses?

Answer 17

Result from a germ line infection which are generaly defective due to mutations and/or deletions so they cannot produce an infection virus.

Question 18

how are HERVs named?

Answer 18

After the tRNA primer

Question 19

How can we estimate the time of HERV insertion?

Answer 19

Assume two LTRs are identical when integrated, usually show 3-12% divergence. most recent was HERV-K113 (1mil y.a)

Question 20

Give 2 benificial effects of HERVs?

Answer 20

1. Ability to digest starch to simple sugars due to insertion of a provirus upstream of duplicated copy of the pancreatic amylase gene
2. Development of placenta, expressed in placental syncytiotrophablasts which have membrane fusion activity important in placental development

Question 21

Give 2 detrimental effects of HERVs

Answer 21

1. Association with autoimmune diseases and cancers, type 1 diabetes
2. Behavioral effects, HERV-K(HML-2) integrated near a gene which regulates dopaminergic activity - addictive traits