Lecture 7- Genome replication- RNA viruses mRNA production

Question 1

Review question: what is the first problem eukaryotic viruses must solve?

Answer 1

The end problem in replication

Question 2

What is the second problem for eukaryotic viruses?

Answer 2

Second problem is producing mRNA from viral genome (first AUG rule)

Question 3

Viruses with divided genomes- 2 types

Answer 3

2 types where genomes come in pieces

1) Segmented genomes: get genome segments. Within each virion there is a series of segments all found in capsid. Reovirus has 10 segments, influenza virus has 8
2) Multipartite genomes. Each genome segment separately encapsidated. Genome is the sum each segment carried in separate particles. All three of these need to be present for infection. Examples: alfalfa mosaic virus (tripartite), cowpea virus (bipartite)

Question 4

Replication vs transcription: How are DNA templates replicated transcribed vs RNA templates?

Answer 4

DNA templates: replication by DNA polymerase, transcription by RNA polymerase
RNA templates: Replication by RNA polymerase/replicase, Transcription by RNA polymerase (transcriptase)

Question 5

RNA polymerases and RNA synthesis

Answer 5

DNA templates have a promoter, which tells which way to go (left or right)
Have a DNA-dependent RNA polymerase. Top is template b/c it is antiparallel to strand being synth
Nascent RNA chain grows out, original two chains rebase pair behind growing chain as moves to the right, still have about 12 bases paired to DNA strand. Nascent RNA displaced as you move.

Question 6

Viral RNA Replication

Answer 6

Genome runs 5’ on left, to 3’ on right. Antigenome (synthesized from genome) runs right to left: 5’ on right, 3’ on left. Antigenome is int complementary RNA. Complementary sequence runs in opposite direction in terms of polarity. Then make genome again (pattern). No end problem in RNA replication because RNA polymerase does not require a primer.

Question 7

RNA polymerases and RNA synthesis

Answer 7

RNA template (strand) acted on by RNA-dependent RNA polymerase. RNA polymerase initiates new chain by assembling new nucleotides depending right at 3’ end and going from left to right. As it continues, nascent RNA chain becomes displaced. With DNA, made sense that nascent RNA chain was being displaced- why does it get displaced here?

Question 8

Synthesis of Viral mRNAs- Initial considerations

Answer 8

1) To program the cell to make proteins, viruses must synthesize mRNAs
2) The mRNAs must conform to the rules dictated by the host cell ribosomes
3) The importance of the first mRNA (the aug problem)
Viruses are completely dependent on cellular ribosomes to make proteins so must make mana that conforms to the rules dictated by the host cells
Ribosomes more difficult for eukaryotes than prokaryotes.

Question 9

Baltimore Classification Scheme

Answer 9

For each virus, there will be three considerations concerning the relationship between the viral genome and the synthesis of mRNA

1) Is the viral genome itself an mRNA?
2) If not, can the cell synthesize an mRNA starting from the viral genome?
3) If not, then the virus must provide the transcriptase (an RNA pol) that is carried within the virion.

Question 10

Baltimore Classification Scheme Diagram:

Answer 10

Draw out on white board

Question 11

Synth of viral mRNAs summary

Answer 11

1) Generally cellular RNA polymerases synthesize the mRNA for viruses with ds or ss DNA genomes (poxviruses an exception)

Question 12

Isolated nucleic acid genome not infectious if:

Answer 12

1) essential enzymes are enclosed w/in capsid
2) the genome of the virus is segmented
3) DNA genome is too large to remain intact during isolation