Lecture 6: Stages 3, 4, 5

Question 1

Describe a productive infection.

Answer 1

infection of a cell that results in virus replication.

Question 2

What is viral titer?

Answer 2

The amount of viral load within each cell. Increases by several orders of magnitude.

Question 3

What is a persistent infection?What are the two types of persistent infections?

Answer 3

An infection that allows the virus and host to coexist. The two types are true viral latency and chronic infection.

Question 4

What are key ideas about true viral latency?

Answer 4

The virus is dormant. The viral genome is inactive. A specific transcribed gene typically triggers latency. The host cell functions as normal. This can turn “on and off,” meaning the virus will go through stages of latency, then reactivation leading to viral production, followed by more latency.

Question 5

What are key ideas about chronic viral infections?

Answer 5

The virus is being replicated. Viral particles are being released by infected cells. There is no “latency gene.” Infected cells may enter a resting state where they will stop dividing enter clinical latency. The SYMPTOMS are latent, not the virus replication.

Question 6

What is the difference between true viral latency and clinical latency?

Answer 6

Viral latency = Virus is NOT ACTIVE

Clinical latency = SYMPTOMS ARE NOT ACTIVE

Question 7

What is genome integration? How is it related to persistent viral infections?

Answer 7

Viral genome can integrate into host genome during both types of persistent infections; but viral genome stays silent.

Question 8

What is genome integration called in bacterial cells?

Answer 8

In bacterial cells, genome integration into host genome is called lysogeny.

Question 9

What is genome integration called in animal/plant cells? What are the two classifications of viruses that can do genome integration?

Answer 9

If it’s non-oncogenic viruses, it’s just called genome integration. If it’s an oncogenic viruses, it can be called oncogenic transformation.

Question 10

What is apoptosis?

Answer 10

Programmed cell death. Individual cells will suicide, energy is required for this process. Cells will shrink, collapse, and contents will be contained. This process activates the immune system without releasing the virus.

Question 11

What is necrosis?

Answer 11

Mass cell death. Groups of cells are killed by some injury, cancer, inflammation, or infection. Cells will swell, rupture, then release its contents. The cell will try to delay necrosis.

Question 12

What is the biggest difference between spherical and cylindrical virus assembly (step 6)?

Answer 12

In spherical viruses, the empty capsid and genome are generated separately. Then, the genome is packed with a molecular motor, requiring energy. Cylindrical viruses pack their genome and assemble their capsid SIMULTANEOUSLY.

Question 13

Describe how cylindrical viruses assemble? What do their capsids look like?

Answer 13

Rod-like capsid. Genome is folded in the center, and lockwasher protein subunits bind and extend to the growing spiral surrounding it. Capsid formation is complete when all the RNA is packaged.

Question 14

Define a protomer

Answer 14

The smallest repeating structural unit of a capsid with a defined stoichiometry. Individual proteins come together to create one protomer.

Question 15

Define a capsomere/capsomer

Answer 15

What forms when protomers are put together. The intermediate geometric subunit.

Question 16

Define a penton, octon, and hexon.

Answer 16

A capsomere made of 5 protomers, 8 protomers, and 6 protomers. In a soccer-ball shaped capsid, pentons are knitted together with hexons (detail).

Question 17

Define a procapsid

Answer 17

What forms when capsomeres are put together. The highest order of geometric subunit.

Question 18

Define a concatemer

Answer 18

A long string of multiple copies of a genome linked in series. Unavoidable product of rolling circle DNA replication.

Question 19

How do bacteriophages pack their genomes into pre-formed heads?

Answer 19

Genome concatemer will pack a ‘headful’ of DNA into the partially formed head, the concatemer is cleaved, then the remaining tail and tail fibers are added.

Question 20

What is the first step in assembly of a large icosahedral virus (ie. bacteriophage)?

Answer 20

Procapsid will be built around an internal scaffolding protein. At its vertex will be a molecular motor. Motor is made of portal proteins and pilot proteins. Pilot protein guides genome through the portal, later forming a plug. Lastly, terminase binds to the vertex of the procapsid, bringing with it the viral genome.

Question 21

What is the second step in assembly of a large icosahedral virus?

Answer 21

The internal scaffolding proteins will degrade (proteolysis) SIMULTANEOUSLY as the portal protein starts to pump genome in via ATP hydrolysis.

Question 22

What is the third step in assembly of a large icosahedral virus?

Answer 22

When the head is full, temrinase will cut the genome concatemer and release from the capsid.

Question 23

What is the fourth step in assembly of a large icosahedral virus?

Answer 23

Tail proteins are added, leading to a mature phage.

Question 24

Describe the structure of the lockwasher.

Answer 24

It consists of 17 long protomers arranged into a single disk. Two disks form a capsomere, called the washer. Upon the correct conditions, the washer will dislocate and look like two turns of a protein helix.

Question 25

Describe the capsid of a rod-like virus.

Answer 25

Capsid consists of the protomer in one long protein helix plus one molecule of single-starnded RNA.

Question 26

What happens to the capsomer double disk in a test tube at pH 7 (no genome)?

Answer 26

The washer is formed, but no assembly occurs.

Question 27

What happens to the capsomer double disk in a test tube at pH 5?

Answer 27

The capsomer SPONTANEOUSLY dislocates into a lockwasher, looking like two turns of a protein helix. These are stackable and can form into a long, continuous protein helix.

Question 28

What happens to the capsomer double disk in a test tube at pH 7 (with genome)?

Answer 28

The dislocation of the first washer SPONTANEOUSLY occurs by contact between the washer hole and the initiation region of the DNA, about 900-1300 bases from the 3’ end of the genome. The first lockwasher nucleates the sequential stacking of the next washer. The 5’ end is drawn upward until all the RNA is packaged.

Question 29

Define egress

Answer 29

Virus exit

Question 30

How does a phage virus egress from the host cell?

Answer 30

Produces lysozyme to lyse bacterial cell wall from inside. Necrosis and expulsion of virions.

Question 31

How does an animal virus egress from the host cell?

Answer 31

Necrosis for non-enveloped viruses. Budding for enveloped viruses, where spike and matrix proteins are produced and expressed on host plasma membrane in a patch.

Question 32

How does the host plasma membrane become virus-specific?

Answer 32

Virus-encoded transmembrane proteins are synthesized and packaged using the normal ER/golgi apparatus.

Question 33

What are polarized cells? How are they involved in viral infection?

Answer 33

A cell where its upper and lower surfaces contain different amounts of plasma membrane proteins. This is common in respiratory epithelium, where the virus enters from one surface and buds from another.

Question 34

What is freeze fracture EM? What does this technique show?

Answer 34

A stained-EM sample preparation technique that images MEMBRANE SURFACE TOPOGRAPHY. Freeze sample, subject it to IMPACT, membranes will fracture to divide the inner and outer leaflet of phospholipid bilayer. Transmembrane proteins stay with one leaflet. Thus, this method also reveals RESOLUTION of SINGLE TRANSMEMBRANE PROTEINS.

Question 35

What are the steps involved in freeze fracture EM?

Answer 35

1. Freeze sample.
2. Split the membranes.
3. Cover w/ thin film of heavy metal.
4. Deposit carbon to add structure.
5. Degrade original sample w/ bleach.
6. Wash metal/carbon replica.
7. Image ONLY the replica via EM

Question 36

How is freeze-etch EM different from freeze-fracture EM?

Answer 36

Freeze-etch includes an etching step right before metal is deposited. This sharpens the image by etching on fake ice crystals from the fracture surface. Can reveal actin cytoskeleton pushing out mature viruses.