Chapter 2 - V

Question 1

Key steps in virus replication

Answer 1

1. Virus adsorption and attachment to susceptible host, occurs by chance
   - Adsorption: 2 solid things bind tgt
2. Inject genomes into cell - genome enters cell and any other needed proteins
3. This step varies
   - stnthesis of viral mRNA
   - Synthesis of viral proteins
   - Synthesis of viral genome (for next generation?)
4. Assembly of virus particle from parts
5. release from host

Question 2

Virus structure

Answer 2

All virus genomes are surrounded by proteins

the virus particle is a gene delivery system - it moves the virus’ genome from one host cell to another host cell

The genome is packaged in a protein known as capsid

Some viruses, the virion consist of only genome and the capsid (nucleocapsid, capsid with enclosed genome)

others may have an additional layer

• lipid bilayer envelope
• layer of protein between capsid and envelope (matrix)
• internal lipid laters
• protein occlusion bodies

Question 3

Capsid protein

Answer 3

Capsid proteins are structural proteins

the function of the proteins that make up the capsid include:

1. protection of chemically labile virus genome
   - from nucleases in the environment
   - from agents that might damage the base of nucleotide
   - shearing of viral genome from mechanical forces
2. Delivery of the virus’ genome to the location within the cel where the genome can be replicated and expressed

if virus is naked particle, the capsid has the structure to bind to host cell

If enveloped, the envelope has the structure to bind to host cell.- capsid does not have the correct structure (aka if envelope gone, no invasion)

Question 4

Step 1: Virus - host cell Invasion

Answer 4

Adsorption and attachment to susceptible host:

Virus and host cell must have complementary structures to be able to interact - chance event (recognition specificity)

Host cell has a virus receptor - a normal cell structure that virus exploits as a binding site

the cell structure can be - protein or carbohydrate attached to a protein

Virus has an anti-receptor -
- structure that bind to host cell

A host is susceptible if it has the proper virus receptor

Question 5

Step 1: Virus entry

Answer 5

The virus’ genome and replicative enzyme needs to enter the cell

Different strategies for different viruses,
but a given species of virus always use the same method

After binding to cell’s virus receptor, for some viruses

• only genome enters
• the whole virus enters the cell in an endosome
• the virus envelope fuses to the cell membrane releasing the genome into cytoplasm

Question 6

Virus entry is necessary cuz

Answer 6

Viral genome needs to gain access to hosts translation machinery for replication

Some genomes have their own transcription machinery but no virus has translation machinery

Question 7

Virus entry

Answer 7

Virus particle itself is inert with respect to its metabolism

The virus particle is designed to interact with host cell receptor to initiate the process of infection

for virus particles, the capsid is stable enough to persist in the environment but unstable enough to allow the release of the genome into cells cytoplasm

Question 8

What type of interactions exist between the protein subunits in the capsid

Answer 8

interactions with other proteins

changes in pH

cleavage of proteins

Unfolding proteins

Question 9

Susceptibility and permissiveness

Answer 9

For viruses to become successful in nature, it must encounter host cell that is both susceptible to viral infection and permissive to viral replication

Susceptibility

• host cells structure - viral receptor that specific virus can attach to for delivering the virus and its genome into the host ell
• virus has an anti-receptor structure that recognizes the virus receptor on the cell

Question 10

Susceptibility and permissiveness - polio eg

Answer 10

poliovirus in human cells and mouse

• polio virus known to only infect human cells
• certain human cells express protein CD155 (aka poliovirus receptor) also act as virus receptor for polio

Polio binds to CD155 infecting them and kills them - increase in virus particle (spreads)

While in mouse, they have their own version of CD155, but slightly different than humans

• polio still cannot bind to mice CD155, so it cannot infect mouse cells
• mouse cell survives and three is no increase in number of virus particles

Question 11

Susceptibility and permissiveness: Are mice really resistant to polio or not permissive to its replication

Answer 11

Determining if mouse cells are permissive:

isolate the mRNA from poliovirus and then find a way to introduce them into mouse cell

results: poliovirus are produced in both human and mouse cells
- mouse are permissive to polio replication even though they are not susceptible

Question 12

Permissive

Answer 12

Ability to replicate the viral genome in the cell and

produce more viral particles

Question 13

Susceptible

Answer 13

Ability of a given virus to infect

- virus receptor fits with anti receptor

Question 14

Determining if mouse cells are permissive:

-II

Answer 14

Another experiment that we can do to confirm mouse cells permissiveness to polio

• transfect the mouse cells with the gene for the human version of CD155
• Transfecting means that we introduce a gene so that it can be expressed by the new cell - they have a transgene
• allow mouse cells to have human CD155 as well
  result: transfected mice are susceptible to polio - we see an increase in the number of the virus particle

Question 15

Susceptibility and permissiveness

Answer 15

For viruses to be successful, they need to:

• infect susceptible and permissive host
• package functional enzymes involved in the replication of its genome
• be able to remain intact and capable of infecting a new host when it is in the environment ( exposure to the environment should not damage the virus in such a way that it can no longer infect new host)
• virus must have multiple structures to interact with the surface of the host cell - ensures that in the event that some are damaged (by oxidation/enzymes) the virus can stull infect a new cell

Question 16

Susceptibility and permissiveness

Answer 16

Susceptibility and permissiveness can be demonstrated experimentally

The virus that persist in natures have been successful at encountering host cells that are both susceptible and permissive

Question 17

Baltimore classification of viruses

Answer 17

based on how viral mRNA is made

- group by starting material dna or RNA ds or ss

Question 18

Key concepts - review

Answer 18

Polymerases work the same way in all cells - applies to both DNA and RNA pol

• All read a template in the 3-5’ direction
• all synthesize a complementary strand in the 5’-3’ direction

Question 19

Differences between viruses and cells

Answer 19

Cell template is always DNA, virus can be DNA or RNA

Cells does transcription in nucleus,

Question 20

Key concepts - review

Answer 20

All ribosomes work the same way

• read mRNA template 5-3’ direction
• Synthesize the polypeptide from N to C direction

with viruses they differ in how they start process of translation:

can be cap-dependent or cap independent

Question 21

Viral Replication Enzymes

Answer 21

Some viruses package their own replicative enzymes in their capsids for 2 main reasons

1. some species of viruses replicate their genomes in the cytoplasm of the cell
   - can’t use cell’s DNA or RNA polymerases that are located inside the nucleus
   - these viruses also must encode the genes for the polymerase
2. For RNA viruses, the Cell’s RNA polymerase cannot read RNA template (only DNA)

All RNA viruses must have in their genome a gene that encodes a polymerase that can read RNA as a template and synthesize RNA or DNA - some even have a functional enzyme copy readily inside the capsid
(RDRP)

Question 22

Viral Genomes

Answer 22

RNA viruses need an enzyme that can read RNA as a template and make RNA for both their genomes and mRNA - excluding RT

The cell do not have a polymerase that can read RNA as a template

The virus must have a gene that encomds such an enzyme:
+ sense RNA can synthesize it right after infecting the cell- sense RNA viruses must have some packaged within their capsid

RNA dependent RNA polymerase

Question 23

+ and - Sense

Answer 23

normal mRNA is defined as being + sense (same as host cell mRNA

+ sense is like normal mRNA can be translated right away and the RDRP produced and cango back to transcribe the - sense mRNA from the + sense and then more + again

While - sense needs to have RDRP packaged with it so it can use to make the + sense

Question 24

Retroviruses

Answer 24

RT use an alternative approach –> transcribe the ssRNA to dsRNA first

dsDNA is transcribed to makes more of the ssRNA viral genome

need enzyme that reads RNA and synthesizes DNA = RT

Question 25

RT

Answer 25

RT has 3 distinct enzyme activities

1. RNA dependent DNA polymerase
   - result in DNA-RNA hybrid molecule
2. RNaseH
   - an enzyme that removes RNA in the DNA-RNA hybrid molecules
3. DNA dependent DNA polymerase
   - makes the complementary DNA strand so that the ssDNA becomes ds DNA

Question 26

mRNA synthesis

Answer 26

DNA viruses that replicate their genomes in cells nucleus can use the cells DNA polymerase
- DNA viruses can then use the cells RNA polymerase to transcribe their DNA

Retroviruses that have copied their genome to DNA can use the cell’s RNA pol to transcribe their DNA
- so now they can make multiple copies of that RNA

All DNA viruses that replicate in the cytoplasm and are all RNA viruses must encode their own RNA polymerase (except Retroviruses cuz they make DNA then use RNA pol in nucleus)

• they must package itw tih genome

Question 27

Translation of viral m RNA

Answer 27

All viruses translate their mRNA (+sense RNA) on the cell’s ribosome

Eukaryotic ribosomes like:

• mRNA with 5’ cap, start codon, single ORF, stop codon, poly A tail
• cellular mRNA has these more favorable

Sometimes assembly of the mRNA on the ribosome is a big challenge, eukaryotic ribosomes are designed to translate eukaryotic mRNA not viral mRNA

some types of viruses have mRNA without the 5’cap - they use secondary structure instead
- not as efficient as 5’ cap but adequate

although some viruses do have 5’cap and poly-A tail

Question 28

Translation of viral m RNA

Answer 28

monocistronic mRNA - one gene

Polycistronic mRNA multiple genes, multiple reading frames (multiple start stop codon)
- common in bacteria/viruses

Question 29

Translation of viral m RNA

Answer 29

Eukaryotic ribosomes are not good at translating polycistronic mRNA

Only 1 gene would be translated. the ribosome would dissociate at the first stop codon and would not reassemble on the downstream start codon
- not good for viruses that need to make several proteins

Prokaryotic ribosomes might be able to reassemble on such mRNA but eukaryotic ribosomes cannot
- the cap is essential in ribosome assembly on mRNA

In cap-independent translation, a secondary mRNA structure takes place of the 5’ cap

Question 30

Translation of viral m RNA

Answer 30

Viruses have evolved a simple strategy to overcome the problem of translating multiple proteins in a eukaryotic cell

• entire mRNA is translated into a polyprotein, which gets cleaved up into several individual proteins
• one ORF (like operons

Question 31

SARS - CoV -2

Answer 31

Coronaviruses have one ss + sense RNA as their genome

they replicate and express their genomes in the cytoplasm of the infected cell
- they cannot use host cells RNA pol to replicate/transcribe their genomes
We would not find RDRP packaged in capsids cus plus sense