Lecture 3

Question 1

Virus envelopes (composition)

Answer 1

• derived from cell membranes
• composed of lipid bilayers and viral glycoproteins (sugar attached to protein) embedded in the lipid bilayer
• can have a variety of shapes, not always symmetrical
• many are acquired at the cell membrane through budding

Question 2

Virus envelopes (function)

Answer 2

• provides permeability barrier
• controls what goes in and out of the particle
• protects viral genome from the environment
• allows for “less perfect” packaging of proteins

Question 3

Creation of virus envelopes

Answer 3

• through budding
• viral glycoproteins become inserted into the cell membrane
• the nucleocapsid is wrapped in a membrane into which viral glycoproteins have been inserted
  Happens in 2 ways:
  1. nucleocapsid interacts directly with tails of the glycoproteins, also known as viral spikes
  2. matrix proteins are sent to the plasma membrane and interact with the viral spikes and the nucleocapsids

Question 4

Characteristics of envelope glycoproteins

Answer 4

1. large glycosylated (sugar molecules have been attached) domain (ectodomain)
2. Hydrophobic transmembrane anchor domain (helps anchor protein in to the membrane)
3. Short internal cytoplasmic tail

Question 5

Why is the glycosylation of the external domain of the envelope proteins important?

Answer 5

• prevents dehydration of the external surfaces of virus particles
• reduces protein-protein interactions to prevent viral aggregation (individual units come together to form a larger structure)

Question 6

Role of the ectodomain of envelope proteins

Answer 6

binds to cell receptors and mediates fusion between the viral envelope and cell membrane

Question 7

where are envelope proteins synthesized

Answer 7

• synthesized on ribosomes in the ER
• inserted into the plasma membrane via standard export pathways for cell-surface proteins

Question 8

Type 1 integral membrane proteins

Answer 8

• N- terminus facing lumen of ER or extracellular space
• Transmembrane anchor nearest C-terminus

Question 9

Type 2 integral membrane proteins

Answer 9

• C- terminal faces the lumen or extracellular space
• N-terminal near the anchor, faces the cytosol

Question 10

membrane insertion of envelope proteins

Answer 10

• signal sequence on the envelope protein directs it
• Type 1 membrane proteins have N-terminal signal sequence that is cleaved by a peptidase when it is inserted in the ER during synthesis
• Type 2 use the transmembrane anchor as the signal sequence

Question 11

scaffolding proteins

Answer 11

• assist with formation of procapsid (particle that does not have DNA in it)
• not included in the final, mature virion

Question 12

Packaging of the Herpesvirus and other dsDNA genomes

Answer 12

• terminase binds to the viral genome
• terminase attracts to the poral protein at the entrance of the empty procapsid
• once they interact, they start using energy to pull the genomes inside
• ATP is used to insert genome in to the capsid
• once it reaches 1 genome equivalent inside the capsid, it will cut off the rest

Question 13

concatemer

Answer 13

• several pieces of DNA joined end-to-end
• has repeated viral genomes

Question 14

Packaging signals

Answer 14

• direct specificity for incorporation of viral genomes into virions
• nucleotide sequence of DNA or RNA viral genome that gets recognized by packaging proteins
• has highly specific interactions with capsid proteins
• interactions between negatively charged phosphate groups on nucleic acids with positive charges on the proteins

Question 15

Packaging of the genome of helical RNA viruses

Answer 15

• helical structure forms like a zipper around the remainder of the RNA molecule
• selective for viral RNA
• advantageous for the virus because it will select for viral RNA and not package the cell RNA because the cell RNA does not have the right nucleotide sequences
• For many negative strand RNA viruses, packaging happens as RNA is synthesized

Question 16

core proteins

Answer 16

• associated with the DNA viral genome inside the capsid
• nucleoprotein complex (nucleic acid + protein)
• neutralizes the negative charges on DNA
• involved in condensing the viral DNA for optimal packaging
• can use cellular and/or viral histones to form nucleosomes

Question 17

Virus envelopes and budding

Answer 17

• involves membrane curvature
  Different mechanisms
  1. nucleocapsids can assemble in the cytoplasm and approach membrane via binding to cytoplasmic tails of envelope proteins, once proteins are inserted into the plasma membrane it tends to bend it to form a curvature
  2. lots of helical nucleocapsids use internal matrix proteins
  3. some generate empty envelopes and budding is driven by envelope glycoproteins, they cluster and change membrane curvature that triggers cellular machinery
• many viruses require cell proteins for final pinch off

Question 18

matrix proteins

Answer 18

• encoded by viruses
• present just underneath the envelope
• forms connecting bridges between envelope and nucleocapsid, keeps everything in place

Question 19

retrovirus budding

Answer 19

• assembly of nucleocapsids directly at the membrane during budding
• a precursor protein (gap protein) iS cleaved to generate matrix and nucleocapsid proteins
• synthesis of a large polypeptide and it is then cleaved at specific sequences to get matrix and nucleocapsid proteins
• can also make ‘bald proteins’, do not have envelopes and layer of gag proteins on inner plasma membrane surface interact strongly with lipid bilayer that drives budding

Question 20

ESCRT proteins and budding

Answer 20

• help pinch off in budding
• assembles in concentric spirals, which constrict the neck of the vesicle and it eventually cuts it off
• can happen in the ER or the plasma membrane

Question 21

mechanisms for release of the viral genome (virion disassembly)

Answer 21

1. proteolytic cleavage of capsid proteins
   - most common
   - self cleavage, pH dependent, membrane fusion
2. unspooling of genome into the cell
   - particle stays outside and injects genetic material into the cell
   - common in bacteriophages
3. Interaction of genome with cytoplasmic components
   - DNA is folded in certain way in the capsid
   - may need to undergo structural changes
   - opens it up to start replicating or using the genome

Question 22

virus classification is based on…

Answer 22

• molecular architecture
• genetic relatedness
• host organism

Question 23

viruses are grouped into…

Answer 23

• species
• genera
• families

Question 24

Names of families end in…

Answer 24

-viridae

Question 25

Names of genera end in…

Answer 25

• virus

Question 26

molecular criteria used to classify viruses

Answer 26

1. type of nucleic acid genome (DNA or RNA)
2. strandedness of nucleic acids (single or double-stranded)
3. Topology of nucleic acids (linear, circular, fragmented)
4. Capsid symmetry (icosahedral, helical, none)
5. Presence or absence of an envelope

Question 27

genetic relatedness can be determined by…

Answer 27

• comparing genomic sequences
• comparing amino acid sequences
• looking at order of genes
• mechanisms of encoding mRNA
• use of reverse transcriptase

Question 28

Categories of virus hosts

Answer 28

1. bacteria
2. archaea
3. lower eukaryotes (fungi, protozoa, algae)
4. Plants
5. Invertebrates
6. Vertebrates (including humans)

Question 29

Virus species

Answer 29

• share a high degree of nucleic acid homology
• share similar amino acid sequences
• share antigenic properties
• infect limited organisms or specific target cells/tissues
• have common genetic lineage

Question 30

Virus genera

Answer 30

• share genome organization and size
• share tructure of the virion
• share replication strategies
• related by evolution but may have divergences in nucelotide and amino acid sequences
• they infect different organisms or cells/tissues within an organism

Question 31

Virus families

Answer 31

• share overall/general genome organization
• share virion structure
• share replication mechanisms
• can vary greatly in virion size and genome length
• may have unique genes not present in other family members
• could have evolved separately, with limited homology of nucleotide or amino acid sequences

Question 32

bacteriophages naming convention

Answer 32

• include name of the host bacterial genus and an arbitrary number
• normally just referred to as “phage” or “bacteriophage” followed by a number
  ex. Baccilus phage SP01

Question 33

plant virus naming convention

Answer 33

named after host plant species in which the virus was first isolated and disease symptoms caused by virus
ex. Tobacco mosaic virus

Question 34

insect virus naming convention

Answer 34

include Latin name of the insect species from which the virus was isolated followed by virus genus name
ex. Aedes aegypti denosvirus

Question 35

vertebrate virus naming conventions

Answer 35

• number of diff conventions
• host species of origin ex. simian virus 40
• location of isolation ex. Ebola virus
• disease caused ex. measles virus

Question 36

characteristics of viruses with double-stranded DNA genomes

Answer 36

• have vide variation in genome size
• have unfragmented genomes
• some are circular, some are linear
• the larger viruses tend to have envelopes
• no know dsDNA viruses infect plants

Question 37

characteristics of viruses with positive strand RNA genomes

Answer 37

• linear genomes
• most genomes are <19kb expect for coronaviruses
• if genome is <10kb = not enveloped
• found in most plant viruses and many vertebrate viruses
• very susceptible to degradation via nucleases or divalent cation attack which tends to limit genome size

Question 38

characteristics of viruses with negative stand RNA genomes

Answer 38

• have helical nucleocapsids
• some have fragmented genomes
• some of most widespread and deadly diseases are caused by negative strand RNA
• all -ve strand viruses that infect vertebrates are enveloped
• no known -ve strand viruses of prokaryotes
• linear genomes

Question 39

characteristics of viruses with double-stranded RNA genomes

Answer 39

• fragmented genomes
• capsids with icosahedral symmetry
• linear genomes
• multiple genome segments that each code a single mRNA and usually a single viral protein
• tend to have broad host range because they have their own RNA replication machinery so they are less dependent on specific cell environment
• unique mechanism of transcription where capsids act as tiny intracellular machines

Question 40

transcription of viruses with double-stranded RNA genomes

Answer 40

• capsid/subviral particle remains intact intracellularly while mRNAs are transcribed from each individual segment
• virus uses RNA polymerase that is packaged in the virus particle along with the genome to make mRNA from the genomic segments
• capsid provides a structure to position RNA polymerase with each segment and allows for directional transfer of mRNAs to cytoplasm
• reads RNA template 3’ to 5’ and synthesizes 5’ to 3’