Viral Pathogens: Classification, Biology, Diseases I

Question 1

How are viruses classified

Answer 1

They are classified based on their viral genome

Question 2

What viral genome types exist

Answer 2

• Single-stranded RNA (ssRNA)
• Double-strandedRNA (dsRNA)
• Single-stranded DNA (ssDNA)
• Double-stranded RNA (dsDNA)
• Double-stranded genomes have complementary base pairing
• RNA genomes can be linear and segmented i.e. more than one RNA per capsid
• DNA genomes can be linear or circular.

Question 3

What is the central dogma

Answer 3

• DNA polymerase replicates DNA into DNA
• RNA polymerase converts DNA into RNA
• Ribosomes convert RNA into proteins

Question 4

How do viruses use the central dogma

Answer 4

• Viruses use reverse transcriptase to covert RNA into DNA
• RNA replication to form sense or antisense strands of RNA, using RNA dependant RNA polymeases

Question 5

What is the Baltimore classification

Answer 5

Classification of the viral genome in how they replicate
- Has 7 different groups based on the viral genome

Question 6

Describe the structure of HIV

Answer 6

• Nucleic acid covered by Proteins
• Has a capsid and membrane proteins all around it.

Question 7

Describe the structure of the mature HIV-1 particle

Answer 7

• The outer envelope of HIV consists of a lipid bilayer with protruding Env spikes (heterotrimers of SU 3TM3).
• Inside the envelope lie shells of Gag proteins. In the immature particle, Gag itself forms a single shell.
• MA associates with the membrane CA forms the conical capsid NC coats the viral RNA genome.
• The core contains two genomic RNA strands (plus strand), tRNALys3, and ~50 copies of each viral enzyme (PR, RT, and IN)

Question 8

Describe the genome organisation of Retroviruses

Answer 8

Retroviruses synthesise 3 polyproteins:

• Gag; group specific antigen; viral core proteins; MA (matrix), CA (capsid), NC (nucleocapsid)
• Pol; viral enzymes; protease (PR), reverse transcriptase (RT) and integrase (IN)
• Env; envelope glycoprotein; gp120 SU (surface); gp41 TM (transmembrane)

RNA is converted into DNA in the host cell

Question 9

List the Regulatory/Accessory proteins in HIV-1

Answer 9

• Tat - potent activator of viral transcription
• Rev - mediates unspliced RNA nuclear export
• Vif - critical regulator of virus infectivity
• Nef - immune modulator, T-cell activation, virus spread
• Vpu - immune modulator, virus release
• Vpr - cell cycle, virus nuclear import

Question 10

How does HIV fuse with the host cell

Answer 10

• Spike proteins (GP120) binds to CD4 proteins on host cell membrane
• GP41 subunits sites on the viral membrane bind CoR on the host cell
• This helps orient the virus towards the host
• The protein folds in on another and brings together both membranes and hips it fuse together

Question 11

How does HIV enter into the host cell

Answer 11

• After fusion, the viral content must make its way into the nucleus
• The viral contents use the mitochondrial trafficking process to help it move closer to the nucleus - Active process which is fast and orients the contents for the nucleus
• The contents enter the nucleus via the Nuclear pre receptors

Question 12

How does HIV enter via the NPC

Answer 12

Capsid proteins interact with proteins on the nuclear pore that help send the capsid to different parts of the nucleus

This allows it to be sent to regions that are more transcriptionally active

Question 13

Describe the function of Reverse Transcriptase

Answer 13

RT is a heterodimer of p66 and p51 subunits.
Catalytic properties are in p66 subunit, p51 serves structural role and lacks RNAse H domain

RT displays three distinct enzymatic activities:

• RNA-dependent DNA polymerase
• RNAse H (cleaves RNA from RNA/DNA hybrid)
• DNA-dependent DNA polymerase

Question 14

Describe the basic steps of reverse transcriptase

Answer 14

• Makes an RNA primer strand using the existing RNA
• Makes a DNA strand from the RNA primer
• Use the made DNA to make a double strand

Question 15

How does the HIV DAN genome integrate into host chromosomes

Answer 15

• Finds a target region within the chromosome
• Uses the integrase enzyme to put the DNA into the host cell genome

Question 16

How does the viral integrase enzyme bind both host and viral DNA

Answer 16

• The enzyme mimics normal cell DNA checking
• Finds broken parts of the DNA
• Integrase anneals viral DNA into cellular DNA

Question 17

How does integrase also help with targeting chromatin

Answer 17

Cellular proteins are bound to integrase such as LEDGF/P75 and are directed to transcriptionally active parts of the DNA

Question 18

How is the viral DNA transcribed in the nucleus

Answer 18

Cellular proteins bind viral transcription factors that produce viral RNA

Question 19

What is the purpose of the TAT protein

Answer 19

The TAT protein ensures that viral DNA gets transcribed and not cellular DNA
- Binding of TAT protein to TAR RNA enhances elongation of RNA pol II

Question 20

How does HIV-1 provirus generate different mRNAs for the viral proteins

Answer 20

Alternative splicing leads to the production of different viral proteins

Question 21

How does the viral proteins mediate alternative splicing

Answer 21

REV protein produced by viral DNA mediates the splicing of mRNA to produce different proteins

Question 22

What is RRE and REV used in

Answer 22

Essential for nuclear export of intron-containing viral mRNAs

• RRE has lariat structures that act as recognition and binding sites of proteins like REV

Question 23

How are the produced proteins and mRNA transported to make a new HIV virus

Answer 23

• HIV membrane is made and processed in the ER and Golgi and transported into the membrane
• Proteins and mRNA are transported using microtubule proteins
• The virus is assembled at the plasma membrane where all the components are brought together

Question 24

What is the function of Gag and Gag-Pol proteins in HIV production

Answer 24

Help assemble viral particle at the membrane

Gag-pol protein is generated by -1 ribosomal frameshifting induced by a ‘slippery’ sequence and an RNA hairpin structure

Question 25

What is the function of ESCRT machinery in a normal cell

Answer 25

Used in cytokinesis to form daughter cells.

Causes the splitting of membrane

Question 27

What function does the ESCRT perform when it is hijacked HIV

Answer 27

• Sorts out cellular cargo to package out of the cell
• Used to form viral particles in the membrane to Budd out of the celll

Question 28

How are polyproteins organised before HIV budding

Answer 28

• Polyproteins are cut by proteases to make individual proteins for the viral capsid
• ESCR organises the proteins
• Proteases also form RT and other enzymes that the virus needs

Question 29

Describe the maturation process of HIV after budding from the host

Answer 29

• Immature HIV has proteins in the wrong orientation which makes it not very infective
• Maturation of HIV makes it not infect previous host cell
• Viral protease enzymes modifies proteins in the cased and matures it
• Ready for infection