Retroviruses (HIV) Flashcards
All retroviruses have:
1) 2 copies of (+) RNA genome
2) 5’ CAP
3) Poly A tail
4) Envelope
5) Package reverse transcriptase
(T/F) Retroviruses make a dsDNA intermediate which incorporates into our genome (hides from the immune system). We are not aware if we are infected at this stage. It also requires warm temps to be a host.
True!
What is MLV?
MLV stands for murine leukemia virus. It is the 2nd most studied retrovirus.
All mouse strains carry genetic information for MuLV-related viruses (endogenous) in their chromosomes.
It is part of Class 1 of retrovirus classification.
(T/F) MMTV (murine mammary tumor virus) belongs to class III of retrovirus classification, while HIV (human immunodeficiency virus) belongs to class II.
False!
MMTV (murine mammary tumor virus) belongs to class II of retrovirus classification, while HIV (human immunodeficiency virus) belongs to class III.
Why did we think that MLV were causing cancer?
1) Uncontrolled division of the progenitor cells
2) Removal of a “break” of the cell cycle
Match the following terms to their definitions:
1) Retrovirus
2) Retroviral reverse transcription
3) Retroviral transcription
A) To convert a ssRNA genome into a dsDNA provirus. Done in the CYTOSOL.
B) Process by which the cell’s RNA polymerase II transcribes integrated proviral DNA into mRNA in the NUCLEUS.
C) A virus that converts its single-stranded (+) RNA genome into dsDNA through the action of virally encoded REVERSE TRANSCRIPTASE prior to INTEGRATION into the host cell’s genomic DNA.
Retrovirus: A virus that converts its single-stranded (+) RNA genome into dsDNA through the action of virally encoded REVERSE TRANSCRIPTASE prior to INTEGRATION into the host cell’s genomic DNA.
2) Retroviral reverse transcription: To convert a ssRNA genome into a dsDNA provirus. Done in the CYTOSOL.
3) Retroviral transcription: Process by which the cell’s RNA polymerase II transcribes integrated proviral DNA into mRNA in the NUCLEUS.
Briefly answer the following questions regarding HIV:
1) What is the family, the subfamily and the genus?
2) How many species are there? Differentiate both.
3) What receptors do they interact with for entry (cell tropism)?
1) Family - retroviridase, subfamily - orthoretrovirinase, genus - lentivirus.
2) There are two: HIV1 & HIV2. Both are recombinant viruses of SIVs transmitted by ZOONOSIS to humans. HIV1 (chimpanzee SIV) & HIV2 (Sooty mangabey)
3) They require CO-RECEPTORS for entry. CCR5 (macrophages) and CXCR4 (CD4 + T cells).
Which one of the statements regarding HIV is false?
1) HIV brings its own proteins to produce a successful infection.
2) HIV has a lot of surface proteins and is not enveloped. Due to these factors, it is noticed by the host immune system.
3) Immature virus is 150x less effective in infection.
2!
HIV has very little surface proteins and it has host membrane in its membrane. Due to these factors, it is not noticed by the host immune system.
Match the following proteins (3) that are produced through pro-protein GAG to their definitions:
1) Nucleocapsid (NC)
2) Capsid (CA)
3) Matrix (MA)
A) p17 - forms the matrix or inner layer to the envelope
B) p7 - binds the 2 RNA copies
C) p24 - forms the conical capsid
Nucleocapsid (NC): p7 - binds the 2 RNA copies
Capsid (CA): p24 - forms the conical capsid
Matrix (MA): p17 - forms the matrix or inner layer to the envelope
5’-MA-CA-NC-…3’
HIV has a _______, ______, _______ ssRNA (+) genome.
________ and ______ are present.
monopartite, linear, dimeric
5’ cap; polyA tail
What is LTR? What does it contain?
LTR: long terminal repeat.
It contains the U3, R and U5 regions of HIV.
It is at the 5’ and the 3’ of the RNA.
What are the 3 main genes of the HIV genome?
1) Group Specific Antigen GAG (structural pro-proteins)
2) Pol (enzymes)
3) Env (receptor ligand)
Gag-pol polypeptide is made by ___ _________ ______ at the P6 region of ____ (part of GAG that overlaps pol)
Env is in the same reading frame as _________. It is generated by _______ ________.
-1 Ribosomal Frameshift; NC
Gag-Pol; Alternative splicing
What are the 4 enzymes that POL gene encodes for?
1) Protease (pro)
2) Reverse transcriptase (RT)
3) RNase H (cleaves DNA/RNA intermediates)
4) Integrase (IN)
VPU and ENV are generated from ________ _______.
The other accessory genes are generated by __________ ______.
_____ is the last protein by the HIV genome.
Leaky scanning
Alternative splicing
Nef
Match HIV-1 proteins to their definitions:
1) LTR
2) gag
3) pol
4) vif
5) vpr
6) rev
7) vpu
8) env
9) tat
10) nef
A) encodes several enzymes
B) promotes cell-cycle arrest and facilitates infection of macrophages
C) cleaved into two proteins: gp120 mediates viral binding to the cell, and gp41 promotes fusion of the viral and cellular membranes
D) promotes CD4 lymphocyte activation, blocks cell suicide, enhances infectivity, and is associated with progression of disease
E) contains transcriptional regions and is required for the transcription of the viral genome
F) promotes CD4 degradation and helps virion release
G) enhances viral RNA transcription and promotes CD4 lymphocyte activation
H) regulates viral RNA nuclear export
I) building block of the virus particle core
J) overcomes inhibitions by host factors and promotes more stable reverse transcription complexes
1) LTR: contains transcriptional regions and is required for the transcription of the viral genome
2) gag: building block of the virus particle core
3) pol: encodes several enzymes
4) vif: overcomes inhibitions by host factors and promotes more stable reverse transcription complexes
5) vpr: promotes cell-cycle arrest and facilitates infection of macrophages
6) rev: regulates viral RNA nuclear export
7) vpu: promotes CD4 degradation and helps virion release
8) env: cleaved into two proteins: gp120 mediates viral binding to the cell, and gp41 promotes fusion of the viral and cellular membranes
9) tat: enhances viral RNA transcription and promotes CD4 lymphocyte activation
10) nef: promotes CD4 lymphocyte activation, blocks cell suicide, enhances infectivity, and is associated with progression of disease
Give an overview of HIV-1 entry (step 1 of its life cycle).
HIV-1 enters target cells (macrophages and CD4+ T cells) by receptor (CD4 with coreceptors) mediated FUSION and needs to cross and/or interact with the ACTIN CORTEX underlying the plasma membrane for productive infection to take place.
POST-FUSION UNCOATING DOES NOT TAKE PLACED AT THIS STAGE.
HIV entry can be divided into THREE key steps, all mediated by the viral surface protein ENV, which is a trimer of GP120 and GP41 heterodimers.
What are the 3 steps?
1) The virion binds to the host cell which can facilitate by non-specific cell attachment factors and allows Env (gp120) to bind CD4.
2) CD binding induces CONFORMATION CHANGES in Env that enables engagement of a co-receptor (ex. CCR5 or CXCR4). This step TRIGGERS MEMBRANE FUSION.
3) GP41 subunits of Env enable fusion of the viral and host membranes allowing delivery of the viral particles.
What are the three main entry pathways that exist for HIV?
1) Endocytosis: HIV particles and receptors are internalized, and undergo lipid mixing with the vesicle membrane and deliver their contents into the cytoplasm.
2) Endocytosis but fusion occurs at the plasma membrane (DYNAMIN DEPENDENT FUSION WITH ENDOSOMES) and proceed at least to the stage of hemi-fusion (lipid mixing occurs). *receptor not internalized
3) Fusion events at the plasma membrane that do not result in any subsequent mixing; no productive infection.
1&2 are the main entry pathways for HIV (clathrin- and dynamin- dependent).
Three outcomes possible during HIV uncoating; a) uncoating and nuclear transport, b) no uncoating, c) premature uncoating.
Describe each and what they lead to.
A) Uncoating and nuclear transport: After entry into the cytoplasm, HIV-1 capsids that are on a path of PRODUCTIVE INFECTION remain intact and are transported towards the nucleus along the cytoskeleton. They uncoat at the nuclear membrane upon completion of reverse transcription.
B) Compromised uncoating, in the case of incomplete reverse transcription or of hyperstable capsid mutant leads to DEAD-END INFECTION.
C) Premature uncoating, in the case of TRIM5alpha restriction/unstable capsid mutants, leads to ABORTIVE INFECTION.
Describe the steps that follow HIV entering the host cell until it reaches the nuclear pore complex (i.e, steps 2, 3, 4, and 5 of HIV life cycle).
After entering host cell through receptor mediated fusion,
Step 2: The HIV reverse transcription complex (RTC) undergo RAPID and saltatory MICROTUBULE-DIRECTED movement towards the nuclear compartment
Step 3: RTCs undergo transition from microtubules onto ACTIN FILAMENTS at close proximity to the nuclear compartment
Step 4: Slow actin-directed movement towards the nuclear membrane occurs
Step 5: Docking on the nuclear pore complex
Reverse transcription of HIV is primer __________.
Dependent.
The primer is HOST tRNA that typically carries lysine residues! The primer provides 3’OH so the RT can use it to make a dsDNA intermediate.
*when virus buds out, it actually steals our tRNA
What is PBS found in the HIV genome?
Primer binding site! The tRNA (primer) binds here.
It is located on the 5’ end of the mRNA.
It is critical for initiation of synthesis of dsDNA intermediate. The RT takes the 3’ OH of the tRNA and starts synthesizing a (-) DNA.
It is also copied twice during reverse transcription; once on the (-) DNA from the RNA genome and once on the (+) DNA from the tRNA primer. These sites allow for the annealing of the two dsDNA.
(T/F) The 5’ LTR of HIV gRNA (R-U5) does not have a U3.
True!
5’ : r U5 pbs
3’ r U3 ppt
