Dougherty - Viral Replication Flashcards
What are the three classifications of retroviruses (low yield)
OnCoretroviruses - cause tumors
Lentiviruses - cause immunological and neurological diseases with long latency
Spumaviruses - causes vacuolation of some mammalian cells in culture
What are some of the major proteins involved with the HIV virus?
TM - transmembrane envelope protein SU - surface envelope protein Integrase Protease Reverse transcriptase
What type of nucleic acid does HIV have?
Single stranded RNA. It will then go to double stranded DNA back to ssRNA
What is the shape of the capsid of HIV?
Conal
How many strands of RNA are in HIV?
2
- both positive strands
How do retroviruses (HIV) get into the cell initially?
Receptor mediated endocytosis via the SU (surface envelope protein) binding to the cells that it is targeting.
Retrovirus life cycle
1) virus gets into the cells through receptor mediated endocytosis via the SU glycoprotein on the virus.
2) the virus + strand RNA will be reverse transcribed to make a dsDNA
3) this dsDNA will become part of the “pre-integration” complex and will be transported into the nucleus
4) the dsDNA will then be incorporated into the genome.
5) Once in the genome, it is known as a “provirus” and is transcribed by cellular RNA Pol II
6) This then yields a full length RNA which can either be packaged into a new virion to infect other cells or it can be translated and spliced to make viral proteins.
Retroviral genome
The provirus DNA genome is made up of a long terminal repeat (LTR) on each end with all of the genes in between. The LTR is made up of U3-R-U5 on both ends. The viral genes to make proteins are in between the LTRs and at at the minimum always have the genes gag-pol-env in that order with sometimes other genes in between. Once RNA Pol II transcribes the RNA genome from the provirus, the genome loses U3 on the 5’ end and U5 on the 3’ end.
Where is the transcription start site in the provirus genome?
In between U3 and R on the 5’ end, so when the DNA is transcribed, they lose the U3.
Where does transcription end in the provirus when making RNA?
Between R and U5 there is Polyadenylation, so U5 doesn’t get incorporated into the DNA.
What/where are the retrovirus receptors?
It depends on what the retrovirus is. Most retroviruses act on the cell surface receptors. For HIV, the cell surface receptor is CD4.
What is the co-receptor for HIV?
CCR5 and CXCR4 - members of the chemokine receptor family.
What is the basic mechanism of reverse transcriptase in the first place to make the DNA that was incorporated into the “provirus”
tRNA is used to prime synthesis. It starts at the 5’ end to “reverse transcribe” the 5’ LTR region because if you remember that part is key for viral transcription from the DNA. Then, the tRNA essentially takes that piece and brings it over to the 3’ side and continues on from there so there is a full complement of U3-R-U5. There is then degradation of the RNA as you go along by RNAase.
Recombination during reverse transcription
In the event that a cell is infected with two different RNA viruses, there will be a very high rate of recombination between the two.
Also, retro viral reverse transcriptase is much more error prone relative to cellular DNA synthesis.
How is the dsDNA integrated into the genome?
Now we have the dsDNA, the virus has been encoded, now it just has to be integrated. The sequences linked to each end of the genome are essential as well as the reverse transcriptase. They promote efficient integration into the genome of the infected cell.
The viruses integrate relatively randomly. They can integrate pretty much much anywhere but they have preferences. So for instance, HIV prefers actively transcribed genes. Others might want to be in other places. When a retrovirus integrates, what it’ll do is that the cellular DNA sequence that it is targeting will be duplicated. The amount of nucleotides that are duplicated are usually between 4 and 6 (HIV is 5). Then, the DNA will be inserted in between the two duplicates and in the process, two nucleotides will be lost from the ends of the RNA.
