Human immunodeficiency virus type 1 (HIV-1) is responsible for the acquired immunodeficiency syndrome (AIDS), a leading cause of death worldwide infecting millions of people each year. Despite intensive research in vaccine development, therapies against HIV-1 infection are not curative and the huge genetic variability of HIV-1 challenges drug development. Current animal models for HIV-1 research present important limitations, impairing the progress of in vivo approaches. Macaques require CD8+ depletion or large portions of the genome to be replaced by sequences derived from simian immunodeficiency viruses to progress to AIDS, and the maintenance cost is high. Mice are a cheaper alternative, but need to be 'humanized' and breeding is not possible and knockout experiments are difficult. The development of an HIV-1 clone able to replicate in mice is a challenging proposal. The lack of human co-factors in mice impedes function of the HIV-1 accessory proteins Tat and Rev, hampering HIV-1 replication. The Tat and Rev function can be replaced by constitutive/chimeric promoters, codon-optimized genes and the constitutive transport element (CTE), generating a novel HIV-1 clone able to replicate in mice without disrupting the amino acid sequence of the virus. By minimally manipulating the genomic 'identity' of the virus, we propose the generation of an HIV-1 clone able to replicate in mice to assist in antiviral drug development. My results have determined that murine NIH 3T3 cells are able to generate pseudotyped HIV-1 particles, but they are not infectious. Codon-optimized HIV-1 constructs are efficiently made in human HEK-293T cells in a Tat and Rev independent manner and capable of packaging a competent genome in trans. CSGW (an HIV-1 vector genome) efficiently generates infectious particles in the absence of Tat and Rev in human cells when 4 copies of the CTE are placed preceding the 3’LTR. HIV-1 replication competent genomes lacking tat expression and encoding different promoters are functional during the first cycle of replication when Tat is added in trans. Finally, we developed a replication competent HIV-1 clone lacking tat and rev genes and encoding 4CTEs that could be a future candidate for HIV research. My results shown that the development of an HIV-1 Tat-Rev independent clone could become a candidate for HIV research in a near future, but further investigations are necessary before proposing our model as an alternative yet.
| Identifer | oai:union.ndltd.org:bl.uk/oai:ethos.bl.uk:754382 |
| Date | January 2018 |
| Creators | Vera Ortega, Walter |
| Publisher | University of Glasgow |
| Source Sets | Ethos UK |
| Detected Language | English |
| Type | Electronic Thesis or Dissertation |
| Source | http://theses.gla.ac.uk/30755/ |
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