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Title: Investigating the role of stem-loop 1 in the assembly process of HIV-1
Author: Hellmund, Christopher James
ISNI:       0000 0004 7660 9349
Awarding Body: University of Cambridge
Current Institution: University of Cambridge
Date of Award: 2019
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An important step in the production of infectious HIV-1 particles is maturation of the virus core. This process is completed by cleavage of the capsid (CA) domain of Gag, from its precursor, CA-SP1, by the viral protease. Large deletions in stem-loop 1 (SL1) in the 5' untranslated region (UTR) of HIV-1 genomic RNA (gRNA) delay CA-SP1 processing. SL1 harbours the dimerisation initiation site (DIS) palindrome suggesting that efficient Gag processing may be linked to gRNA dimerization as shown in HIV-2. However, a dimerisation mutant with normal Gag processing was identified. Gag processing defects are hallmarks of late domain mutants, and SL1 mutation was found to result in reduced virus release. HIV-1 hijacks the host's endosomal complexes required for transport (ESCRT) pathway to enable budding. An ESCRT-associated protein, ALIX, is known to be capable of binding to the nucleocapsid (NC) domain of Gag using lipids or RNA as a 'bridge' in vitro. It was hypothesised that SL1 mutation disrupts an RNA-dependent interaction that occurs during virus assembly. Consistent with this, an intact SL1 was found to be required for efficient ALIX function. Increasing the abundance of gRNA in the cell by expressing it in trans accelerated CA-SP1 processing in a manner that required ALIX's binding motif in p6. Gag processing could also be accelerated by introducing previously identified compensatory mutations into the SP1 and NC domains of Gag, in a manner reminiscent of the actions of maturation inhibitor resistance mutations. The effects of the compensatory mutations were also dependent on intact late domain motifs. These data suggest that gRNA is involved in regulating virus budding and maturation through interaction with ALIX. A model is proposed whereby the packaging signal (psi) region of gRNA acts as a bridge between Gag and ALIX, acting as a checkpoint mechanism to promote Gag processing and optimise release of virions that have successfully packaged gRNA.
Supervisor: Lever, Andrew Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
Keywords: HIV-1 ; RNA ; Gag ; Virus