Compensatory Point Mutations in the Human Immunodeficiency Virus Type 1 Gag Region That Are Distal from Deletion Mutations in the Dimerization Initiation Site Can Restore Viral Replication

Abstract

The dimerization initiation site (DIS), downstream of the long terminal repeat within the human immunodeficiency virus type 1 (HIV-1) genome, can form a stem-loop structure (SL1) that has been shown to be involved in the packaging of viral RNA. In order to further determine the role of this region in the virus life cycle, we deleted the 16 nucleotides (nt) at positions +238 to +253 within SL1 to generate a construct termed BH10-LD3 and showed that this virus was impaired in viral RNA packaging, viral gene expression, and viral replication. Long-term culture of these mutated viruses in MT-2 cells, i.e., 18 passages, yielded revertant viruses that possessed infectivities similar to that of the wild type. Cloning and sequencing showed that these viruses retained the original 16-nt deletion but possessed two additional point mutations, which were located within the p2 and NC regions of the Gag coding region, respectively, and which were therefore named MP2 and MNC. Site-directed mutagenesis studies revealed that both of these point mutations were necessary to compensate for the 16-nt deletion in BH10-LD3. A construct with both the 16-nt deletion and the MP2 mutation, i.e., LD3-MP2, produced approximately five times more viral protein than BH10-LD3, while the MNC mutation, i.e., construct LD3-MNC, reversed the defects in viral RNA packaging. We also deleted nt +261 to +274 within the 3′ end of SL1 and showed that the diminished infectivity of the mutated virus, termed BH10-LD4, could also be restored by the MP2 and MNC point mutations. Therefore, compensatory mutations within the p2 and NC proteins, distal from deletions within the DIS region of the HIV genome, can restore HIV replication, viral gene expression, and viral RNA packaging to control levels.

Compensatory adaptation to the deleterious effect of antibiotic resistance in Salmonella typhimurium

“…The presented results suggest that, even for a highly conserved and essential target such as the ribosome, there is a plethora of ways to restore fitness by compensatory mutations. Similar conclusions can be drawn from other experimental systems in which compensatory mutations can conceal the effects of deleterious mutations (Olsthoorn and van Duin, 1996;Liang et al, 1998;Burch and Chao, 1999;Bull et al, 2000;Moore et al, 2000;Rokyta et al, 2002). Thus, it is possible that most deleterious mutations, irrespective of their location and target, can be partly or fully compensated by additional mutations (however, for an interesting exception, see Crill et al, 2000).…”

HIV-1 Pr55Gagbinds genomic and spliced RNAs with different affinity and stoichiometry

“…86 Interestingly, this genomic site generated a consensus over the last years, since Pr55 Gag binding sites in immature virions and NCp7 binding sites in mature virions, 17,69 fit with present findings and our previous in vitro data (Table S1). 85,86 Other studies showed that deletion of the lower part of SL1 101,102 or mutation of its internal loop 32 reduces gRNA packaging and viral infectivity. The effect of mutations in the SL1 internal loop on in vivo gRNA dimerization 32,103-105 is probably a direct consequence of the weaker binding of Pr55 Gag to the mutated SL1, the apical loop of which mediates gRNA dimerization.…”

Evolution of Cell Recognition by Viruses: A Source of Biological Novelty with Medical Implications

“…Small numbers of mutations in components of mutant spectra are easily attainable by diversifying populations of viruses during acute or chronic infections. Even when a mutation that confers a phenotypic change results in a modest fitness decrease, compensatory mutations can have an opportunity to rescue genomes with normal or nearly normal fitness values (Cassady et al, 2002;Escarmís et al, 1999Escarmís et al, , 2002Lá zaro et al, 2002;Liang et al, 1998;Nijhuis et al, 1999;Wang et al, 1996;Yuan and Shih, 2000). a Based on many published studies reviewed in Domingo et al (1985Domingo et al ( , 1999Domingo et al ( , 2001, Crandall (1999), Flint et al (2000), and DeFilippis and Villarreal (2001).…”