Positive selection pressure introduces secondary mutations at Gag cleavage sites in human immunodeficiency virus type 1 harboring major protease resistance mutations

Abstract

Human immunodeficiency virus type 1 (HIV-1) protease inhibitors (PIs) specifically target the HIV-1 protease enzyme. Mutations in the enzyme can result in PI resistance (termed PI mutations); however, mutations in the HIV-1 gag region, the substrate for the protease enzyme, might also lead to PI resistance. We analyzed gag and pol sequence data from the following 313 HIV-1-infected patients: 160 treatment-naïve patients, 93 patients failing antiretroviral treatment that included a PI (with no major PI mutations), and 60 patients failing antiretroviral treatment that included a PI (with major PI mutations). Additional sequences from 13 patients were included for longitudinal analysis. We assessed positive selection pressure on the gag/protease region using a test for the overall influence of positive selection and a total of five tests to identify positively selected single codons. We found that positive selection pressure was the driving evolutionary force for the gag region in all three patient groups. An increase in positive selection was observed in gag cleavage site regions p7/p1/p6 only after the acquisition of major PI mutations, suggesting that amino acids in gag cleavage sites under positive selection pressure could function as compensatory mutations for major PI mutations in the protease region. Isolated gag mutations did not appear to confer PI resistance, but mutations in the gag cleavage sites could substitute for minor PI resistance mutations in the protease region.

FIG. 1.

The gag selection model and partial amino acid alignment for gag residues 428 to 453 including both gag CS p1/p7 and p7/p6. In the selection model, all amino acid sites under positive selection pressure according to the MEC model and results from clade analyses shown in Table 4 are shown. The partial amino acid alignment shows (red boxes) unique sites that are only under positive selection in treated patients with major PI mutations. Sites in yellow or red boxes are under positive selection according to model 8.

FIG. 2.

Phylogenetic tree for gag residues showing synonymous (blue), nonsynonymous (red), or both (orange) mutations. Mutations were inferred using GARLI (29). (A) gag site 452. (B) gag site 453. (C) gag site 449. (D) gag site 451. pos, position.

FIG. 3.

Longitudinal data for patient 01. Appearance of mutations in the gag gene and the protease gene are shown, as well as viral load and PI treatment history.