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The variable loop 3 in the envelope glycoprotein is critical for the atypical coreceptor usage of an HIV-1 strain.

Xiang Y, Liu W, Chen Y, Zhang C, Su W, Zhang Y, Sun J, Gao F, Jiang C - PLoS ONE (2014)

Bottom Line: The G306A and S322A mutations significantly reduced the replication capacity of YU2.6248V3 in GPR15+ cells, while all other alanine substitutions at positions 307, 314, 315, 316, 317 and 318 completely abrogated the infectivity of YU2.6248V3 in GPR15+ cells.The E314A mutation, as the E314G mutation reported before, also rendered the YU2.6248V3 infectious in CCR5+ cells, while none of other alanine mutants could infect CCR5+ cells.These results demonstrated that amino acids in ZP6248 V3 might form a unique conformation that was critical for the interaction with GPR15 while the amino acids at position 314 in the V3 crown of ZP6248 played a key role in interaction with both CCR5 and GPR15.

View Article: PubMed Central - PubMed

Affiliation: National Engineering Laboratory For AIDS Vaccine, College of Life Science, Jilin University, Changchun, Jilin, China.

ABSTRACT
The majority of HIV-1 strains enter CD4+ T cells using the CCR5 and/or CXCR4 co-receptor. However, we recently identified a transmitted/founder (T/F) virus (ZP6248) that efficiently used an alternative coreceptor GPR15, rather than commonly used CXCR4 and CCR5, to establish clinical infection. To understand which regions in the env gene were critical for the atypical coreceptor usage, we generated a set of V3 mutants and determined their infectivity in GHOST cells that expressed different coreceptors. When the variable loop 3 (V3) in YU2 was replaced with the ZP6248 V3 (YU2.6248V3), the chimera YU2.6248V3 infected GPR15+ cells but not CCR5+ cells. To determine which amino acids in V3 was responsible for this phenotype change, each of the eight amino acids that differed from the subtype B consensus V3 was substituted with alanine. The G306A and S322A mutations significantly reduced the replication capacity of YU2.6248V3 in GPR15+ cells, while all other alanine substitutions at positions 307, 314, 315, 316, 317 and 318 completely abrogated the infectivity of YU2.6248V3 in GPR15+ cells. The E314A mutation, as the E314G mutation reported before, also rendered the YU2.6248V3 infectious in CCR5+ cells, while none of other alanine mutants could infect CCR5+ cells. These results demonstrated that amino acids in ZP6248 V3 might form a unique conformation that was critical for the interaction with GPR15 while the amino acids at position 314 in the V3 crown of ZP6248 played a key role in interaction with both CCR5 and GPR15. The unique phenotypes of ZP6248 can serve as a model to understand how HIV-1 explores the diverse coreceptor reservoir through novel genetic variants to establish clinical infection.

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Viability of cells infected with ZP6248 and its mutants.The GHOST GPR15 cells were infected with ZP6248 and its mutants and cultured for 10(day 10) was determined by CellTiter-Glo Luminescent Cell Viability Assay. Each virus was assayed in triplicate and mean ± standard deviation is shown. Similar results were obtained in two independent experiments and the results from one experiment are shown.
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pone-0098058-g004: Viability of cells infected with ZP6248 and its mutants.The GHOST GPR15 cells were infected with ZP6248 and its mutants and cultured for 10(day 10) was determined by CellTiter-Glo Luminescent Cell Viability Assay. Each virus was assayed in triplicate and mean ± standard deviation is shown. Similar results were obtained in two independent experiments and the results from one experiment are shown.

Mentions: Examination of the cell viability showed that wild type ZP6248 was cytopathic in GPR15+ cells. The cell death was first observed at day 4 in the culture (Table 1) and about 50% of the cells died at day 10 (Fig. 4). The ZP6248 V3 rendered YU2 more cytopathic than the wild type ZP6248 (p = 0.001); only 26% of cells were still viable in the YU2.ZP6248V3 culture at day 10 (Fig. 4). Both G306A and S322A mutants caused significantly more cell death than wild type ZP6248 (p<0.001) as well as YU2.6248V3 (p = 0.003). The cell death was observed at day 2 in the culture (Table 1) and about 90% of the cells died at day 10 (Fig. 4). These results suggested that the low levels of p24 detected in the culture of YU2.6248V3 as well as both G306A and S322A mutants in GPR15+ cells was partly due to the higher numbers of dead cells.


The variable loop 3 in the envelope glycoprotein is critical for the atypical coreceptor usage of an HIV-1 strain.

Xiang Y, Liu W, Chen Y, Zhang C, Su W, Zhang Y, Sun J, Gao F, Jiang C - PLoS ONE (2014)

Viability of cells infected with ZP6248 and its mutants.The GHOST GPR15 cells were infected with ZP6248 and its mutants and cultured for 10(day 10) was determined by CellTiter-Glo Luminescent Cell Viability Assay. Each virus was assayed in triplicate and mean ± standard deviation is shown. Similar results were obtained in two independent experiments and the results from one experiment are shown.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4045670&req=5

pone-0098058-g004: Viability of cells infected with ZP6248 and its mutants.The GHOST GPR15 cells were infected with ZP6248 and its mutants and cultured for 10(day 10) was determined by CellTiter-Glo Luminescent Cell Viability Assay. Each virus was assayed in triplicate and mean ± standard deviation is shown. Similar results were obtained in two independent experiments and the results from one experiment are shown.
Mentions: Examination of the cell viability showed that wild type ZP6248 was cytopathic in GPR15+ cells. The cell death was first observed at day 4 in the culture (Table 1) and about 50% of the cells died at day 10 (Fig. 4). The ZP6248 V3 rendered YU2 more cytopathic than the wild type ZP6248 (p = 0.001); only 26% of cells were still viable in the YU2.ZP6248V3 culture at day 10 (Fig. 4). Both G306A and S322A mutants caused significantly more cell death than wild type ZP6248 (p<0.001) as well as YU2.6248V3 (p = 0.003). The cell death was observed at day 2 in the culture (Table 1) and about 90% of the cells died at day 10 (Fig. 4). These results suggested that the low levels of p24 detected in the culture of YU2.6248V3 as well as both G306A and S322A mutants in GPR15+ cells was partly due to the higher numbers of dead cells.

Bottom Line: The G306A and S322A mutations significantly reduced the replication capacity of YU2.6248V3 in GPR15+ cells, while all other alanine substitutions at positions 307, 314, 315, 316, 317 and 318 completely abrogated the infectivity of YU2.6248V3 in GPR15+ cells.The E314A mutation, as the E314G mutation reported before, also rendered the YU2.6248V3 infectious in CCR5+ cells, while none of other alanine mutants could infect CCR5+ cells.These results demonstrated that amino acids in ZP6248 V3 might form a unique conformation that was critical for the interaction with GPR15 while the amino acids at position 314 in the V3 crown of ZP6248 played a key role in interaction with both CCR5 and GPR15.

View Article: PubMed Central - PubMed

Affiliation: National Engineering Laboratory For AIDS Vaccine, College of Life Science, Jilin University, Changchun, Jilin, China.

ABSTRACT
The majority of HIV-1 strains enter CD4+ T cells using the CCR5 and/or CXCR4 co-receptor. However, we recently identified a transmitted/founder (T/F) virus (ZP6248) that efficiently used an alternative coreceptor GPR15, rather than commonly used CXCR4 and CCR5, to establish clinical infection. To understand which regions in the env gene were critical for the atypical coreceptor usage, we generated a set of V3 mutants and determined their infectivity in GHOST cells that expressed different coreceptors. When the variable loop 3 (V3) in YU2 was replaced with the ZP6248 V3 (YU2.6248V3), the chimera YU2.6248V3 infected GPR15+ cells but not CCR5+ cells. To determine which amino acids in V3 was responsible for this phenotype change, each of the eight amino acids that differed from the subtype B consensus V3 was substituted with alanine. The G306A and S322A mutations significantly reduced the replication capacity of YU2.6248V3 in GPR15+ cells, while all other alanine substitutions at positions 307, 314, 315, 316, 317 and 318 completely abrogated the infectivity of YU2.6248V3 in GPR15+ cells. The E314A mutation, as the E314G mutation reported before, also rendered the YU2.6248V3 infectious in CCR5+ cells, while none of other alanine mutants could infect CCR5+ cells. These results demonstrated that amino acids in ZP6248 V3 might form a unique conformation that was critical for the interaction with GPR15 while the amino acids at position 314 in the V3 crown of ZP6248 played a key role in interaction with both CCR5 and GPR15. The unique phenotypes of ZP6248 can serve as a model to understand how HIV-1 explores the diverse coreceptor reservoir through novel genetic variants to establish clinical infection.

Show MeSH
Related in: MedlinePlus