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Suppression of a Natural Killer Cell Response by Simian Immunodeficiency Virus Peptides.

Schafer JL, Ries M, Guha N, Connole M, Colantonio AD, Wiertz EJ, Wilson NA, Kaur A, Evans DT - PLoS Pathog. (2015)

Bottom Line: The functional effects of these peptide variants on NK cell responses also corresponded to their effects on Mamu-A1*002 tetramer binding to Mamu-KIR3DL05.In assays with mixtures of inhibitory and disinhibitory peptides, low concentrations of inhibitory peptides dominated to suppress NK cell responses.Consistent with the inhibition of Mamu-KIR3DL05+ NK cells by viral epitopes presented by Mamu-A1*002, SIV replication was significantly higher in Mamu-A1*002+ CD4+ lymphocytes co-cultured with Mamu-KIR3DL05+ NK cells than with Mamu-KIR3DL05- NK cells.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology and Immunobiology, Harvard Medical School, New England Primate Research Center, Southborough, Massachusetts, United States of America.

ABSTRACT
Natural killer (NK) cell responses in primates are regulated in part through interactions between two highly polymorphic molecules, the killer-cell immunoglobulin-like receptors (KIRs) on NK cells and their major histocompatibility complex (MHC) class I ligands on target cells. We previously reported that the binding of a common MHC class I molecule in the rhesus macaque, Mamu-A1*002, to the inhibitory receptor Mamu-KIR3DL05 is stabilized by certain simian immunodeficiency virus (SIV) peptides, but not by others. Here we investigated the functional implications of these interactions by testing SIV peptides bound by Mamu-A1*002 for the ability to modulate Mamu-KIR3DL05+ NK cell responses. Twenty-eight of 75 SIV peptides bound by Mamu-A1*002 suppressed the cytolytic activity of primary Mamu-KIR3DL05+ NK cells, including three immunodominant CD8+ T cell epitopes previously shown to stabilize Mamu-A1*002 tetramer binding to Mamu-KIR3DL05. Substitutions at C-terminal positions changed inhibitory peptides into disinhibitory peptides, and vice versa, without altering binding to Mamu-A1*002. The functional effects of these peptide variants on NK cell responses also corresponded to their effects on Mamu-A1*002 tetramer binding to Mamu-KIR3DL05. In assays with mixtures of inhibitory and disinhibitory peptides, low concentrations of inhibitory peptides dominated to suppress NK cell responses. Consistent with the inhibition of Mamu-KIR3DL05+ NK cells by viral epitopes presented by Mamu-A1*002, SIV replication was significantly higher in Mamu-A1*002+ CD4+ lymphocytes co-cultured with Mamu-KIR3DL05+ NK cells than with Mamu-KIR3DL05- NK cells. These results demonstrate that viral peptides can differentially affect NK cell responses by modulating MHC class I interactions with inhibitory KIRs, and provide a mechanism by which immunodeficiency viruses may evade NK cell responses.

No MeSH data available.


Related in: MedlinePlus

Signals from inhibitory peptides dominate to suppress NK cell activation.721.221-ICP47-A1*002 cells were pulsed with mixtures of Gag GY9 and GY9 L8W (A, B & E) or Env RY8 and RY8 V7W (C, D & F) and tested for susceptibility to killing by Mamu-KIR3DL05+ NK cells in CAM cytotoxicity assays. Representative data (A & C) and mean percent specific lysis (B & D) are shown for three independent experiments using NK cells from different animals. In panels A-D, the percentages of inhibitory versus disinhibitory peptides varied, keeping the total peptide concentration constant at 0.5 μM for GY9/GY9 L8W and 5 μM for RY8/RY8 V7W. Error bars indicate +1 SD and asterisks indicate significant differences in the lysis of target cells pulsed with GY9 or RY8 compared to target cells pulsed with the indicated peptide mixtures (*p<0.05, ****p<0.001 by two-way ANOVA with Dunnett’s test). In panels E-F, target cells were pulsed with increasing concentrations of Gag GY9 (E) or Env RY8 (F) in combination with a fixed concentration of their respective disinhibitory variants (0.375 μM GY9 L8W or 3.75 μM RY8 V7W). The dashed line indicates 50% inhibition where GY9 or RY8 alone defines 100% inhibition and GY9 L8W or RY8 V7W alone defines 0% inhibition. Mamu-A1*002 stabilization on the surface of 721.221-ICP47-A1*002 cells was verified by flow cytometry using the MHC class I-specific monoclonal antibody W6/32 (S5 Fig).
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ppat.1005145.g006: Signals from inhibitory peptides dominate to suppress NK cell activation.721.221-ICP47-A1*002 cells were pulsed with mixtures of Gag GY9 and GY9 L8W (A, B & E) or Env RY8 and RY8 V7W (C, D & F) and tested for susceptibility to killing by Mamu-KIR3DL05+ NK cells in CAM cytotoxicity assays. Representative data (A & C) and mean percent specific lysis (B & D) are shown for three independent experiments using NK cells from different animals. In panels A-D, the percentages of inhibitory versus disinhibitory peptides varied, keeping the total peptide concentration constant at 0.5 μM for GY9/GY9 L8W and 5 μM for RY8/RY8 V7W. Error bars indicate +1 SD and asterisks indicate significant differences in the lysis of target cells pulsed with GY9 or RY8 compared to target cells pulsed with the indicated peptide mixtures (*p<0.05, ****p<0.001 by two-way ANOVA with Dunnett’s test). In panels E-F, target cells were pulsed with increasing concentrations of Gag GY9 (E) or Env RY8 (F) in combination with a fixed concentration of their respective disinhibitory variants (0.375 μM GY9 L8W or 3.75 μM RY8 V7W). The dashed line indicates 50% inhibition where GY9 or RY8 alone defines 100% inhibition and GY9 L8W or RY8 V7W alone defines 0% inhibition. Mamu-A1*002 stabilization on the surface of 721.221-ICP47-A1*002 cells was verified by flow cytometry using the MHC class I-specific monoclonal antibody W6/32 (S5 Fig).

Mentions: MHC class I molecules present a diverse repertoire of peptides derived from viral and cellular antigens on the surface of virus-infected cells, of which some may stabilize and some may disrupt interactions with any given KIR. We therefore asked which signal dominates when a mixture of peptides that do or do not suppress NK cell activity is presented on the cell surface. 721.221-ICP47-A1*002 cells were pulsed with mixtures of Gag GY9 and GY9 L8W or Env RY8 and RY8 V7W at concentrations that did not over-saturate binding to Mamu-A1*002 (S5A Fig), and were tested for susceptibility to lysis by Mamu-KIR3DL05+ NK cells. Cell surface stabilization of Mamu-A1*002 was similar for GY9 versus GY9 L8W and RY8 versus RY8 V7W individually (S5A Fig), or in combination (S5B Fig), and none of the peptide mixtures affected Mamu-KIR3DL05- NK cell responses (S5C Fig). At 25% of the total peptide concentration, Gag GY9 and Env RY8 dominated over their respective disinhibitory variants to fully suppress the cytolytic activity of Mamu-KIR3DL05+ NK cells (Fig 6A–6D). To determine the threshold at which the inhibitory effects of these peptides are observed, increasing concentrations of Gag GY9 and Env RY8 were tested in the presence of a fixed, sub-saturating concentration of Gag GY9 L8W and Env RY8 V7W. Half-maximal inhibition of killing was achieved at approximately 5% of the total peptide concentration for both Gag GY9 and Env RY8 (Fig 6E and 6F). Thus, under conditions where inhibitory peptides and their corresponding disinhibitory variants are presented simultaneously by Mamu-A1*002, the inhibitory peptides dominate to suppress Mamu-KIR3DL05+ NK cell responses.


Suppression of a Natural Killer Cell Response by Simian Immunodeficiency Virus Peptides.

Schafer JL, Ries M, Guha N, Connole M, Colantonio AD, Wiertz EJ, Wilson NA, Kaur A, Evans DT - PLoS Pathog. (2015)

Signals from inhibitory peptides dominate to suppress NK cell activation.721.221-ICP47-A1*002 cells were pulsed with mixtures of Gag GY9 and GY9 L8W (A, B & E) or Env RY8 and RY8 V7W (C, D & F) and tested for susceptibility to killing by Mamu-KIR3DL05+ NK cells in CAM cytotoxicity assays. Representative data (A & C) and mean percent specific lysis (B & D) are shown for three independent experiments using NK cells from different animals. In panels A-D, the percentages of inhibitory versus disinhibitory peptides varied, keeping the total peptide concentration constant at 0.5 μM for GY9/GY9 L8W and 5 μM for RY8/RY8 V7W. Error bars indicate +1 SD and asterisks indicate significant differences in the lysis of target cells pulsed with GY9 or RY8 compared to target cells pulsed with the indicated peptide mixtures (*p<0.05, ****p<0.001 by two-way ANOVA with Dunnett’s test). In panels E-F, target cells were pulsed with increasing concentrations of Gag GY9 (E) or Env RY8 (F) in combination with a fixed concentration of their respective disinhibitory variants (0.375 μM GY9 L8W or 3.75 μM RY8 V7W). The dashed line indicates 50% inhibition where GY9 or RY8 alone defines 100% inhibition and GY9 L8W or RY8 V7W alone defines 0% inhibition. Mamu-A1*002 stabilization on the surface of 721.221-ICP47-A1*002 cells was verified by flow cytometry using the MHC class I-specific monoclonal antibody W6/32 (S5 Fig).
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Related In: Results  -  Collection

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ppat.1005145.g006: Signals from inhibitory peptides dominate to suppress NK cell activation.721.221-ICP47-A1*002 cells were pulsed with mixtures of Gag GY9 and GY9 L8W (A, B & E) or Env RY8 and RY8 V7W (C, D & F) and tested for susceptibility to killing by Mamu-KIR3DL05+ NK cells in CAM cytotoxicity assays. Representative data (A & C) and mean percent specific lysis (B & D) are shown for three independent experiments using NK cells from different animals. In panels A-D, the percentages of inhibitory versus disinhibitory peptides varied, keeping the total peptide concentration constant at 0.5 μM for GY9/GY9 L8W and 5 μM for RY8/RY8 V7W. Error bars indicate +1 SD and asterisks indicate significant differences in the lysis of target cells pulsed with GY9 or RY8 compared to target cells pulsed with the indicated peptide mixtures (*p<0.05, ****p<0.001 by two-way ANOVA with Dunnett’s test). In panels E-F, target cells were pulsed with increasing concentrations of Gag GY9 (E) or Env RY8 (F) in combination with a fixed concentration of their respective disinhibitory variants (0.375 μM GY9 L8W or 3.75 μM RY8 V7W). The dashed line indicates 50% inhibition where GY9 or RY8 alone defines 100% inhibition and GY9 L8W or RY8 V7W alone defines 0% inhibition. Mamu-A1*002 stabilization on the surface of 721.221-ICP47-A1*002 cells was verified by flow cytometry using the MHC class I-specific monoclonal antibody W6/32 (S5 Fig).
Mentions: MHC class I molecules present a diverse repertoire of peptides derived from viral and cellular antigens on the surface of virus-infected cells, of which some may stabilize and some may disrupt interactions with any given KIR. We therefore asked which signal dominates when a mixture of peptides that do or do not suppress NK cell activity is presented on the cell surface. 721.221-ICP47-A1*002 cells were pulsed with mixtures of Gag GY9 and GY9 L8W or Env RY8 and RY8 V7W at concentrations that did not over-saturate binding to Mamu-A1*002 (S5A Fig), and were tested for susceptibility to lysis by Mamu-KIR3DL05+ NK cells. Cell surface stabilization of Mamu-A1*002 was similar for GY9 versus GY9 L8W and RY8 versus RY8 V7W individually (S5A Fig), or in combination (S5B Fig), and none of the peptide mixtures affected Mamu-KIR3DL05- NK cell responses (S5C Fig). At 25% of the total peptide concentration, Gag GY9 and Env RY8 dominated over their respective disinhibitory variants to fully suppress the cytolytic activity of Mamu-KIR3DL05+ NK cells (Fig 6A–6D). To determine the threshold at which the inhibitory effects of these peptides are observed, increasing concentrations of Gag GY9 and Env RY8 were tested in the presence of a fixed, sub-saturating concentration of Gag GY9 L8W and Env RY8 V7W. Half-maximal inhibition of killing was achieved at approximately 5% of the total peptide concentration for both Gag GY9 and Env RY8 (Fig 6E and 6F). Thus, under conditions where inhibitory peptides and their corresponding disinhibitory variants are presented simultaneously by Mamu-A1*002, the inhibitory peptides dominate to suppress Mamu-KIR3DL05+ NK cell responses.

Bottom Line: The functional effects of these peptide variants on NK cell responses also corresponded to their effects on Mamu-A1*002 tetramer binding to Mamu-KIR3DL05.In assays with mixtures of inhibitory and disinhibitory peptides, low concentrations of inhibitory peptides dominated to suppress NK cell responses.Consistent with the inhibition of Mamu-KIR3DL05+ NK cells by viral epitopes presented by Mamu-A1*002, SIV replication was significantly higher in Mamu-A1*002+ CD4+ lymphocytes co-cultured with Mamu-KIR3DL05+ NK cells than with Mamu-KIR3DL05- NK cells.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology and Immunobiology, Harvard Medical School, New England Primate Research Center, Southborough, Massachusetts, United States of America.

ABSTRACT
Natural killer (NK) cell responses in primates are regulated in part through interactions between two highly polymorphic molecules, the killer-cell immunoglobulin-like receptors (KIRs) on NK cells and their major histocompatibility complex (MHC) class I ligands on target cells. We previously reported that the binding of a common MHC class I molecule in the rhesus macaque, Mamu-A1*002, to the inhibitory receptor Mamu-KIR3DL05 is stabilized by certain simian immunodeficiency virus (SIV) peptides, but not by others. Here we investigated the functional implications of these interactions by testing SIV peptides bound by Mamu-A1*002 for the ability to modulate Mamu-KIR3DL05+ NK cell responses. Twenty-eight of 75 SIV peptides bound by Mamu-A1*002 suppressed the cytolytic activity of primary Mamu-KIR3DL05+ NK cells, including three immunodominant CD8+ T cell epitopes previously shown to stabilize Mamu-A1*002 tetramer binding to Mamu-KIR3DL05. Substitutions at C-terminal positions changed inhibitory peptides into disinhibitory peptides, and vice versa, without altering binding to Mamu-A1*002. The functional effects of these peptide variants on NK cell responses also corresponded to their effects on Mamu-A1*002 tetramer binding to Mamu-KIR3DL05. In assays with mixtures of inhibitory and disinhibitory peptides, low concentrations of inhibitory peptides dominated to suppress NK cell responses. Consistent with the inhibition of Mamu-KIR3DL05+ NK cells by viral epitopes presented by Mamu-A1*002, SIV replication was significantly higher in Mamu-A1*002+ CD4+ lymphocytes co-cultured with Mamu-KIR3DL05+ NK cells than with Mamu-KIR3DL05- NK cells. These results demonstrate that viral peptides can differentially affect NK cell responses by modulating MHC class I interactions with inhibitory KIRs, and provide a mechanism by which immunodeficiency viruses may evade NK cell responses.

No MeSH data available.


Related in: MedlinePlus