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Identities of P2 and P3 Residues of H-2Kb-Bound Peptides Determine Mouse Ly49C Recognition.

Marquez EA, Kane KP - PLoS ONE (2015)

Bottom Line: Despite this ability, little is understood regarding the properties that enable some peptides, when bound to MHC-I molecules, to support Ly49 recognition, but not others.Using RMA-S target cells expressing MHC-I molecules loaded with individual peptides and effector cells expressing the ectodomain of the inhibitory Ly49C receptor, we found that two adjacent amino acid residues, P2 and P3, both buried in the peptide binding groove of H-2Kb, determine mouse Ly49C specificity.If both are aliphatic residues, this is supportive.

View Article: PubMed Central - PubMed

Affiliation: Medical Microbiology and Immunology, University of Alberta, Edmonton, Alberta, Canada.

ABSTRACT
Ly49 receptors can be peptide selective in their recognition of MHC-I-peptide complexes, affording them a level of discrimination beyond detecting the presence or absence of specific MHC-I allele products. Despite this ability, little is understood regarding the properties that enable some peptides, when bound to MHC-I molecules, to support Ly49 recognition, but not others. Using RMA-S target cells expressing MHC-I molecules loaded with individual peptides and effector cells expressing the ectodomain of the inhibitory Ly49C receptor, we found that two adjacent amino acid residues, P2 and P3, both buried in the peptide binding groove of H-2Kb, determine mouse Ly49C specificity. If both are aliphatic residues, this is supportive. Whereas, small amino acids at P2 and aromatic amino acids at the P3 auxiliary anchor residue are detrimental to Ly49C recognition. These results resemble those with a rat Ly49 where the identity of a peptide anchor residue determines recognition, suggesting that dependence on specific peptide residues buried in the MHC-I peptide-binding groove may be fundamental to Ly49 peptide selectivity and recognition.

No MeSH data available.


Related in: MedlinePlus

Ly49C peptide dependent recognition of H-2Kb decreases with peptides bearing non-polar aliphatic auxiliary anchor residues.(A) RMA-S cell surface expression of H-2Kb when incubated with the indicated peptides, or no peptide. Shaded histogram corresponds to isotype control. (B) Cytotoxicity assay using RMA-S cells loaded with peptides that have aromatic R groups at P3 co-incubated with RNK.49WC effector cells. (C) Statistically significant changes in Ly49W/C recognition of H-2Kb, with respect to the positive control for RNK.49W/C recognition (SIINFEKL-loaded RMA/S cells), were performed at the 12.5:1 E:T ratio from cytotoxicity assays (*p < 0.05, **p < 0.005). Results plotted are the mean of three independent experiments with error bars representing SD.
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pone.0131308.g002: Ly49C peptide dependent recognition of H-2Kb decreases with peptides bearing non-polar aliphatic auxiliary anchor residues.(A) RMA-S cell surface expression of H-2Kb when incubated with the indicated peptides, or no peptide. Shaded histogram corresponds to isotype control. (B) Cytotoxicity assay using RMA-S cells loaded with peptides that have aromatic R groups at P3 co-incubated with RNK.49WC effector cells. (C) Statistically significant changes in Ly49W/C recognition of H-2Kb, with respect to the positive control for RNK.49W/C recognition (SIINFEKL-loaded RMA/S cells), were performed at the 12.5:1 E:T ratio from cytotoxicity assays (*p < 0.05, **p < 0.005). Results plotted are the mean of three independent experiments with error bars representing SD.

Mentions: From the initial set of peptides used in this study we found that along with SIINFEKL, peptides KIITYRNL, KVITFIDL, and YAMIYRNL stabilized H-2Kb on RMA/S cells (Fig 1A) and similarly supported Ly49W/C recognition (Fig 1B and 1C). On the other hand, although capable of yielding similar H-2Kb expression on RMA/S cells (Fig 2A), peptides ANYDFICV, INFDFPKL, and ISFKFDHL did not, or poorly supported H-2Kb and Ly49W/C interaction, (Fig 2B and 2C) resembling RGYVYQGL and AAYAYAAL peptides previously shown to be unsupportive of H-2Kb and Ly49C interaction [22]. Peptides that supported H-2Kb and Ly49W/C interaction, SIINFEKL, KIITYRNL, KVITFIDL and YAMIYRNL have the same P8 residues, conserved aromatic P5 residues and non-polar aliphatic P3 residues. While peptides that did not confer Ly49W/C and H-2Kb interaction, ANYDFICV, INFDFPKL, ISFKFDHL, RGYVYQGL and AAYAYAAL have similar P8 and P5 residues, but have aromatic residues at P3. Therefore, given the correlation between the identity of auxiliary anchor residue at P3 and Ly49W/C recognition, as noted in Table 3, we hypothesized that peptides bearing non-polar aliphatic residues at P3 are supportive of receptor and ligand interaction, while aromatic residues at P3 are detrimental for H-2Kb and Ly49W/C association.


Identities of P2 and P3 Residues of H-2Kb-Bound Peptides Determine Mouse Ly49C Recognition.

Marquez EA, Kane KP - PLoS ONE (2015)

Ly49C peptide dependent recognition of H-2Kb decreases with peptides bearing non-polar aliphatic auxiliary anchor residues.(A) RMA-S cell surface expression of H-2Kb when incubated with the indicated peptides, or no peptide. Shaded histogram corresponds to isotype control. (B) Cytotoxicity assay using RMA-S cells loaded with peptides that have aromatic R groups at P3 co-incubated with RNK.49WC effector cells. (C) Statistically significant changes in Ly49W/C recognition of H-2Kb, with respect to the positive control for RNK.49W/C recognition (SIINFEKL-loaded RMA/S cells), were performed at the 12.5:1 E:T ratio from cytotoxicity assays (*p < 0.05, **p < 0.005). Results plotted are the mean of three independent experiments with error bars representing SD.
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4493100&req=5

pone.0131308.g002: Ly49C peptide dependent recognition of H-2Kb decreases with peptides bearing non-polar aliphatic auxiliary anchor residues.(A) RMA-S cell surface expression of H-2Kb when incubated with the indicated peptides, or no peptide. Shaded histogram corresponds to isotype control. (B) Cytotoxicity assay using RMA-S cells loaded with peptides that have aromatic R groups at P3 co-incubated with RNK.49WC effector cells. (C) Statistically significant changes in Ly49W/C recognition of H-2Kb, with respect to the positive control for RNK.49W/C recognition (SIINFEKL-loaded RMA/S cells), were performed at the 12.5:1 E:T ratio from cytotoxicity assays (*p < 0.05, **p < 0.005). Results plotted are the mean of three independent experiments with error bars representing SD.
Mentions: From the initial set of peptides used in this study we found that along with SIINFEKL, peptides KIITYRNL, KVITFIDL, and YAMIYRNL stabilized H-2Kb on RMA/S cells (Fig 1A) and similarly supported Ly49W/C recognition (Fig 1B and 1C). On the other hand, although capable of yielding similar H-2Kb expression on RMA/S cells (Fig 2A), peptides ANYDFICV, INFDFPKL, and ISFKFDHL did not, or poorly supported H-2Kb and Ly49W/C interaction, (Fig 2B and 2C) resembling RGYVYQGL and AAYAYAAL peptides previously shown to be unsupportive of H-2Kb and Ly49C interaction [22]. Peptides that supported H-2Kb and Ly49W/C interaction, SIINFEKL, KIITYRNL, KVITFIDL and YAMIYRNL have the same P8 residues, conserved aromatic P5 residues and non-polar aliphatic P3 residues. While peptides that did not confer Ly49W/C and H-2Kb interaction, ANYDFICV, INFDFPKL, ISFKFDHL, RGYVYQGL and AAYAYAAL have similar P8 and P5 residues, but have aromatic residues at P3. Therefore, given the correlation between the identity of auxiliary anchor residue at P3 and Ly49W/C recognition, as noted in Table 3, we hypothesized that peptides bearing non-polar aliphatic residues at P3 are supportive of receptor and ligand interaction, while aromatic residues at P3 are detrimental for H-2Kb and Ly49W/C association.

Bottom Line: Despite this ability, little is understood regarding the properties that enable some peptides, when bound to MHC-I molecules, to support Ly49 recognition, but not others.Using RMA-S target cells expressing MHC-I molecules loaded with individual peptides and effector cells expressing the ectodomain of the inhibitory Ly49C receptor, we found that two adjacent amino acid residues, P2 and P3, both buried in the peptide binding groove of H-2Kb, determine mouse Ly49C specificity.If both are aliphatic residues, this is supportive.

View Article: PubMed Central - PubMed

Affiliation: Medical Microbiology and Immunology, University of Alberta, Edmonton, Alberta, Canada.

ABSTRACT
Ly49 receptors can be peptide selective in their recognition of MHC-I-peptide complexes, affording them a level of discrimination beyond detecting the presence or absence of specific MHC-I allele products. Despite this ability, little is understood regarding the properties that enable some peptides, when bound to MHC-I molecules, to support Ly49 recognition, but not others. Using RMA-S target cells expressing MHC-I molecules loaded with individual peptides and effector cells expressing the ectodomain of the inhibitory Ly49C receptor, we found that two adjacent amino acid residues, P2 and P3, both buried in the peptide binding groove of H-2Kb, determine mouse Ly49C specificity. If both are aliphatic residues, this is supportive. Whereas, small amino acids at P2 and aromatic amino acids at the P3 auxiliary anchor residue are detrimental to Ly49C recognition. These results resemble those with a rat Ly49 where the identity of a peptide anchor residue determines recognition, suggesting that dependence on specific peptide residues buried in the MHC-I peptide-binding groove may be fundamental to Ly49 peptide selectivity and recognition.

No MeSH data available.


Related in: MedlinePlus