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The D0 domain of KIR3D acts as a major histocompatibility complex class I binding enhancer.

Khakoo SI, Geller R, Shin S, Jenkins JA, Parham P - J. Exp. Med. (2002)

Bottom Line: In contrast point mutations in the D1 and D2 domains of KIR3DL1, made from knowledge of KIR2D:HLA-C interactions, disrupted binding to Bw4(+) HLA-B.The results are consistent with a model in which D1 and D2 make the principal contacts between KIR3DL1 and HLA-B while D0 acts through a different mechanism to enhance the interaction.This modulatory role for D0 is compatible with natural loss of expression of the D0 domain, a repeated event in the evolution of functional KIR genes.

View Article: PubMed Central - PubMed

Affiliation: Department of Structural Biology, Stanford University School of Medicine, CA 94305, USA.

ABSTRACT
In contrast to the KIR2D:HLA-C interaction, little is known of KIR3DL1's interaction with HLA-B or the role of D0, the domain not present in KIR2D. Differences in the strength and specificity for major histocompatibility complex class I of KIR3DL1 and its common chimpanzee homologue Pt-KIR3DL1/2 were exploited to address these questions. Domain-swap, deletion, and site-directed mutants of KIR3DL1 were analyzed for HLA-B binding using a novel, positively signaling cell-cell binding assay. Natural 'deletion' of residues 50 and 51 from its D0 domain causes Pt-KIR3DL1/2 to bind Bw4(+) HLA-B allotypes more avidly than does KIR3DL1. Deletion of these residues from KIR3DL1, or their substitution for alanine, enhanced binding of Bw4(+) HLA-B. None of 15 different point mutations in D0 abrogated KIR3DL1 binding to Bw4(+) HLA-B. In contrast point mutations in the D1 and D2 domains of KIR3DL1, made from knowledge of KIR2D:HLA-C interactions, disrupted binding to Bw4(+) HLA-B. The results are consistent with a model in which D1 and D2 make the principal contacts between KIR3DL1 and HLA-B while D0 acts through a different mechanism to enhance the interaction. This modulatory role for D0 is compatible with natural loss of expression of the D0 domain, a repeated event in the evolution of functional KIR genes.

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The D0 domain of Pt-KIR3DL1/2 stabilizes the interaction with MHC-B. Domain swap constructs of KIR3DL1ζ and Pt-KIR3DL1ζ were tested against a panel of 721.221 transfectants expressing various HLA-B allotypes in SEAP assays. The Ig domain composition of the three constructs tested are shown. Human Ig domains are indicated with stippled boxes and chimpanzee Ig domains with filled boxes. The reactivities of the constructs in the SEAP assays are shown in comparison to stimulation with plate bound anti-CD3. The means and standard errors of three independent experiments are shown.
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fig3: The D0 domain of Pt-KIR3DL1/2 stabilizes the interaction with MHC-B. Domain swap constructs of KIR3DL1ζ and Pt-KIR3DL1ζ were tested against a panel of 721.221 transfectants expressing various HLA-B allotypes in SEAP assays. The Ig domain composition of the three constructs tested are shown. Human Ig domains are indicated with stippled boxes and chimpanzee Ig domains with filled boxes. The reactivities of the constructs in the SEAP assays are shown in comparison to stimulation with plate bound anti-CD3. The means and standard errors of three independent experiments are shown.

Mentions: To assess the contribution of individual Ig-like domains to differences in HLA-B recognition by KIR3DL1 and Pt-KIR3DL1/2, we made six constructs from KIR3DL1ζ and Pt-KIR3DL1/2ζ in which the sequences encoding individual human and chimpanzee domains were recombined in all possible combinations. Of these, three failed to give a detectable cell-surface protein upon transfection into Jurkat cells. The three expressed recombinant mutants analyzed for HLA-B binding consisted of HCCζ and CHHζ in which the D0 domains were swapped, and HCHζ in which the D1 domain of KIR3DL1 was replaced with that of Pt-KIR3DL1/2 (Fig. 3) . Mutant CHHζ, in which chimpanzee D0 replaces human D0, exhibited stronger reactions with all three Bw4+ HLA-B allotypes tested, but no reactivity with the Bw6+ HLA-B*1502. The reciprocal mutant, in which human D0 is associated with chimpanzee D1 and D2, gave a pattern of reactivity B*3801 >> B*5801 > B*1513, comparable to that of KIR3DL1ζ and lacking reaction with B*1502. Thus, for Pt-KIR3DL1/2, reactivity with Bw6+ B*1502, but not Bw4+ HLA-B, is dependent upon the D0 domain and upon features that distinguish the chimpanzee D0 domain from its human counterpart. The reactivity of mutant HCHζ was B*3801 > B*5801 ≈ B*1513 ≈ B*1502, a pattern distinct from that of KIR3DL1 and which includes the Bw6+ allotype B*1502. This suggests that substitutions in the chimpanzee D1 domain also contribute to the recognition of HLA-B*1502.


The D0 domain of KIR3D acts as a major histocompatibility complex class I binding enhancer.

Khakoo SI, Geller R, Shin S, Jenkins JA, Parham P - J. Exp. Med. (2002)

The D0 domain of Pt-KIR3DL1/2 stabilizes the interaction with MHC-B. Domain swap constructs of KIR3DL1ζ and Pt-KIR3DL1ζ were tested against a panel of 721.221 transfectants expressing various HLA-B allotypes in SEAP assays. The Ig domain composition of the three constructs tested are shown. Human Ig domains are indicated with stippled boxes and chimpanzee Ig domains with filled boxes. The reactivities of the constructs in the SEAP assays are shown in comparison to stimulation with plate bound anti-CD3. The means and standard errors of three independent experiments are shown.
© Copyright Policy
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC2194033&req=5

fig3: The D0 domain of Pt-KIR3DL1/2 stabilizes the interaction with MHC-B. Domain swap constructs of KIR3DL1ζ and Pt-KIR3DL1ζ were tested against a panel of 721.221 transfectants expressing various HLA-B allotypes in SEAP assays. The Ig domain composition of the three constructs tested are shown. Human Ig domains are indicated with stippled boxes and chimpanzee Ig domains with filled boxes. The reactivities of the constructs in the SEAP assays are shown in comparison to stimulation with plate bound anti-CD3. The means and standard errors of three independent experiments are shown.
Mentions: To assess the contribution of individual Ig-like domains to differences in HLA-B recognition by KIR3DL1 and Pt-KIR3DL1/2, we made six constructs from KIR3DL1ζ and Pt-KIR3DL1/2ζ in which the sequences encoding individual human and chimpanzee domains were recombined in all possible combinations. Of these, three failed to give a detectable cell-surface protein upon transfection into Jurkat cells. The three expressed recombinant mutants analyzed for HLA-B binding consisted of HCCζ and CHHζ in which the D0 domains were swapped, and HCHζ in which the D1 domain of KIR3DL1 was replaced with that of Pt-KIR3DL1/2 (Fig. 3) . Mutant CHHζ, in which chimpanzee D0 replaces human D0, exhibited stronger reactions with all three Bw4+ HLA-B allotypes tested, but no reactivity with the Bw6+ HLA-B*1502. The reciprocal mutant, in which human D0 is associated with chimpanzee D1 and D2, gave a pattern of reactivity B*3801 >> B*5801 > B*1513, comparable to that of KIR3DL1ζ and lacking reaction with B*1502. Thus, for Pt-KIR3DL1/2, reactivity with Bw6+ B*1502, but not Bw4+ HLA-B, is dependent upon the D0 domain and upon features that distinguish the chimpanzee D0 domain from its human counterpart. The reactivity of mutant HCHζ was B*3801 > B*5801 ≈ B*1513 ≈ B*1502, a pattern distinct from that of KIR3DL1 and which includes the Bw6+ allotype B*1502. This suggests that substitutions in the chimpanzee D1 domain also contribute to the recognition of HLA-B*1502.

Bottom Line: In contrast point mutations in the D1 and D2 domains of KIR3DL1, made from knowledge of KIR2D:HLA-C interactions, disrupted binding to Bw4(+) HLA-B.The results are consistent with a model in which D1 and D2 make the principal contacts between KIR3DL1 and HLA-B while D0 acts through a different mechanism to enhance the interaction.This modulatory role for D0 is compatible with natural loss of expression of the D0 domain, a repeated event in the evolution of functional KIR genes.

View Article: PubMed Central - PubMed

Affiliation: Department of Structural Biology, Stanford University School of Medicine, CA 94305, USA.

ABSTRACT
In contrast to the KIR2D:HLA-C interaction, little is known of KIR3DL1's interaction with HLA-B or the role of D0, the domain not present in KIR2D. Differences in the strength and specificity for major histocompatibility complex class I of KIR3DL1 and its common chimpanzee homologue Pt-KIR3DL1/2 were exploited to address these questions. Domain-swap, deletion, and site-directed mutants of KIR3DL1 were analyzed for HLA-B binding using a novel, positively signaling cell-cell binding assay. Natural 'deletion' of residues 50 and 51 from its D0 domain causes Pt-KIR3DL1/2 to bind Bw4(+) HLA-B allotypes more avidly than does KIR3DL1. Deletion of these residues from KIR3DL1, or their substitution for alanine, enhanced binding of Bw4(+) HLA-B. None of 15 different point mutations in D0 abrogated KIR3DL1 binding to Bw4(+) HLA-B. In contrast point mutations in the D1 and D2 domains of KIR3DL1, made from knowledge of KIR2D:HLA-C interactions, disrupted binding to Bw4(+) HLA-B. The results are consistent with a model in which D1 and D2 make the principal contacts between KIR3DL1 and HLA-B while D0 acts through a different mechanism to enhance the interaction. This modulatory role for D0 is compatible with natural loss of expression of the D0 domain, a repeated event in the evolution of functional KIR genes.

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