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CLEC9A is a novel activation C-type lectin-like receptor expressed on BDCA3+ dendritic cells and a subset of monocytes.

Huysamen C, Willment JA, Dennehy KM, Brown GD - J. Biol. Chem. (2008)

Bottom Line: We describe here the first characterization of CLEC9A, a group V C-type lectin-like receptor located in the "Dectin-1 cluster" of related receptors, which are encoded within the natural killer (NK)-gene complex.CLEC9A is expressed at the cell surface as a glycosylated dimer and can mediate endocytosis, but not phagocytosis.These data indicate that CLEC9A functions as an activation receptor.

View Article: PubMed Central - PubMed

Affiliation: Division of Immunology, Institute of Infectious Disease and Molecular Medicine, University of Cape Town, Observatory, 7925, South Africa.

ABSTRACT
We describe here the first characterization of CLEC9A, a group V C-type lectin-like receptor located in the "Dectin-1 cluster" of related receptors, which are encoded within the natural killer (NK)-gene complex. Expression of human CLEC9A is highly restricted in peripheral blood, being detected only on BDCA3(+) dendritic cells and on a small subset of CD14(+)CD16(-) monocytes. CLEC9A is expressed at the cell surface as a glycosylated dimer and can mediate endocytosis, but not phagocytosis. CLEC9A possesses a cytoplasmic immunoreceptor tyrosine-based activation-like motif that can recruit Syk kinase, and we demonstrate, using receptor chimeras, that this receptor can induce proinflammatory cytokine production. These data indicate that CLEC9A functions as an activation receptor.

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hCLEC9A can induce pro-inflammatory cytokine production and can recruit and signal via Syk kinase. A, quantitation of zymosan (zym) induced TNF production by transduced RAW264.7 macrophages, in the presence or absence of soluble β-glucan (β-glu), as indicated, demonstrating the ability of the chimeric receptor to induce proinflammatory cytokine production. Shown are the mean ± S.D. of one representative experiment of three. B, Western blotting of immunoprecipitates using phosphorylated (pY) and unphosphorylated peptides (Y) corresponding to the cytoplasmic tail of hCLEC9A or Dectin-1 from RAW264.7 cell lysates. Blots were probed with anti-phosphotyrosine (αPY), anti-Syk, and anti-Lyn, as indicated. C, IL-2 production following zymosan stimulation of Syk-deficient (Syk-) and Syk-sufficient (Syk+) cells, transduced with the chimeric receptor or vector-only control, as indicated, showing that cytokine production induced by the chimeric receptor in response to zymosan requires Syk. The data shown are the mean ± S.D. and are representative of three independent experiments. *, p < 0.05 versus control (Student's t test).
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fig6: hCLEC9A can induce pro-inflammatory cytokine production and can recruit and signal via Syk kinase. A, quantitation of zymosan (zym) induced TNF production by transduced RAW264.7 macrophages, in the presence or absence of soluble β-glucan (β-glu), as indicated, demonstrating the ability of the chimeric receptor to induce proinflammatory cytokine production. Shown are the mean ± S.D. of one representative experiment of three. B, Western blotting of immunoprecipitates using phosphorylated (pY) and unphosphorylated peptides (Y) corresponding to the cytoplasmic tail of hCLEC9A or Dectin-1 from RAW264.7 cell lysates. Blots were probed with anti-phosphotyrosine (αPY), anti-Syk, and anti-Lyn, as indicated. C, IL-2 production following zymosan stimulation of Syk-deficient (Syk-) and Syk-sufficient (Syk+) cells, transduced with the chimeric receptor or vector-only control, as indicated, showing that cytokine production induced by the chimeric receptor in response to zymosan requires Syk. The data shown are the mean ± S.D. and are representative of three independent experiments. *, p < 0.05 versus control (Student's t test).

Mentions: CLEC9A Can Induce Inflammatory Cytokine Production—In addition to phagocytosis, the tyrosine-based motif of Dectin-1 can induce a variety of cellular responses, including the production of proinflammatory cytokines in response to zymosan (27). So we next explored the possibility that signaling via CLEC9A could similarly induce TNF production in the RAW264.7 macrophages transduced with the chimeric receptor. As described above, the expression of Dectin-1 and the chimeric receptor resulted in increased levels of zymosan binding in these cells (Fig. 5). Furthermore, both Dectin-1 (23) and the chimeric receptor induced high levels of TNF in response to zymosan (Fig. 6A). As we have previously shown that Dectin-1 lacking a cytoplasmic tail can mediate zymosan binding, but not the induction of TNF (23), our results indicate that the cellular response induced by the chimeric receptor is due to the presence of the CLEC9A cytoplasmic tail. Thus these data suggest that, like Dectin-1, CLEC9A signaling can induce a proinflammatory response.


CLEC9A is a novel activation C-type lectin-like receptor expressed on BDCA3+ dendritic cells and a subset of monocytes.

Huysamen C, Willment JA, Dennehy KM, Brown GD - J. Biol. Chem. (2008)

hCLEC9A can induce pro-inflammatory cytokine production and can recruit and signal via Syk kinase. A, quantitation of zymosan (zym) induced TNF production by transduced RAW264.7 macrophages, in the presence or absence of soluble β-glucan (β-glu), as indicated, demonstrating the ability of the chimeric receptor to induce proinflammatory cytokine production. Shown are the mean ± S.D. of one representative experiment of three. B, Western blotting of immunoprecipitates using phosphorylated (pY) and unphosphorylated peptides (Y) corresponding to the cytoplasmic tail of hCLEC9A or Dectin-1 from RAW264.7 cell lysates. Blots were probed with anti-phosphotyrosine (αPY), anti-Syk, and anti-Lyn, as indicated. C, IL-2 production following zymosan stimulation of Syk-deficient (Syk-) and Syk-sufficient (Syk+) cells, transduced with the chimeric receptor or vector-only control, as indicated, showing that cytokine production induced by the chimeric receptor in response to zymosan requires Syk. The data shown are the mean ± S.D. and are representative of three independent experiments. *, p < 0.05 versus control (Student's t test).
© Copyright Policy - open-access
Related In: Results  -  Collection

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fig6: hCLEC9A can induce pro-inflammatory cytokine production and can recruit and signal via Syk kinase. A, quantitation of zymosan (zym) induced TNF production by transduced RAW264.7 macrophages, in the presence or absence of soluble β-glucan (β-glu), as indicated, demonstrating the ability of the chimeric receptor to induce proinflammatory cytokine production. Shown are the mean ± S.D. of one representative experiment of three. B, Western blotting of immunoprecipitates using phosphorylated (pY) and unphosphorylated peptides (Y) corresponding to the cytoplasmic tail of hCLEC9A or Dectin-1 from RAW264.7 cell lysates. Blots were probed with anti-phosphotyrosine (αPY), anti-Syk, and anti-Lyn, as indicated. C, IL-2 production following zymosan stimulation of Syk-deficient (Syk-) and Syk-sufficient (Syk+) cells, transduced with the chimeric receptor or vector-only control, as indicated, showing that cytokine production induced by the chimeric receptor in response to zymosan requires Syk. The data shown are the mean ± S.D. and are representative of three independent experiments. *, p < 0.05 versus control (Student's t test).
Mentions: CLEC9A Can Induce Inflammatory Cytokine Production—In addition to phagocytosis, the tyrosine-based motif of Dectin-1 can induce a variety of cellular responses, including the production of proinflammatory cytokines in response to zymosan (27). So we next explored the possibility that signaling via CLEC9A could similarly induce TNF production in the RAW264.7 macrophages transduced with the chimeric receptor. As described above, the expression of Dectin-1 and the chimeric receptor resulted in increased levels of zymosan binding in these cells (Fig. 5). Furthermore, both Dectin-1 (23) and the chimeric receptor induced high levels of TNF in response to zymosan (Fig. 6A). As we have previously shown that Dectin-1 lacking a cytoplasmic tail can mediate zymosan binding, but not the induction of TNF (23), our results indicate that the cellular response induced by the chimeric receptor is due to the presence of the CLEC9A cytoplasmic tail. Thus these data suggest that, like Dectin-1, CLEC9A signaling can induce a proinflammatory response.

Bottom Line: We describe here the first characterization of CLEC9A, a group V C-type lectin-like receptor located in the "Dectin-1 cluster" of related receptors, which are encoded within the natural killer (NK)-gene complex.CLEC9A is expressed at the cell surface as a glycosylated dimer and can mediate endocytosis, but not phagocytosis.These data indicate that CLEC9A functions as an activation receptor.

View Article: PubMed Central - PubMed

Affiliation: Division of Immunology, Institute of Infectious Disease and Molecular Medicine, University of Cape Town, Observatory, 7925, South Africa.

ABSTRACT
We describe here the first characterization of CLEC9A, a group V C-type lectin-like receptor located in the "Dectin-1 cluster" of related receptors, which are encoded within the natural killer (NK)-gene complex. Expression of human CLEC9A is highly restricted in peripheral blood, being detected only on BDCA3(+) dendritic cells and on a small subset of CD14(+)CD16(-) monocytes. CLEC9A is expressed at the cell surface as a glycosylated dimer and can mediate endocytosis, but not phagocytosis. CLEC9A possesses a cytoplasmic immunoreceptor tyrosine-based activation-like motif that can recruit Syk kinase, and we demonstrate, using receptor chimeras, that this receptor can induce proinflammatory cytokine production. These data indicate that CLEC9A functions as an activation receptor.

Show MeSH
Related in: MedlinePlus