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Identification and molecular characterization of NKp30, a novel triggering receptor involved in natural cytotoxicity mediated by human natural killer cells.

Pende D, Parolini S, Pessino A, Sivori S, Augugliaro R, Morelli L, Marcenaro E, Accame L, Malaspina A, Biassoni R, Bottino C, Moretta L, Moretta A - J. Exp. Med. (1999)

Bottom Line: This novel receptor is associated with CD3zeta chains that become tyrosine phosphorylated upon sodium pervanadate treatment of NK cells.Molecular cloning of NKp30 cDNA revealed a member of the immunoglobulin superfamily, characterized by a single V-type domain and a charged residue in the transmembrane portion.Moreover, we show that NKp30 is encoded by the previously identified 1C7 gene, for which the function and the cellular distribution of the putative product were not identified in previous studies.

View Article: PubMed Central - PubMed

Affiliation: Istituto Nazionale per la Ricerca sul Cancro, 16132 Genova, Italy.

ABSTRACT
Two major receptors involved in human natural cytotoxicity, NKp46 and NKp44, have recently been identified. However, experimental evidence suggested the existence of additional such receptor(s). In this study, by the generation of monoclonal antibodies (mAbs), we identified NKp30, a novel 30-kD triggering receptor selectively expressed by all resting and activated human natural killer (NK) cells. Although mAb-mediated cross-linking of NKp30 induces strong NK cell activation, mAb-mediated masking inhibits the NK cytotoxicity against normal or tumor target cells. NKp30 cooperates with NKp46 and/or NKp44 in the induction of NK-mediated cytotoxicity against the majority of target cells, whereas it represents the major triggering receptor in the killing of certain tumors. This novel receptor is associated with CD3zeta chains that become tyrosine phosphorylated upon sodium pervanadate treatment of NK cells. Molecular cloning of NKp30 cDNA revealed a member of the immunoglobulin superfamily, characterized by a single V-type domain and a charged residue in the transmembrane portion. Moreover, we show that NKp30 is encoded by the previously identified 1C7 gene, for which the function and the cellular distribution of the putative product were not identified in previous studies.

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NKp30 cooperates with NKp46 and NKp44 in the induction of NK-mediated cytotoxicity against tumor or normal autologous target cells. (A) The representative NK clone MIL69 was analyzed for cytolytic activity against FO-1 or A549 FcγR-negative target cell lines either in the absence or presence of mAbs to the indicated molecules. The following mAbs were used: KL247 (anti-NKp46), AZ20 (anti-NKp30), and KS38 (anti-NKp44). The E/T ratios were 2:1 (FO-1) and 3:1 (A549). (B) Two NK cell clones (MX361 and P9) were analyzed for cytolytic activity against autologous PHA blasts either in the absence (white bars) or presence of mAbs to the indicated molecules (black bars). The mAbs used were A6-136 (anti–HLA class I [cl I]), KL247 (anti-NKp46), KS38 (anti-NKp44), and AZ20 (anti-NKp30). The E/T ratio was 10:1.
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Figure 6: NKp30 cooperates with NKp46 and NKp44 in the induction of NK-mediated cytotoxicity against tumor or normal autologous target cells. (A) The representative NK clone MIL69 was analyzed for cytolytic activity against FO-1 or A549 FcγR-negative target cell lines either in the absence or presence of mAbs to the indicated molecules. The following mAbs were used: KL247 (anti-NKp46), AZ20 (anti-NKp30), and KS38 (anti-NKp44). The E/T ratios were 2:1 (FO-1) and 3:1 (A549). (B) Two NK cell clones (MX361 and P9) were analyzed for cytolytic activity against autologous PHA blasts either in the absence (white bars) or presence of mAbs to the indicated molecules (black bars). The mAbs used were A6-136 (anti–HLA class I [cl I]), KL247 (anti-NKp46), KS38 (anti-NKp44), and AZ20 (anti-NKp30). The E/T ratio was 10:1.

Mentions: Analysis of the same NK clones in cytolytic assays against other tumor target cells such as SMMC and A549 (Fig. 5) revealed a balanced contribution of NKp46 and NKp30 to the induction of cytotoxicity. Indeed, while mAb-mediated masking of NKp46 or NKp30 alone had a moderate inhibitory effect, the simultaneous masking of the two molecules resulted in a significant inhibition. These results indicate that the two receptors may exert an additive effect in the induction of cytotoxicity against certain target cells. Cooperation in NK cell triggering was previously demonstrated for NKp46 and NKp44 23. Further analysis revealed that NKp30 could exert an additive effect in the induction of NK-mediated cytotoxicity, not only with NKp46, but also with NKp44. Fig. 6 A shows the cytolytic activity of the representative NK clone MIL69 against FO-1 or A549 tumor cells. Target cell lysis was only partially inhibited by mAb-mediated masking of NKp30, NKp44, or NKp46 receptors. However, the combined masking of two receptors resulted in a higher inhibitory effect, whereas the simultaneous masking of the three receptors gave the maximal inhibition. Isotype-matched anti-CD56 mAb had no inhibitory effect either when used alone or in combination with other mAbs (data not shown).


Identification and molecular characterization of NKp30, a novel triggering receptor involved in natural cytotoxicity mediated by human natural killer cells.

Pende D, Parolini S, Pessino A, Sivori S, Augugliaro R, Morelli L, Marcenaro E, Accame L, Malaspina A, Biassoni R, Bottino C, Moretta L, Moretta A - J. Exp. Med. (1999)

NKp30 cooperates with NKp46 and NKp44 in the induction of NK-mediated cytotoxicity against tumor or normal autologous target cells. (A) The representative NK clone MIL69 was analyzed for cytolytic activity against FO-1 or A549 FcγR-negative target cell lines either in the absence or presence of mAbs to the indicated molecules. The following mAbs were used: KL247 (anti-NKp46), AZ20 (anti-NKp30), and KS38 (anti-NKp44). The E/T ratios were 2:1 (FO-1) and 3:1 (A549). (B) Two NK cell clones (MX361 and P9) were analyzed for cytolytic activity against autologous PHA blasts either in the absence (white bars) or presence of mAbs to the indicated molecules (black bars). The mAbs used were A6-136 (anti–HLA class I [cl I]), KL247 (anti-NKp46), KS38 (anti-NKp44), and AZ20 (anti-NKp30). The E/T ratio was 10:1.
© Copyright Policy
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC2195691&req=5

Figure 6: NKp30 cooperates with NKp46 and NKp44 in the induction of NK-mediated cytotoxicity against tumor or normal autologous target cells. (A) The representative NK clone MIL69 was analyzed for cytolytic activity against FO-1 or A549 FcγR-negative target cell lines either in the absence or presence of mAbs to the indicated molecules. The following mAbs were used: KL247 (anti-NKp46), AZ20 (anti-NKp30), and KS38 (anti-NKp44). The E/T ratios were 2:1 (FO-1) and 3:1 (A549). (B) Two NK cell clones (MX361 and P9) were analyzed for cytolytic activity against autologous PHA blasts either in the absence (white bars) or presence of mAbs to the indicated molecules (black bars). The mAbs used were A6-136 (anti–HLA class I [cl I]), KL247 (anti-NKp46), KS38 (anti-NKp44), and AZ20 (anti-NKp30). The E/T ratio was 10:1.
Mentions: Analysis of the same NK clones in cytolytic assays against other tumor target cells such as SMMC and A549 (Fig. 5) revealed a balanced contribution of NKp46 and NKp30 to the induction of cytotoxicity. Indeed, while mAb-mediated masking of NKp46 or NKp30 alone had a moderate inhibitory effect, the simultaneous masking of the two molecules resulted in a significant inhibition. These results indicate that the two receptors may exert an additive effect in the induction of cytotoxicity against certain target cells. Cooperation in NK cell triggering was previously demonstrated for NKp46 and NKp44 23. Further analysis revealed that NKp30 could exert an additive effect in the induction of NK-mediated cytotoxicity, not only with NKp46, but also with NKp44. Fig. 6 A shows the cytolytic activity of the representative NK clone MIL69 against FO-1 or A549 tumor cells. Target cell lysis was only partially inhibited by mAb-mediated masking of NKp30, NKp44, or NKp46 receptors. However, the combined masking of two receptors resulted in a higher inhibitory effect, whereas the simultaneous masking of the three receptors gave the maximal inhibition. Isotype-matched anti-CD56 mAb had no inhibitory effect either when used alone or in combination with other mAbs (data not shown).

Bottom Line: This novel receptor is associated with CD3zeta chains that become tyrosine phosphorylated upon sodium pervanadate treatment of NK cells.Molecular cloning of NKp30 cDNA revealed a member of the immunoglobulin superfamily, characterized by a single V-type domain and a charged residue in the transmembrane portion.Moreover, we show that NKp30 is encoded by the previously identified 1C7 gene, for which the function and the cellular distribution of the putative product were not identified in previous studies.

View Article: PubMed Central - PubMed

Affiliation: Istituto Nazionale per la Ricerca sul Cancro, 16132 Genova, Italy.

ABSTRACT
Two major receptors involved in human natural cytotoxicity, NKp46 and NKp44, have recently been identified. However, experimental evidence suggested the existence of additional such receptor(s). In this study, by the generation of monoclonal antibodies (mAbs), we identified NKp30, a novel 30-kD triggering receptor selectively expressed by all resting and activated human natural killer (NK) cells. Although mAb-mediated cross-linking of NKp30 induces strong NK cell activation, mAb-mediated masking inhibits the NK cytotoxicity against normal or tumor target cells. NKp30 cooperates with NKp46 and/or NKp44 in the induction of NK-mediated cytotoxicity against the majority of target cells, whereas it represents the major triggering receptor in the killing of certain tumors. This novel receptor is associated with CD3zeta chains that become tyrosine phosphorylated upon sodium pervanadate treatment of NK cells. Molecular cloning of NKp30 cDNA revealed a member of the immunoglobulin superfamily, characterized by a single V-type domain and a charged residue in the transmembrane portion. Moreover, we show that NKp30 is encoded by the previously identified 1C7 gene, for which the function and the cellular distribution of the putative product were not identified in previous studies.

Show MeSH
Related in: MedlinePlus