Moretta A, Vitale M, Sivori S, Bottino C, Morelli L, Augugliaro R, Barbaresi M, Pende D, Ciccone E, Lopez-Botet M, Moretta L
Istituto di Istologia ed Embriologia Generale, Università di Genova, Italy.
J Exp Med. 1994 Aug 1;180(2):545-55. doi: 10.1084/jem.180.2.545.
GL183 or EB6 (p58) molecules have been shown to function as receptors for different HLA-C alleles and to deliver an inhibitory signal to natural killer (NK) cells, thus preventing lysis of target cells. In this study, we analyzed a subset of NK cells characterized by a p58-negative surface phenotype. We show that p58-negative clones, although specific for class I molecules do not recognize HLA-C alleles. In addition, by the use of appropriate target cells transfected with different HLA-class I alleles we identified HLA-B7 as the protective element recognized by a fraction of p58-negative clones. In an attempt to identify the receptor molecules expressed by HLA-B7-specific clones, monoclonal antibodies (mAbs) were selected after mice immunization with such clones. Two of these mAbs, termed XA-88 and XA-185, and their F(ab')2 fragments, were found to reconstitute lysis of B7+ target cells by B7-specific NK clones. Both mAbs were shown to be directed against the recently clustered Kp43 molecule (CD94). Thus, mAb-mediated masking of Kp43 molecules interferes with recognition of HLA-B7 and results in target cell lysis. Moreover, in a redirected killing assay, the cross-linking of Kp43 molecules mediated by the XA185 mAb strongly inhibited the cytolytic activity of HLA-B7-specific NK clones, thus mimicking the functional effect of B7 molecules. Taken together, these data strongly suggest that Kp43 molecules function as receptors for HLA-B7 and that this receptor/ligand interaction results in inhibition of the NK-mediated cytolytic activity. Indirect immunofluorescence and FACS analysis of a large number of random NK clones showed that Kp43 molecules (a) were brightly expressed on a subset of p58-negative clones, corresponding to those specific for HLA-B7; (b) displayed a medium/low fluorescence in the p58-negative clones that are not B7-specific as well as in most p58+ NK clones; and (c) were brightly expressed as in the p58+ clone ET34 (GL183-/EB6+, Cw4-specific). Functional analysis revealed that Kp43 functioned as an inhibitory receptor only in NK clones displaying bright fluorescence. These studies also indicate that some NK clones (e.g., the ET34) can coexpress two distinct receptors (p58 and Kp43) for different class I alleles (Cw4 and B7). Finally, we show that Kp43 molecules function as receptors only for some HLA-B alleles and that still undefined receptor(s) must exist for other HLA-B alleles including B27.
GL183或EB6（p58）分子已被证明可作为不同HLA - C等位基因的受体，并向自然杀伤（NK）细胞传递抑制信号，从而防止靶细胞裂解。在本研究中，我们分析了一组具有p58阴性表面表型的NK细胞亚群。我们发现，p58阴性克隆虽然对I类分子具有特异性，但不识别HLA - C等位基因。此外，通过使用转染了不同HLA - I类等位基因的合适靶细胞，我们确定HLA - B7是一部分p58阴性克隆所识别的保护元件。为了鉴定由HLA - B7特异性克隆表达的受体分子，在用此类克隆免疫小鼠后选择了单克隆抗体（mAb）。发现其中两种单克隆抗体，称为XA - 88和XA - 185，及其F（ab'）2片段，可重建B7特异性NK克隆对B7 +靶细胞的裂解作用。这两种单克隆抗体均显示针对最近聚类的Kp43分子（CD94）。因此，mAb介导的Kp43分子掩盖会干扰对HLA - B7的识别并导致靶细胞裂解。此外，在重定向杀伤试验中，XA185单克隆抗体介导的Kp43分子交联强烈抑制了HLA - B7特异性NK克隆的细胞溶解活性，从而模拟了B7分子的功能效应。综上所述这些数据强烈表明，Kp43分子作为HLA - B7的受体，并且这种受体/配体相互作用导致NK介导的细胞溶解活性受到抑制。对大量随机NK克隆进行间接免疫荧光和FACS分析表明，Kp43分子：（a）在一部分p58阴性克隆上高表达，这些克隆对应于对HLA - B7具有特异性的克隆；（b）在非B7特异性的p58阴性克隆以及大多数p58 + NK克隆中显示中/低荧光；（c）与p58 +克隆ET34（GL183 - /EB6 +，Cw4特异性）一样高表达。功能分析表明，Kp43仅在显示高荧光的NK克隆中作为抑制性受体起作用。这些研究还表明，一些NK克隆（例如ET34）可以共表达针对不同I类等位基因（Cw4和B7）的两种不同受体（p58和Kp43）。最后，我们表明Kp43分子仅作为某些HLA - B等位基因的受体，而对于包括B27在内的其他HLA - B等位基因，必定存在尚未确定的受体。
