Mold C, Nemerow G R, Bradt B M, Cooper N R
University of New Mexico, Department of Medicine, Albuquerque 87131.
J Immunol. 1988 Mar 15;140(6):1923-9.
Antibody-independent activation of the alternative C pathway by human lymphoblastoid cell lines latently infected with EBV has been recognized for some time, although the mechanisms involved and the specific cell surface molecule(s) recognized by the C system have not been identified. The present studies, carried out with the purified proteins of the alternative pathway have addressed these questions. Activation of the purified proteins of the alternative pathway by Raji lymphoblastoid cells was found to be antibody independent, confirming earlier findings with serum. Surprisingly, activation was highly dependent on properdin. In other models properdin has been found to augment alternative pathway activation and to be required for lysis of virus infected cells. Molecules which activate the alternative pathway provide binding sites on which C3 breakdown by regulatory proteins is impeded; therefore intact C3b accumulates on the activator. Immunoprecipitation studies with either anti-CR2 or anti-C3 have identified CR2, the R for C3d,g and EBV, as a major covalent and noncovalent binding site for C3 deposition on Raji cells during alternative pathway activation. Covalently bound C3b was dissociated from CR2 by hydroxylamine, indicating attachment via an ester bond. C3b binding after activation was not reduced by an anti-CR2 mAb which blocks CR2 R function, indicating that it was probably not mediated by C3d,g R epitopes on CR2. Direct confirmation of the ability of CR2 to trigger the alternative pathway came from studies with purified CR2 which was found to activate the alternative C pathway in serum or in mixtures of the purified proteins of the pathway. This work provides conclusive evidence that CR2 is a C activator and functions in this capacity on Raji cells.
一段时间以来，人们已经认识到，被EBV潜伏感染的人淋巴母细胞系可通过抗体非依赖性方式激活补体替代途径（C途径），尽管其中涉及的机制以及补体系统识别的特定细胞表面分子尚未明确。目前使用替代途径的纯化蛋白开展的研究解决了这些问题。发现Raji淋巴母细胞系对替代途径纯化蛋白的激活是抗体非依赖性的，这证实了早期血清研究的结果。令人惊讶的是，激活高度依赖备解素。在其他模型中，已发现备解素可增强替代途径的激活，并且是病毒感染细胞裂解所必需的。激活替代途径的分子提供了结合位点，在这些位点上调节蛋白对C3的分解受到阻碍；因此完整的C3b在激活剂上积累。用抗CR2或抗C3进行的免疫沉淀研究已确定CR2（C3d,g和EBV的受体）是替代途径激活过程中C3在Raji细胞上沉积的主要共价和非共价结合位点。通过羟胺可使共价结合的C3b从CR2上解离，表明其通过酯键连接。激活后C3b的结合并未因阻断CR2受体功能的抗CR2单克隆抗体而减少，这表明其可能不是由CR2上的C3d,g受体表位介导的。对CR2触发替代途径能力的直接证实来自对纯化CR2的研究，发现其可在血清或该途径纯化蛋白混合物中激活补体替代途径。这项工作提供了确凿证据，证明CR2是一种补体激活剂，并在Raji细胞上发挥这种作用。
