Park L S, Friend D J, Schmierer A E, Dower S K, Namen A E
Immunex Corporation, Seattle, Washington 98101.
J Exp Med. 1990 Apr 1;171(4):1073-89. doi: 10.1084/jem.171.4.1073.
A murine cell line (IxN/2b) absolutely dependent upon exogenous IL-7 for continued growth has been obtained that expresses lymphoid precursor and class I MHC antigens and also contains a rearranged mu heavy chain. This cell line has been used to define the binding and structural characteristics of the murine IL-7 receptor using 125I-labeled recombinant murine IL-7. 125I-IL-7 binding to IxN/2b cell was rapid and saturable at both 4 degrees and 37 degrees C. Equilibrium binding studies produced curvilinear Scatchard plots at both temperatures with high and low affinity Ka values of approximately 1 x 10(10) M-1 and 4 x 10(8) M-1, respectively, and a total of 2,000-2,500 IL-7 binding sites expressed per cell. Experiments measuring inhibition of binding of 125I-IL-7 by unlabeled IL-7 also produced data consistent with the existence of two classes of IL-7 receptors. Evidence concerning the possible molecular nature of two classes of IL-7 receptors was provided by dissociation kinetics and affinity crosslinking experiments. The dissociation rate of 125I-IL-7 was markedly increased when measured in the presence of unlabeled IL-7 at both 37 degrees and 4 degrees C, which is diagnostic of a receptor population displaying negative cooperativity. Crosslinking studies showed that under both reducing and nonreducing conditions, the major crosslinked species observed corresponded to a receptor size of 75-79 kD while a less intense higher molecular mass crosslinked species was also seen which corresponded to a receptor size approximately twice as large (159-162 kD). Both types of experiments suggest that the IL-7 receptor may form noncovalently associated dimers in the membrane. The IL-7 receptor was expressed on pre-B cells, but not detected on several murine B cell lines or primary mature B cells. It was also expressed on murine thymocytes, some T lineage cell lines, and on bone marrow-derived macrophage. All cells binding 125I-IL-7 exhibited curvilinear Scatchard plots. No cytokines or growth factors tested were able to inhibit binding of 125I-IL-7 to its receptor. These results define the initial binding and structural characteristics, and the cellular distribution, of the murine IL-7 receptor.
已获得一种鼠细胞系（IxN/2b），其持续生长绝对依赖外源性白细胞介素-7（IL-7），该细胞系表达淋巴样前体和I类主要组织相容性复合体（MHC）抗原，并且还含有重排的μ重链。该细胞系已被用于利用125I标记的重组鼠IL-7来确定鼠IL-7受体的结合和结构特征。在4℃和37℃下，125I-IL-7与IxN/2b细胞的结合迅速且具有饱和性。平衡结合研究在两个温度下均产生了曲线型Scatchard图，高亲和力和低亲和力的解离常数（Ka）值分别约为1×10(10) M-1和4×10(8) M-1，每个细胞表达的IL-7结合位点总数为2000 - 25oo个。测量未标记的IL-7对125I-IL-7结合的抑制作用的实验也产生了与两类IL-7受体的存在相一致的数据。解离动力学和亲和交联实验提供了有关两类IL-7受体可能的分子性质的证据。在37℃和4℃下，当在未标记的IL-7存在下测量时，125I-IL-7的解离速率显著增加，这是显示负协同性的受体群体的特征。交联研究表明，在还原和非还原条件下，观察到的主要交联产物对应于75 - 79 kD的受体大小，同时也看到了强度较低的更高分子量的交联产物，其对应于大约两倍大的受体大小（159 - 162 kD）。这两类实验均表明IL-7受体可能在膜中形成非共价结合的二聚体。IL-7受体在前B细胞上表达，但在几种鼠B细胞系或原代成熟B细胞上未检测到。它也在鼠胸腺细胞、一些T系细胞系以及骨髓来源的巨噬细胞上表达。所有结合125I-IL-7的细胞均呈现曲线型Scatchard图。所测试的细胞因子或生长因子均不能抑制125I-IL-7与其受体的结合。这些结果确定了鼠IL-7受体的初始结合和结构特征以及细胞分布。
