REGULATION OF IMMUNOGLOBULIN GENE EXPRESSION

当我们将克隆的kappa免疫球蛋白基因注射到非洲爪哇卵母细胞中时， 没有观察到kappa启动子启动的特异性转录。 当我们联合注射产生抗体的细胞核或核提取物时 骨髓瘤细胞与克隆的基因一起，高水平转录自 出现了kappa启动子。因此，细胞类型的特定因素 激活免疫球蛋白在淋巴细胞中的转录必须能够 在卵母细胞中发挥作用，为这些因子提供了体内检测方法。我们希望 用这种卵母细胞互补试验纯化细胞起始 免疫球蛋白基因转录因子。在中进行实验 使用合适的质粒底物的活体系统，以及后来的DNA结合 对纯化的激活因子的研究，将使我们能够确定 这些因素是作用于kappa启动子本身还是作用于下游 增强元素或两者兼而有之。 B淋巴细胞型细胞可以产生膜结合或分泌的 一种IgM抗体形式，具有从第一种抗体切换到第二种抗体的程序 在细胞发育过程中形成。膜的重链和 分泌形式是由不同的mRNA转录而来的 基因，两个mRNAs利用不同的聚腺苷酸化(Poly A) 网站。我们希望分离和研究导致这种疾病的细胞因素。 在发育过程中从膜结合抗体转变为分泌抗体， 显然是通过确定使用哪个聚A信号来实现的。为此， 来自不同发育阶段的淋巴样细胞的核将是 与克隆的Mu重链基因一起注射到卵母细胞中。如果 可以使细胞核改变转录本的多聚腺苷模式。 通过基因延伸，我们将使用这种方法来纯化因子 参与选择Poly A站点。

When we injected a cloned kappa immunoglobulin gene into Xenopus oocytes, no specific transcription initiating at the kappa promoter was observed. When we coinjected nuclei or nuclear extracts from antibody-producing myeloma cells together with the cloned gene, high level transcription from the kappa promoter occurred. Hence, the cell-type specific factors that activate immunoglobulin transcription in lymphoid cells must be able to function in oocytes, providing an in vivo assay for these factors. We wish to use this oocyte complementation assay to purify the cellular initiation factors for transcription of immunoglobulin genes. Experiments with the in vivo system using suitable plasmid substrates, and later DNA binding studies with the purified activating factors, will allow us to determine whether the factors act on the kappa promotor itself or on the downstream enhancing element or both. Cells of B-lymphocyte type can produce either a membrane-bound or secreted form of IgM antibody, with a program to switch from the first to the second form during cellular development. The heavy chains of the membrane and secreted form are synthesized from different mRNAs transcribed off the same gene, with the two mRNAs utilizing different polyadenylation (poly A) sites. We wish to isolate and study the cellular factors that cause the switch from membrane-bound to secreted antibody during development, evidently by determining which poly A signal is used. For this purpose, nuclei from lymphoid cells at various stages of development will be injected into oocytes together with a cloned Mu heavy chain gene. If the nuclei can be made to alter the polyadenylation pattern of transcripts extending through the gene, we will use this assay to purify the factors involved in selecting the poly A site.