DNA SEQUENCES INVOLVED IN THE HEAVY CHAIN SWITCH

在抗原驱动的免疫球蛋白分化过程中- 产生细胞，从微米到伽马，埃，或重 可以进行链合成。这种蛋白质转换是一种 DNA缺失将V区从供体Cµ移动到C Gamma、C epsilon或Cα基因。此删除操作从开始到结束 在开关片段中，由重复序列组成的DNA区域 序列。我们建议从分子上克隆和测序 在六到十个重排的伽马3和 杂交瘤DNA中的伽马1基因。伽马3和伽马1 每个开关区包括特征的49个碱基对 简单重复的序列。我们之前的数据表明， Gamma 3交换机均匀分布在其49mer 一致的序列，但S伽马1开关是 定位于其49mer共识序列的一部分。 此外，伽马开关位置附近的序列表明， 单一酶可能介导所有四种酶的切割/宗教 老鼠体内的伽马基因。我们将使用我们的新序列数据 来检验这些暂定结论。我们 将确定交换机重组是否在交换机中发生 仅限重复序列，或者如果它可以出现在非49聚体中 S伽马1区的层序我们还将使用派生数据 从克隆的开关区的核苷酸序列来理解 这两个交换机上的交换站点中是否存在任何关联关系 单个免疫球蛋白表达细胞的染色体。 在同一组实验中，我们将检查 从简单的细胞到重新排列的开关区域的结构 交换模式(典型交换)和复杂交换 模式(非典型开关)。我们还将检查交换机 小鼠杂交瘤的区域内容用α- 表达骨髓瘤伴侣。我们会问两个脾细胞- 衍生的免疫球蛋白基因座倾向于重排到相同的开关区域，并且 重链开关中同型特异性的指征 重组事件。这样我们就可以理解DNA Switch的序列识别、特异性和遗传学 重组酶我们建议选择一种符合构成的细胞系 产生这些酶。

During antigen-driven differentiation of immunoglobulin- producing cells, a switch from mu to gamma, epsilon, or a heavy chain synthesis can occur. This protein switch is the result of a DNA deletion that moves a V region from a donor C mu to a C gamma, C epsilon, or C alpha gene. This deletion begins and ends in switch segments, DNA regions composed of tandemly repeated sequences. We propose to molecularly clone and sequence the switch recombination sites in six to ten rearranged gamma 3 and gamma 1 genes from hybridoma DNA. The gamma 3 and gamma 1 switch regions each include a characterisitic 49 base pair tandemly repeated sequence. Our previous data indicate that the gamma 3 switches are uniformly distributed throughout its 49mer consensus sequence, but that the S gamma 1 switches are localized to a part of its 49mers consensus sequence. Furthermore, sequences near gamma switch sites suggest that a single enzyme may mediate the cutting/religation for all four gamma genes in the mouse. We will use our new sequence data from gamma switch sites to test these tentative conclusions. We will determine if switch recombination occurs in the tandemly repeated sequences only, or if it can occur in non-49mer sequences in the S gamma 1 region. We will also use data derived from nucleotide sequences of cloned switch regions to understand any correlative relationships in switch sites on the two chromosomes from a single immunoglobulin expressing cell. Under the same group of experiments, we will examine the structure of rearranged switch regions from cells with simple switching patterns (typical switches) and complex switching patterns (atypical switches). We will also examine the switch region content of murine hybridomas generated using an alpha- expressing myeloma partner. We will ask if the two spleen cell- derived Igh loci tend to rearrange to the same switch region, and indication of isotype specificity in the heavy chain switch recombinational event. So that we might understand the DNA sequence recognition, specificity, and genetics of switch recombinases we propose to select a cell line that constitutively produces these enzymes.