Roles for Transcription in Antibody Diversity

免疫球蛋白转换mu区中的重复DNA序列形成含RNA的二级结构，并通过活化诱导的脱氨酶（AID）进行超突变。 为了研究DNA结构如何影响转录和超突变，我们将RNA聚合酶II分子的位置和跨越内含子增强子的5 kb区域的突变映射到恒定的mu基因。 对于RNA聚合酶II，通过离体刺激的B细胞中的核连续和染色质免疫沉淀测定来确定其分布。 RNA聚合酶被发现在高密度的DNA侧翼两侧的1 kb的重复序列，形成开关区的核心。 我们提出，改变DNA结构的开关区域暂停RNA聚合酶II。 我们目前正在研究二级结构和转录模式的可变区，使用敲入小鼠含有一个重排的可变基因的两个等位基因。

Repetitive DNA sequences in the immunoglobulin switch mu region form RNA-containing secondary structures and undergo hypermutation by activation-induced deaminase (AID). To examine how DNA structure affects transcription and hypermutation, we mapped the position of RNA polymerase II molecules and mutations across a 5 kb region spanning the intronic enhancer to the constant mu gene. For RNA polymerase II, the distribution was determined by nuclear run-on and chromatin immunoprecipitation assays in B cells stimulated ex vivo. RNA polymerases were found at a high density in DNA flanking both sides of a 1 kb repetitive sequence that forms the core of the switch region. We propose that altered DNA structure in the switch region pauses RNA polymerase II. We are currently examining secondary structure and transcription patterns in the variable region, using knock-in mice containing one rearranged variable gene on both alleles.