SOMATIC MUTATION AND SELECTION OF MEMORY B CELLS

描述（由申请人提供）：表达高亲和力的记忆B细胞 由体细胞突变的IG编码的表面免疫球蛋白（sIG）受体 可变（V）区基因在抗感染性疾病的保护中起主要作用。 也有助于自身免疫。尽管如此，超过三个 经过几十年的深入研究，我们对这一问题的理解存在许多空白。 记忆B细胞的细胞起源和亲和力依赖性选择， 产生体细胞突变的分子机制。这两种模式 累积的突变和双链DNA断裂的可能性是一个潜在的风险。 中间体的突变，导致了假设，易错 DNA修复可能是导致V区碱基替换的原因 DNA.由于其作为易错聚合酶的已知作用， zeta（polzeta）被认为是这一进程中的关键调解人。的小鼠 表达转基因编码的pol zeta RNA特异性反义RNA， 低水平的pol zeta RNA尽管在免疫后这些小鼠产生 强烈的GC反应，同种型转换抗体的正常滴度，以及正常的 大量的记忆B细胞，它们的抗体亲和力成熟缓慢， 抗体V区积累的体细胞突变比正常的少， 亲和力增强突变的减少。这是一份申请 进一步这些反义转基因小鼠，以精确地定义低表达的影响。 pol zeta水平对体细胞突变和亲和力成熟的影响 用多种抗原免疫。因为体内选择可以 极大地偏离了累积突变的数量和质量，细胞从 这些小鼠也将在克隆水平上进行体外免疫评价 反应性、耐受性、敏感性及其积累 突变。最后，前体细胞混合物的过继转移研究 从常规的、绿色荧光蛋白转基因的和反义p01 将进行zeta转基因小鼠，以评估 记忆B细胞和正常突变与 亚正常突变细胞在选择细胞进入记忆B细胞 池如果成功的话，这些实验将在以下方面取得重要进展： 了解负责B细胞的分子和细胞现象 记忆

DESCRIPTION (provided by applicant): Memory B cells that express high affinity surface immunoglobulin (sIg) receptors encoded by somatically mutated Ig variable (V) region genes play a major role in protection against infectious agents and contribute to autoimmunity as well. In spite of this, and over three decades of intensive investigation, many gaps exist in our understanding of the cellular origins and affinity dependent selection of memory B cells, and the molecular mechanisms that generate somatic mutations. Both the pattern of accumulated mutations and the possibility that double strand DNA breaks are an intermediate in somatic mutation, have led to the hypothesis that error-prone DNA repair may be responsible for introducing base substitutions in V region DNA. Because of its known role as an error-prone polymerase, DNA polymerase zeta (pol zeta) has been suggested as a key mediator in this process. Mice that express transgene encoded antisense RNA specific for pol zeta RNA display very low levels of pol zeta RNA. Although upon immunization these mice generate vigorous GC reactions, normal titers of isotype switched antibodies, and normal numbers of memory B cells, their antibodies affinity-mature slowly and their antibody V regions accumulate fewer than normal somatic mutations with a marked reduction in affinity enhancing mutations. This is a application to study further these antisense transgenic mice to define precisely the impact of low pol zeta levels on somatic mutation and affinity maturation with time after immunization with a variety of antigens. Because in vivo selection could greatly bias the quantity and quality of accumulated mutations, cells from these mice will also be evaluated at the clonal level in vitro for their immune responsiveness, tolerance susceptibility and their accumulation of somatic mutations. Finally, adoptive transfer studies of mixtures of precursor cells from conventional, green fluorescence protein transgenic, and antisense p01 zeta transgenic mice will be carried out to evaluate the cellular origins of memory B cells and any competitive advantage of normally mutating vs. sub-normally mutating cells in the selection of cells into the memory B cell pool. If successful, these experiments will provide important advances in understanding the molecular and cellular phenomena responsible for B cell memory.