Evaluating the Role of Senataxin in Class Switch Recombination: A Model for Biological Diversity

研究计划摘要 免疫球蛋白（IG）基因座是生物系统的一个迷人的例子，它已经进化出多种策略来实现多样性。这个基因座的进化及其调节可以教会我们很多关于多样性是如何产生和生物学调节的。因此，我们的研究计划是由三个主要问题：1）什么因素已经演变成后生动物，以服务专门的角色，在调节IG多样性？2)不同物种间观察到的IG基因独特多样性策略的基础是什么？3)IG多样性在体细胞水平上是如何调节的？从长远来看，这些研究将提供重要的新的进化洞察多样性是如何实现和调节的IG基因座和这种多样性对多生物体关系的后果（共生与寄生与致病/寄生）。 提案摘要 类别转换重组（CSR）是B细胞用于产生各种同种型抗体的过程。抗体类别（或同种型）由重链恒定区（CH）决定。CSR由导致染色体内缺失的重组过程介导。重组事件发生在位于每个CH基因座上游的开关（S）区域之间，并且需要多轮细胞分裂。这一过程由一组复杂的酶协调，包括：活化诱导的胞苷脱氨酶（AID）和参与DNA损伤反应的蛋白质。 S区的转录是CSR所必需的，并导致形成RNA-DNA杂合体（R环）。Senataxin（SETX）是一种RNA-DNA解旋酶，已被证明可以促进R环的分解。它在DNA损伤反应中也起着重要作用。 我们最近已经证明，SETX在调节对病毒感染的先天免疫应答中起着至关重要的作用。初步研究还表明，SETX缺陷型小鼠在对疫苗接种/感染的反应中经历缺陷型CSR。使用SETX-/-小鼠模型，我们将描述SETX在CSR过程中所起的作用，以更深入地了解转录，DNA复制和重组的过程是如何在免疫球蛋白位点同时调节的。 假设 我们推测，SETX在调节CSR中起着重要的作用，SETX缺陷将损害类转换。 意义 这些研究将提供关于IG基因座如何利用一般细胞因子来执行对多样性的产生和调节至关重要的特定功能的重要的新的进化见解。从长远来看，了解管理多样性的产生和调节的基本机制可能在生物工程和合成生物学中有广泛的应用，其中生物多样性的创造和调节是相当重要的。

Summary of Research Program The Immunoglobulin (Ig) locus is a fascinating example of a biological system that has evolved multiple strategies to achieve diversity. The evolution of this locus and its regulation can teach us a great deal about how diversity is generated and regulated biologically. Our research program is thus guided by three major questions: 1) What factors have evolved in metazoans to serve specialized roles in regulating Ig diversity? 2) What is the basis for the unique diversification strategies of Ig genes observed across species? 3) How is Ig diversity regulated at the somatic level? In the long term, these studies will provide important new evolutionary insight into how diversity is achieved and regulated at the Ig locus and the consequences of this diversity on multi-organism relationships (symbiotic vs. commensal vs. pathogenic/parasitic). Summary of Proposal Class switch recombination (CSR) is the process used by B cells to generate antibodies of various isotypes. Antibody class (or isotype) is determined by the heavy chain constant region (CH). CSR is mediated by a process of recombination that leads to intrachromosomal deletion. The recombination event occurs between switch (S) regions located upstream of each CH locus, and requires multiple rounds of cellular division. This process is coordinated by a complex set of enzymes including: activation-induced cytidine deaminase (AID), and proteins involved in the DNA damage response. Transcription of S regions is required for CSR and results in the formation RNA-DNA hybrids (R-loops). Senataxin (SETX) is an RNA-DNA helicase that has been shown to promote the resolution of R-loops. It also plays an important role in the DNA damage response. We have recently demonstrated that SETX plays an essential role in regulating the innate immune response to viral infection. Preliminary studies have also suggested that SETX-deficient mice undergo defective CSR in response to vaccination/infection. Using the SETX-/- mouse model we will characterize the role played by SETX during CSR in order to gain a deeper understanding of how the processes of transcription, DNA replication and recombination are concurrently regulated at the immunoglobulin locus. Hypothesis We hypothesize that SETX plays an important role in regulating CSR and that SETX deficiency will impair class switching. Significance These studies will provide important new evolutionary insight regarding how the Ig locus has taken advantage of general cellular factors to perform specific functions that are critical for the generation and regulation of diversity. In the long term, understanding the basic mechanisms governing the generation and regulation of diversity is likely to have broad applications in bioengineering and synthetic biology, wherein creation and regulation of biological diversity is of considerable importance.