Ubiquitylation and the Regulation of Immune Homeostasis

描述(申请人提供)：树突状细胞(DC)是一种先天免疫细胞，调节免疫动态平衡和抗原驱动的免疫反应。树突状细胞的功能受Toll样受体(TLRs)启动的信号的高度调控。因此，调节TLR信号通路的细胞内分子对调节宿主免疫反应至关重要。我们最近发现A20是一种泛素修饰酶，可以限制Toll样受体(TLR)诱导的信号。我们正在研究树突状细胞A20表达在调节免疫动态平衡和反应中的作用。为了做到这一点，我们培育了两种以基因为靶标的小鼠：一种是在所有细胞中破坏A20基因，另一种是A20的“开花”等位基因被靶向内源性A20基因。我们已经将CD11-Cre转基因小鼠品系与在DC中高选择性和特异性表达Cre酶的新品系进行了杂交。这些复合的A20FLOX/FLOX CD11c-Cre小鼠将为我们研究DCs中A20表达在调节免疫动态平衡和免疫应答中的作用提供一个非常强大的新试剂。我们建议使用这些A20FLOX/FLOX CD11-cre复合小鼠，以及来自这两个品系小鼠的细胞，在体外和体内确定A20在DC中的表达如何调节它们对TLR配体的反应。第一个目标将集中于了解A20‘S在调节DC中TLR信号转导中的细胞生物学和生化作用，并将阐明A20调控的DC的细胞自主功能。第二个目标将利用A20FLOX/FLOX CD11c-CRE小鼠来确定A20在DC中的表达如何调节其体内的动态平衡和激活。第二个目标还将询问树突状细胞中A20的表达如何调节外周耐受和免疫反应。

DESCRIPTION (provided by applicant): Dendritic cells (DCs) are innate immune cells that regulate both immune homeostasis as well as antigen driven immune responses. The functions of DCs are highly regulated by signals initiated by Toll-like receptors (TLRs). Consequently, intracellular molecules that regulate TLR signaling pathways are pivotal for regulating host immune responses. We have recently found that A20 is a ubiquitin modifying enzyme that restricts toll-like receptor (TLR) induced signals. We are studying the roles of A20 expression in DCs in regulating immune homeostasis and responses. To accomplish this, we have generated two strains of gene targeted mice: one that disrupts the A20 gene in all cells and one in which a "floxed" allele of A20 has been targeted to the endogenous A20 locus. We have interbred the "floxed" mice with a novel strain of CD11-Cre transgenic mouse line that expresses Cre enzyme with high selectivity and specificity in DCs. These compound A20flox/flox CD11c-Cre mice will thus provide us with a very powerful and novel reagent for studying the role of A20 expression in DCs in regulating immune homeostasis and immune responses. We propose to use these A20flox/flox CD11-cre compound mice, as well as cells derived from both strains of mice, to determine how A20 expression in DCs regulates their responses to TLR ligands in vitro and in vivo. The first aim will focus upon understanding the cell biology and biochemistry underlying A20's roles in regulating TLR signaling in DCs, and will elucidate the cell-autonomous functions of DCs that are regulated by A20. The second aim will utilize A20flox/flox CD11c-Cre mice to determine how A20 expression in DCs regulates their homeostasis and activation in vivo. This second aim will also interrogate how A20 expression in DCs regulates peripheral tolerance and immune responses.