Regulation of Innate Immunity by F-Box Proteins

描述(申请人提供)：重症肺炎或败血症引起的肺损伤患者的一个临床特征是由入侵的病原体引发的强烈的急性炎症宿主反应。在这里，在入侵病原体激活免疫效应细胞(T细胞、巨噬细胞等)上的受体后，天然免疫系统被激活以分泌大量促炎细胞因子(即细胞因子风暴)。TRAF家族的接头分子是连接外部表面信号(如通过Toll受体)和细胞因子释放的关键和高度保守的蛋白质，通过依赖于核因子-kB的信号。因此，设计有选择性地调节TRAF家族蛋白的可用性的操作可能会减轻脓毒症的严重程度。然而，到目前为止，人们对TRAF家族在蛋白质稳定性水平上的分子调控知之甚少。在研究细菌败血症的过程中，我们发现一个称为Fbxo3的孤儿泛素E3连接酶亚基被激活，并足以泛素化和介导另一个相对较新的泛素E3连接酶亚基Fbxl2的蛋白酶体降解。此外，我们发现Fbxl2具有抗炎作用，因为它以TRAF家族蛋白为靶点，使其在上皮和单核细胞中处置。因此，我们的初步工作表明，铜绿假单胞菌通过激活Fbxo3，导致Fbxl2泛素化和降解，导致免疫反应TRAF增加，细胞因子产生增加，并损害肺的稳定性。这些数据导致了我们的新假设，即铜绿假单胞菌诱导的细胞因子释放涉及Fbxo3介导的泛素化和Fbxl2的降解，这反过来又增加了促炎症的TRAF蛋白的水平。我们将确定孤儿泛素E3连接酶亚单位Fbxo3是否与GSK3b协调，触发Fbxl2泛素化依赖的降解，进而激活TRAF蛋白以刺激细胞因子释放(目标1)。我们还将确定Fbxo3的突变或抑制是否会减轻细菌诱导的急性肺损伤(目标2)。这一建议提供了一种新的先天免疫模式，因为它与细胞因子信号转导有关。这些研究的实施将为在严重细菌感染期间先天性免疫反应的分子调控方面的重大机制进展奠定基础。这些研究的结果旨在作为战略的基础，旨在开发高选择性的新型Fbxo3小分子抑制剂，以减轻细菌或败血症诱导的急性肺损伤的严重程度。

DESCRIPTION (provided by applicant): One clinical hallmark of patients with severe pneumonia or sepsis-induced lung injury is a robust acute inflammatory host response triggered by the invading pathogens. Here, the innate immunity system is activated to secrete large amounts of pro-inflammatory cytokines (i.e. a cytokine storm) after invading pathogens activate receptors on immune effector cells (T-cells, macrophages, etc.). The TRAF family of adaptors molecules are pivotal and highly conserved proteins that link external surface signals (e.g. via Toll receptors) to cytokine release via NF-kB dependent signaling. Thus, maneuvers designed to selectively modulate the availability of TRAF family proteins might lessen the severity of sepsis. However, to date, very little is known regarding the molecular regulation of the TRAF family at the level of protein stability. In the process of studying bacterial sepsis, we discovere that an orphan ubiquitin E3 ligase subunit, termed FBXO3, is activated and sufficient to ubiquitinate and mediate proteasomal degradation of another relatively new ubiquitin E3 ligase subunit, termed FBXL2. Further, we discovered that FBXL2 is anti-inflammatory as it targets the TRAF family of proteins for their disposal in epithelia and monocytes. Thus, our preliminary work demonstrates that P. aeruginosa, via activation of FBXO3, results in FBXL2 ubiquitination and degradation resulting in increased immunoreactive TRAFs, increased cytokine production, and impaired lung stability. These data led to our novel hypothesis that P. aeruginosa induced cytokine release involves FBXO3 mediated ubiquitination and degradation of FBXL2, which in turn increases levels of TRAF proteins which are pro-inflammatory. We will determine if an orphan ubiquitin E3 ligase subunit, FBXO3, coordinates with GSK3b to trigger ubiquitination dependent degradation of FBXL2, which in turn activates TRAF proteins to stimulate cytokine release (Aim 1). We will also determine if mutation or inhibition of FBXO3 will attenuate bacterial-induced acute lung injury (Aim 2). This proposal provides a new model of innate immunity as it relates to cytokine signaling. Execution of these studies will lay the foundation fo a significant mechanistic advance regarding the molecular regulation of the innate immune response during severe bacterial infection. Results from these studies are intended to serve as the basis for strategies directed at the development of highly selective novel small molecule inhibitors of FBXO3 to lessen the severity of bacterial or sepsis-induced acute lung injury.