摘要:ZNRF1在Toll-like receptor訊息傳導途徑的角色由失控的發炎反應所引起的敗血症是現今在加護病房中造成死亡的主因,但除了抗生素外目前仍無更有效的治療藥物。近期報導指出,有少數超級病菌已對幾乎所有種類的抗生素產生抗藥性。因此尋找新藥與改善藥物治療效果是相當迫切需要的。在微生物感染的過程中,先天性免疫細胞利用關聯分子型態受器(主要是Toll-like receptors, TLRs)辨識來自病原體上具高度保留性的病原關聯分子形態,進而啟動一系列的防禦機制來清除病菌,並且亦控制了適應性免疫系統的啟動。髓樣細胞包括巨噬細胞則在病原關聯分子形態的辨識與啟動和放大免疫反應的過程中扮演了中心角色。 TLR的訊息傳遞路徑主要由以下四種機制來調控: 胞內位置分佈,磷酸化作用,泛素化作用/去泛素化作用以及分子間競爭。近期的研究指出,在TLR4活化之後,其下游調控促發炎細胞激素生產的MyD88複合體,以及參與第一型干擾素表現的TRIF複合體,此兩者的活化是發生在不同的細胞區域。 然而,對於TLR4的胞吞作用與其在胞內的胞膜傳遞機制,目前仍尚不清楚。 研究報告指出,坐落於細胞中內含體/溶酶體的RING finger E3 泛素酵素稱作ZNRF1,已知參與在細胞分泌/胞吞作用的機制中。我們的研究發現,ZNRF1不僅在經過LPS(TLR4的配體)刺激的巨噬細胞中迅速的被誘發表現,亦對於TLR4下游mitogen activated protein kinases (MAPKs) 與IKK的活化相當的重要。根據我們初步的實驗結果,我們認為ZNRF1可能經由調控TLR4在胞內的胞膜傳遞來影響TLR4的訊息傳遞。這項研究將會探討ZNRF1在TLRs訊息傳遞與宿主免疫系統的角色。這項研究計劃包含了以下三個特定的實驗目標。1. 研究ZNRF1對於TLRs訊息傳遞路徑的影響。2. 找出ZNRF1調控TLRs訊息傳遞路徑的分子機制。3. 瞭解ZNRF1在宿主防禦機制(host defense)的生理功能。這項研究將對TLR訊息傳遞路徑提出新的見解。並在敗血症、免疫性與感染性疾病的瞭解與認知,以及治療策略的改善方面,都有相當大的助益。
Abstract: Characterization of the role of ZNRF1 in the Toll-like receptor signaling pathwaysSepsis caused by uncontrolled inflammation is the leading cause of death in intensive care units today, but no effective therapeutic medication is available currently except for antibiotics. Most recently, the emergence of few superbugs has been reported to be resistant to almost all antibiotics. Thus, new and improved medications are urgent needed. During microbial infections, innate immune cells use pattern-recognition receptors (PRRs), mainly Toll-like receptors (TLRs), to detect conserved microbial molecules termed pathogen-associated molecular patterns (PAMPs), and then mount a defense response to clear bacteria and control the initiation and determination of the adaptive immune response. Myeloid cells, including macrophages, play central roles in PAMP recognition and in the initiation and amplification of inflammatory cascades. TLR signaling is mainly modulated by four mechanisms: cellular localization, phosphorylation, Ubiquitination/deubiquitination, and competition. Recent studies have shown that activation of MyD88 complex, resulting in induction of proinflammatory cytokine, and TRIF-complex involved in type I interferon expression occur in different cellular compartments upon TLR4 engagement. However, very little is known about regulation of TLR4 endocytosis and intracellular trafficking. ZNRF1, a RING finger E3 ubiquitin ligase, is located in endosome/lysosome compartment of cells. Research has also proposed that ZNRF1 is involved in the secretory/endocytic pathways. We found that the expression of ZNRF1 is quickly induced in LPS (TLR4 ligand)-induced macrophages. In addition, we demonstrated that ZNRF1 is crucial for TLR4-triggered activation of mitogen activated protein kinases (MAPKs) and IKK. Our preliminary results suggest that ZNRF1 regulates TLR4 signaling possibly via modulation of TLR4 trafficking. This study will explore the functional roles of ZNRF1 in regulation of TLRs signaling and host defense.This proposal consists of the following three specific aims:AIM 1. To define the impact of ZNRF1 in TLRs signaling.AIM 2. To elucidate the molecular mechanism by which ZNRF1 regulates TLRs signaling pathways.AIM 3. To determine the physiological function of the ZNRF1 in host defense.This study will provide new insights into the regulation of TLR signaling pathways. The knowledge generated from this research should contribute to advancing the understanding and therapy of sepsis, inflammatory and infectious diseases.