Zcchc6在先天免疫反應中的角色

Abstract

先天性免疫系統是身體防禦的第一道防線。當人體被感染時，它提供即時的保護功能，進而有利於病原體的清除和調節適應性免疫系統。Toll-like Receptors(TLRs) 是先天性免疫系統的重要組成之一。在遭受微生物感染時，它們扮演著重要的角色，會去辨識pathogen associated molecular patterns，然後啟動下游的發炎和抗病毒反應。然而，炎症反應失調與多種疾病息息相關，如癌症、代謝性疾病和發炎性腸道疾病。因此，TLR所媒介的免疫反應必須受到嚴密的調控。在先前的研究中，我們的實驗室發現了”Zcchc6”，它屬於Terminal Uridylyl Transferase (TUTase)的成員，其表現量可在小鼠巨噬細胞中被LPS誘導提升。TUTase已知是調控RNA穩定性的酵素，其可藉由對 RNA受體末端加U，進而促進RNA 穩定或降解。我們的研究結果顯示，在Zcchc6-缺失的RAW264.7 巨噬細胞中， TLRs ligands刺激會導致許多細胞激素的失調。IL-10 是其中受影響的細胞激素之一，它是一種抗發炎細胞激素，且參與許多免疫調節功能。在TLRs活化4小時後， Zcchc6 缺失的 RAW264.7 巨噬細胞會有較高的IL-10 表現量。在這項研究中，我們進一步的探討了Zcchc6調控IL-10的機制，利用Luciferase reporter assays和primary transcript analysis，我們發現Zcchc6調控LPS所誘導的IL-10表現量，是通過其啟動子-355到+103的區域。此外，在Zcchc6缺失的 RAW264.7細胞中， LPS 增強STAT1的mRNA和蛋白質表現量，而先前的研究中STAT1已被推測參與IL-10的轉錄反應，這表示Zcchc6調節IL-10表現量，可能部分是通過調解STAT1的mRNA穩定性。由於IL-10是一種抗發炎細胞激素，在病原體攻擊時，IL-10的調控對於平衡發炎的影響是非常重要的，以確保清除病原體，並減少炎症導致的組織損傷。我們的研究證實Zcchc6可能會通過改變IL-10的轉錄因子的mRNA穩定度，以調控TLR4所媒介的IL-10表現量。因此，這項研究應可進一步幫助了解炎症反應和感染性疾病之間的調節。

The innate immunity is the first line of defense in the body. This system is vital as it provides immediate protective functions when the body is under infection and thus contributes to the clearance of pathogens and the regulation of the adaptive immunity. Toll-like receptors (TLRs) are an essential component of the innate immunity. They play a vital role in the recognition of pathogen associated molecular patterns upon infection and then initiate the downstream pro-inflammatory and antiviral responses. Dysregulation of the inflammatory response has been associated with a variety of diseases, such as cancer, metabolic diseases and inflammatory bowel disease. Thus, the TLR-mediated responses must be tightly controlled. In the previous study, our lab identified a Terminal Uridylyl Transferase (TUTase) member, Zcchc6, which is upregulated in murine macrophages upon LPS stimulation. TUTases are enzymes that are known to be involved in the regulation of RNA stability by adding uridine tails to the end of their RNA substrates. They have been implicated in both RNA stabilization and destabilization. We found that the deletion of Zcchc6 in RAW 264.7 macrophage cells led to the dysregulation of many cytokines after the stimulation of TLRs ligands. One of the cytokines that was affected was IL-10, which is a potent anti-inflammatory cytokine with many important immunoregulatory functions. Zcchc6-deleted RAW 264.7 macrophages had increased IL-10 expression at 4 hours after TLRs activation. In this study we looked at the underlying mechanisms by which Zcchc6 regulated the expression of IL-10 in response to LPS. Analysis using Luciferase reporter assays and primary transcript analysis, we found that Zcchc6 mediated the LPS-induced IL-10 expression through the -355/+103 region of its promoter. In addition, LPS enhanced the mRNA and protein levels of STAT1, that has been speculated to be involved in Il10 transcription, in the Zcchc6-deleted cells suggesting that Zcchc6 partially regulated IL-10 expression by mediating the STAT1 mRNA stability. Since IL-10 is an anti-inflammatory cytokine, its regulation upon pathogen attack is important in balancing the effects of inflammation to ensure pathogen clearance and to minimize inflammatory-associated tissue damage. Our findings strongly suggests that Zcchc6 plays a crucial role in the regulation of TLR4-mediated IL-10 expression and this possibly occurs through the alteration of the mRNA stability of transcription factors involved in Il10 transcription. Thus, this research can help further the knowledge in the fine-tuning of inflammatory responses and infectious diseases.