終端尿醯苷轉移酶TUT7在類鐸受體4引發發炎反應扮演之角色及機制探討

Abstract

先天性免疫反應之於宿主的重要性在於其能有效地清除病原體及修復受傷的組織，然失去調控的免疫反應則會導致許多疾病，例如:癌症、敗血症及代謝疾病等。目前已知，許多機制已被證實能有效地調節免疫反應，其中包含轉錄後調控(post-transcriptional regulation/PTR)。針對免疫相關基因的表現，轉錄後調控是一種快速且有效率的方式。第七終端尿醯苷轉移酶(Terminal uridyltransferase 7，TUT7)，亦稱為ZCCHC6；目前的研究已知TUT7藉由轉移尿醯苷至其標的核醣核酸(RNA)的方式，調控核糖核酸(RNA)的穩定度及生合成。然而，TUT7在先天性免疫系統中所扮演的角色仍未釐清。於本論文中，我們發現以脂多醣(LPS)刺激第四型類鐸受體(TLR4)的活化，會誘發TUT7的表現。再者，利用慢病毒攜帶短片段RNAi (shTut7)或是特定基因剔除鼠(Tut7-/-)減少或去除巨噬細胞裡TUT7的表現並施以LPS，導致部分的細胞激素表現失衡，如:細胞介白素6 (interleukin-6/IL-6)。另外，我們更進一步證實TUT7利用其尿醯苷轉移之能力，將調節型核糖核酸內切酶(Regulatory riboendonuclease-1/Regnase-1)的RNA進行尿醯化的修飾以減少其蛋白質產生，進而對IL-6 mRNA達到保護作用，最終使IL-6得以適時地產生作用。另外，與先前的研究一致，我們也發現尿醯化的Regnase-1 mRNA具有較短的腺苷鏈。綜合前述，我們證實了TUT7藉由尿醯化調節Regnase-1 mRNA穩定度降低Regnase-1蛋白質的表現，以此使IL-6可以適時地參與TLR4所誘導之免疫反應。

Innate immune responses play a crucial role for the host to effectively clear pathogens and repair damaged tissues, but their dysregulation has been associated with numerous illnesses, such as cancer, sepsis and metabolic disorders. Different mechanisms have been demonstrated to elaborately regulate inflammatory response, including post-transcriptional regulation (PTR), which is a fast and effective way to modulate the expression of inflammatory-related genes. Terminal uridyltransferase 7 (TUT7), also named ZCCHC6, is known to be involved in the regulation of RNA stability or microRNA biogenesis by the addition of uridine tails to the end of their RNA substrates. Nevertheless, the role of TUT7 in the innate immune response remains unclear. Here we reveal that TUT7 is induced upon TLR4 activation, and selectively controls the expression of a subset of inflammatory cytokines. Depletion of TUT7 in macrophages leads to dysregulation of several cytokines production including IL-6 upon LPS challenge. We further demonstrate that TUT7 mediates IL-6 expression by controlling its mRNA stability through Regnase-1 after LPS stimulation. Mechanistically, TUT7 directly interacts with and uridylates Regnase-1 for degradation following stimulation LPS, which required the stem-loop structure in its 3’-UTR. Consistently, Regnase-1 mRNAs possess uridylated ends and shorter poly(A) tails in TUT7-proficient macrophages after LPS treatment, whereas Regnase-1 transcripts carries long poly(A) length without uridylation in Tut7-/- cells. Taken together, our findings reveal the functional role of TUT7 in sculpting TLR4-driven responses by modulating the mRNA stability of certain inflammatory mediators.