利用酵母菌雙雜合系統探討 TRAIL 所引起 T 細胞活化其訊息傳導途徑之研究

Abstract

TRAIL 是屬於第二型細胞膜蛋白質，並且是屬於 TNF 家族裡的一員，透過跟死亡受體的結合可以引起細胞的凋亡。TRAIL 主要是以膜結合型的形式存在，而 T 細胞在受到抗 CD3 的抗體或第一型干擾素的刺激之下可以引發 TRAIL 的表現。有越來越多的證據顯示 TNF 家族的成員，像是 CD40L，CD30L，FasL 和 TRANCE，在跟受體結合後都可以傳遞訊息。我們之前的研究顯示，同時給予 T 細胞抗 CD3 抗體和 DR4 的刺激，可以引起 T 細胞的活化和誘導干擾素-γ的產生。這樣的結果指出，TRAIL 在跟死亡受體結合後，除了會導致細胞的凋亡，也會傳遞反向的訊息導致 T 細胞的活化。然而這樣由 TRAIL 所引起的反向訊息傳導途徑仍然不甚清楚。我們之前的研究發現，在 TRAIL 所引起 T 細胞的活化當中可以觀察到 p38 MAPK 和 PI3K/Akt 的活化，顯示 p38 MAPK 和 PI3K/Akt 可能參與在 TRAIL 所引起的反向訊息傳導途徑之中。目前已知，NF-κB 訊息傳導途徑參與在 CD28 所引起的 T 細胞共活化之中，顯示 NF-κB 訊息傳導途徑對於 T 細胞的共活化相當重要。在這篇研究當中，我們利用酵母菌雙雜合系統來識別跟 TRAIL 的細胞質內區域相結合並且幫助 TRAIL 傳遞訊息的可能的分子。研究的結果顯示，JAK1 可能可以跟 TRAIL 的細胞質內區域相結合並且調節 TRAIL 所引起的訊息傳導而使得 T 細胞活化和產生誘導干擾素-γ。進一步的研究顯示，TRAIL 所引起的 T 細胞活化和誘導干擾素-γ的產生可以藉由加入 JAK 抑制劑而得到顯著的抑制，顯示 JAK-STAT 途徑可能參與在 TRAIL 所引起的 T 細胞活化當中。此外，我們也證明了 NF-κB 訊習傳導途徑在 TRAIL 所引起的 T 細胞共活化當中有被活化的現象。

Tumor Necrosis Factor-related Apoptosis-inducing Ligand (TRAIL), a type Ⅱ transmembrane protein, is one of several members of the TNF superfamily that induces apoptosis through engagement of death receptors. TRAIL exists mainly in membrane-bound form, and its expression on T cells is induced after T cell activation by anti-CD3 or type I interferon (IFN). There is growing evidence that ligands of the TNF family, such as CD40L; CD30L; FasL, and TRANCE, transduced signals after engagement with their receptors. Our previous study has demonstrated that TRAIL stimulated with immobilized DR4, in conjunction with suboptimal immobilized anti-CD3, induced T cell proliferation and enhanced IFN-γ production. This indicates that in addition to direct triggering apoptosis through death receptor, TRAIL can tranduce reverse signals to induce T cell activation. However, the reverse signaling pathways transduced by TRAIL is still unclear. In our previous study, we found that p38 MAPK and PI3K/Akt activation was detected after TRAIL-induced T cell activation, indicating that p38 MAPK and PI3K/Akt pathway is involved in TRAIL reverse signaling pathway. It has been demonstrated that the NF-κB signaling pathway was involved in CD28 costimulation signaling pathway, suggesting that NF-κB signaling pathway is critical in costimulation of T cells. In this study, we use yeast-two-hybrid system to identify the possible molecules associated with TRAIL intracytoplasmic domain to transduce signal. Our results showed that JAK1 may be an associated protein in the cytoplasmic region of TRAIL and modulate the signaling transduction of TRAIL-induced T cell proliferation and IFN-γ production. Furthermore, the effect of TRAIL-induced T cell proliferation and IFN-γ production can be significantly blocked by JAK inhibitor, indicating JAK-STAT pathway is critical in TRAIL-induced T cell activation. Moreover, we also demonstrated that NF-κB pathway is activated during TRAIL-induced costimulation of T cells.