Mef2c的磷酸化調控漿狀樹突細胞的發育

Abstract

樹突細胞作為先天免疫系統以及後天免疫的重要橋梁，其可依照特定的表面抗原以及獨特的功能被分類為漿狀樹突細胞(pDC)、第一型及第二型傳統樹突細胞(cDC1, cDC2)。我們先前已透過廣泛篩選的方式挑選Mef2c 轉錄因子作為研究對象。Mef2c對於心肌細胞發育以及血管生成具有不可或缺的角色。而相較於傳統樹突細胞，其高度表現在pDC中，但Mef2c對於pDC發育的影響卻不甚了解。我們在永生化的造血前驅及幹細胞株(iHSPC)中，發現Mef2c 的缺失致使pDC的分化嚴重受損。而在Mef2c條件式基因剃除小鼠(CD11c-Cre Mef2cf/f, cKO mice) 中，在脾臟以及淋巴結中的pDC比例以及細胞數量皆有明顯下降，而在骨髓中則與對照組無統計差異，這也代表Mef2c不僅調控pDC的發育，同時也調控其遷徙能力。我們首先探討Mef2c 是否會影響pDC細胞凋亡，發現不論是骨髓或是脾臟中的pDC 其細胞凋亡速率無統計差異。但由cKO老鼠CDP或CLP體外培養出的pDC 比例以及數量皆有顯著性下降，顯示Mef2c缺失會對於細胞本身造成影響。而在iHSPC加入會磷酸化Mef2c的活化蛋白p38以及Erk5的抑制劑後，則會降低iHSPC發育成為pDC的能力。相似的性狀也會在活化蛋白基因敲落(knockdown) 的 iHPSC上觀察到，甚而在FL3刺激下，在基因敲落細胞也觀察到Mef2c下游基因,Klf2,表現量降低的性狀。以上結果顯示Mef2c磷酸化對於pDC發育是重要的。總結來說，我們透過體內以及體外實驗發現Mef2c調控前驅細胞如先天免疫CDP 或後天免疫CLP發育成為pDC。更多的是，Mef2c 透過MAPK如p38 或Erk5 活化的訊息路徑對於pDC發育是至關重要的。

Dendritic cells (DCs) bridging innate immunity and adaptive immunity are divided into plasmacytoid DC (pDCs), conventional type 1 DC (cDC1), and conventional type 2 DC (cDC2), which have distinct surface markers and functions. Mef2c, a transcription factor, is known to be critical for cardiac morphogenesis and vascular development. While Mef2c is preferentially expressed in pDC as opposed to cDC, its role in pDC development remains unclear. Mef2c deficiency in immortalized hematopoietic stem and progenitor cells (iHSPC) showed impaired pDC development in vitro. CD11c-Cre Mef2cf/f conditional knockout (cKO) mice decreased pDC percentages and cell numbers in the spleen and lymph nodes, indicating that Mef2c not only regulates pDC development but controls migratory ability of this population in vivo. While the apoptosis rate was comparable between control and Mef2c deficient BM and splenic pDC, in vitro development of pDC from both CDP and CLP of the cKO mice was impaired, suggesting that the defect is intrinsic to the loss of Mef2c. Treatment of inhibitors of p38 or Erk5, two kinases known to activated Mef2c, impaired pDC development in vitro. A similar phenotype was observed when either kinase was selectively knocked down. The expression of Klf2, a downstream target of Mef2c was attenuated in the p38 or Erk5 knockdown iHSPC as opposed to control following FL stimulation. These results suggest that the phosphorylation of Mef2c is important for pDC development. Together, we have defined Mef2c as a transcription factor that regulates pDC development in vitro and in vivo from progenitors of both myeloid and lymphoid lineages. Moreover, the activation of Mef2c by MAPKs, such as p38 and Erk5, is critical for controlling pDC development.