白质精氨酸甲基转移酶5 (Prmt5)调控肺分支发育及其分子机制研究

Histone arginine methylation regulates of chromatin structure, controls of cell differentiation and ontogeny by regulating gene expression. Histone H4R3sme2 methylation is catalyzed mainly by Prmt5. In the preliminary study of this proposal, we found that the Prmt5 is highly expressed in lung epithelial cells, and we has generated Prmt5flox/flox;Shh+/cre mice, which the Prmt5 was specially knockout in lung epithelial cells. We found that Prmt5 knockout resulted in fetal lung branching defect, Prmt5 knockout mice died at P0 due to severe respiratory defects. We also found that the Bmp4 expression was significantly increased in Prmt5 knockout mice, indicates that Prmt5 regulates Bmp4 transcription during lung branching morphogenesis. In this proposal, we tend to investigate the biological roles of Prmt5-mediated H4R3sme2 methylation in Bmp4 regulation and lung branching morphogenesis, through immunohistochemical, immunofluorescence, biochemistry and pathology analysis. Meanwhile, combined with the analysis of clinical samples of lung disease, try to find a correlation between Prmt5 and pulmonary disease. We believe the studies in this proposal will help us to understand the regulation mechanisms of epigenetics on organ development, and has important scientific significance for the pathogenesis of lung disease and interpretation of early diagnosis strategy.

组蛋白精氨酸甲基化修饰调控染色质结构，通过调控基因表达控制细胞分化和个体发育，组蛋白H4R3sme2甲基化修饰主要由Prmt5催化。本项目前期研究发现Prmt5在小鼠肺上皮细胞中高表达，并获得了Prmt5在肺上皮细胞中特异敲除小鼠。对Prmt5敲除小鼠分析发现，Prmt5敲除后导致肺分支发育异常，小鼠在出生当天由于呼吸窘迫死亡；但Prmt5敲除后并不影响肺上皮细胞分化。另外，Prmt5敲除小鼠中Bmp4表达上调，提示Prmt5可能通过调控Bmp4转录调控肺分支发育。在此基础上，本项目拟通过免疫荧光、生物化学和病理等分析，阐明Prmt5及其催化的H4R3sme2对Bmp4和小鼠肺分支发育的调控机制。同时，结合分析临床肺病样本，建立Prmt5和发育缺陷相关肺病间的相关性。通过本项目研究，对我们深入理解表观遗传对器官发育的调控机制，诠释肺发育相关疾病的发病机理以及早期诊疗具有重要科学意义。

肺分支形态建成过程是肺发育成一个具有气体交换功能的呼吸器官的关键步骤。近年来一些研究发现骨形态发生蛋白（BMP）信号通路在肺分支形态建成过程中起重要作用，但对BMP信号的调节机制知之甚少。本项目研究发现蛋白质精氨酸甲基转移酶5（Prmt5）和Prmt5催化的组蛋白H4精氨酸3（H4R3sme2）的对称双甲基化直接与Bmp4染色质结合并抑制其转录。在肺上皮细胞中特异的敲除Prmt5后导致肺分支形态建成缺陷，上皮细胞分化改变和新生小鼠死亡。这些小鼠缺陷伴随着肺上皮细胞凋亡的增加和增殖的减少，这是典型的BMP-Smad1/5/9信号转导升高导致的生物学结果。通过Noggin抑制BMP信号转导过程后，可以在体外恢复Prmt5突变体的肺分支发育以及细胞增殖和凋亡异常。总之，本项目研究鉴定了一种新的分子机制，发现通过Prmt5介导的组蛋白精氨酸甲基化抑制经典的BMP信号通路，进而调节肺分支的形态建成过程。