The role of epithelial Progesterone Receptor Isoforms in embryo implantation [RNAseq-Foxo1dd_LE]

RNA-seq analysis of uterine luminal epithelium at D4.5 indicates that epithelial PGRA and PGRB shares conseved pathways. Constitutive epithelial PGRA and PGRB disrupts the embryo implantation both through the suppressed FOXO1 signaling by excluding FOXO1 from the nuclear at the uterine epithelium. There are three layers of regulation. Firstly, PGRA and PGRB diminishes Lif transcription in uterine glands by blocking ESR1 binding at the Lif promoter at D3.5 which is critical for the FOXO1 nuclear expression at D4.5 through LIF/pSTAT3/FOXO1. Secondly, PGRA and PGRB directly suppres Foxo1 transcription at the uterine epithelium probably through direct binding at Foxo1 promoter. Thirdly, PGRA and PGRB promotes the Sgk1 transcription, the kinases that phosphorylate FOXO1 to translocate it into cytoplasma for degradation. The uterine luminal epithelium were collected from Foxo1f/f and Pgrcre/+ Foxo1f/f mice at pregnancy day 4 by laser capture for RNA-seq analysis

妊娠第4.5天（D4.5）子宫腔上皮细胞的RNA测序（RNA-seq）分析显示，上皮细胞中的孕激素受体A（PGRA）与孕激素受体B（PGRB）共享保守信号通路。上皮细胞中组成型表达的PGRA与PGRB可通过将子宫腔上皮细胞内的叉头框蛋白O1（FOXO1）排出细胞核，抑制FOXO1信号通路，进而干扰胚胎着床。该过程存在三层调控机制：其一，在妊娠第3.5天（D3.5），PGRA与PGRB可通过阻断雌激素受体1（ESR1）结合至白血病抑制因子（LIF）的启动子区域，抑制子宫腺体中LIF的转录；而LIF可通过LIF/pSTAT3/FOXO1信号轴调控妊娠第4.5天FOXO1的细胞核定位。其二，PGRA与PGRB可直接抑制子宫腔上皮细胞内Foxo1的转录，该过程可能通过直接结合至Foxo1的启动子区域实现。其三，PGRA与PGRB可促进血清和糖皮质激素调节激酶1（SGK1）的转录，该激酶可磷酸化FOXO1，使其转位至细胞质并发生降解。本研究通过激光捕获技术，从妊娠第4天的Foxo1f/f及Pgrcre/+ Foxo1f/f小鼠中分离子宫腔上皮细胞，用于后续RNA-seq分析。