Department of Life Sciences, and Ph.D. Program in Translational Medicine, National Chung Hsing University, Taichung 402, Taiwan.
Department of Animal Science and Technology, National Taiwan University, Taipei 106, Taiwan.
Int J Mol Sci. 2021 Jan 5;22(1):460. doi: 10.3390/ijms22010460.
The maternal-to-zygotic transition (MZT), which controls maternal signaling to synthesize zygotic gene products, promotes the preimplantation development of mouse zygotes to the two-cell stage. Our previous study reported that mouse granzyme g (Gzmg), a serine-type protease, is required for the MZT. In this study, we further identified the maternal factors that regulate the promoter activity in the zygote to the two-cell stage of mouse embryos. A full-length promoter from mouse genomic DNA, FL-p (-1696+28 nt), was cloned, and four deletion constructs of this promoter, Δ1-p (-1369+28 nt), Δ2-p (-939+28 nt), Δ3-p (-711+28 nt) and Δ4-p (-417+28 nt), were subsequently generated. Different-sized promoters were used to perform promoter assays of mouse zygotes and two-cell stage embryos. The results showed that Δ4-p promoted the highest expression level of the enhanced green fluorescent protein (EGFP) reporter in the zygotes and two-cell embryos. The data suggested that time-specific transcription factors upregulated by binding cis-elements in the -417+28-nt promoter region. According to the results of the promoter assay, the transcription factor binding sites were predicted and analyzed with the JASPAR database, and two transcription factors, signal transducer and activator of transcription 3 (STAT3) and GA-binding protein alpha (GABPα), were identified. Furthermore, STAT3 and GABPα are expressed and located in zygote pronuclei and two-cell nuclei were confirmed by immunofluorescence staining; however, only STAT3 was recruited to the mouse zygote pronuclei and two-cell nuclei injected with the Δ4-p reporter construct. These data indicated that STAT3 is a maternal transcription factor and may upregulate to promote the MZT. Furthermore, treatment with a STAT3 inhibitor, S3I-201, caused mouse embryonic arrest at the zygote and two-cell stages. These results suggest that STAT3, a maternal protein, is a critical transcription factor and regulates transcription activity in preimplantation mouse embryos. It plays an important role in the maternal-to-zygotic transition during early embryonic development.
母体-合子过渡（MZT）控制母体信号合成合子基因产物，促进小鼠合子在植入前发育到两细胞阶段。我们之前的研究报告称，小鼠颗粒酶 G（Gzmg），一种丝氨酸型蛋白酶，是 MZT 所必需的。在这项研究中，我们进一步鉴定了调节小鼠胚胎合子到两细胞阶段启动子活性的母体因子。从小鼠基因组 DNA 克隆全长 启动子，FL-p（-1696+28nt），随后生成了该 启动子的四个缺失构建体，Δ1-p（-1369+28nt）、Δ2-p（-939+28nt）、Δ3-p（-711+28nt）和 Δ4-p（-417+28nt）。使用不同大小的 启动子对小鼠合子和两细胞期胚胎进行启动子测定。结果表明，Δ4-p 促进了绿色荧光蛋白（EGFP）报告基因在合子和两细胞胚胎中的最高表达水平。数据表明，通过结合 -417+28-nt 启动子区域内的顺式元件，时间特异性转录因子上调。根据启动子测定的结果，使用 JASPAR 数据库预测和分析了转录因子结合位点，并鉴定了两个转录因子，信号转导和转录激活因子 3（STAT3）和 GA 结合蛋白α（GABPα）。此外，通过免疫荧光染色证实 STAT3 和 GABPα 在合子原核和两细胞核中表达并定位；然而，只有 STAT3 被募集到注射了Δ4-p 报告基因构建体的小鼠合子原核和两细胞核中。这些数据表明 STAT3 是一种母体转录因子，可能上调 以促进 MZT。此外，用 STAT3 抑制剂 S3I-201 处理导致小鼠胚胎在合子和两细胞阶段停滞。这些结果表明，STAT3 作为一种母体蛋白，是一种关键的转录因子，调节植入前小鼠胚胎中 的转录活性。它在早期胚胎发育过程中的母体-合子过渡中起着重要作用。
