Marikawa Yusuke, Alarcon Vernadeth B
Yanagimachi Institute for Biogenesis Research, Department of Anatomy, Biochemistry and Physiology, University of Hawaii John A. Burns School of Medicine, Honolulu, HI, 96813, USA.
Yanagimachi Institute for Biogenesis Research, Department of Anatomy, Biochemistry and Physiology, University of Hawaii John A. Burns School of Medicine, Honolulu, HI, 96813, USA.
Differentiation. 2025 Jan-Feb;141:100835. doi: 10.1016/j.diff.2025.100835. Epub 2025 Jan 3.
The trophectoderm (TE) is the first tissue to differentiate during the preimplantation development of the mammalian embryo. It forms the outer layer of the blastocyst and is responsible for generating the blastocoel, a fluid-filled cavity whose expansion is essential for successful hatching and implantation. Here, we investigated the role of the small GTPase RHOA in the morphogenesis of the TE, particularly its relationship with HIPPO signaling, using mouse embryos as a model. Inhibition of RHOA resulted in the failure to form a blastocoel and significantly altered the expression of numerous genes. Transcriptomic analysis revealed that 330 genes were down-regulated and 168 genes were up-regulated by more than two-fold. Notably, 98.4% of these transcriptional changes were reversed by simultaneous inhibition of LATS kinases, indicating that the transcriptional influence of RHOA is primarily mediated through HIPPO signaling. Many of the down-regulated genes are involved in critical processes of TE morphogenesis, such as apical-basal cell polarization, tight junction formation, and sodium and water transport, suggesting that RHOA supports TE development by enhancing the expression of morphogenesis-related genes through HIPPO signaling, specifically via TEAD transcription factors. However, RHOA inhibition also disrupted apical-basal polarity and tight junctions, effects that were not restored by LATS inhibition, pointing to additional HIPPO signaling-independent mechanisms by which RHOA controls TE morphogenesis. Furthermore, RHOA inhibition impaired cell viability at the late blastocyst stage, with partial rescue observed upon LATS inhibition, suggesting that RHOA maintains cell survival through both HIPPO signaling-dependent and -independent pathways. A deeper knowledge of the molecular mechanisms governing TE morphogenesis, including blastocoel expansion and cell viability, could significantly advance assisted reproductive technologies aimed at producing healthy blastocysts.
滋养外胚层（TE）是哺乳动物胚胎植入前发育过程中第一个分化的组织。它形成囊胚的外层，负责产生囊胚腔，这是一个充满液体的腔，其扩张对于成功孵化和植入至关重要。在这里，我们以小鼠胚胎为模型，研究了小GTPase RHOA在TE形态发生中的作用，特别是它与HIPPO信号通路的关系。抑制RHOA导致无法形成囊胚腔，并显著改变了许多基因的表达。转录组分析显示，330个基因下调，168个基因上调超过两倍。值得注意的是，这些转录变化中有98.4%通过同时抑制LATS激酶而逆转，表明RHOA的转录影响主要通过HIPPO信号通路介导。许多下调的基因参与了TE形态发生的关键过程，如顶-基细胞极化、紧密连接形成以及钠和水的运输，这表明RHOA通过HIPPO信号通路，特别是通过TEAD转录因子增强形态发生相关基因的表达来支持TE发育。然而，抑制RHOA也破坏了顶-基极性和紧密连接，而LATS抑制并不能恢复这些效应，这表明RHOA控制TE形态发生还存在其他不依赖HIPPO信号通路的机制。此外，抑制RHOA会损害晚期囊胚阶段的细胞活力，在抑制LATS后观察到部分挽救，这表明RHOA通过依赖和不依赖HIPPO信号通路的途径维持细胞存活。对控制TE形态发生的分子机制有更深入的了解，包括囊胚腔扩张和细胞活力，可能会显著推进旨在产生健康囊胚的辅助生殖技术。
