RASGRP3 orchestrates trophoblast morphogenesis of placental cotyledons via RAP1-dependent signaling in goat (Capra hircus).

The development of placental cotyledons is essential for fetal growth in ruminants, with trophoblasts playing a pivotal role in placental formation and nutrient exchange through tightly regulated molecular mechanisms. RAS guanyl-releasing protein 3 (RASGRP3), a guanine nucleotide exchange factor, is known to regulate cell proliferation and migration; however, its role in ruminant placental development remains unclear. In this study, we investigated the function of RASGRP3 in goat placentation using integrated in vitro and in vivo approaches. RASGRP3 expression was found to peak in trophoblast cells during mid-gestation (days 45-90) and was positively correlated with cotyledon size. Functional assays revealed that knockdown of RASGRP3 inhibited trophoblast proliferation, migration, and invasion, while promoting apoptosis and mitochondrial dysfunction. Mechanistically, RASGRP3 activated the PI3K/AKT/mTOR signaling pathway via RAP1GTP, and pharmacological activation of RAP1 or PI3K/AKT partially restored the impaired cellular functions caused by RASGRP3 depletion. In vivo, adenovirus-mediated gene silencing of RASGRP3 in pregnant goats led to reduced cotyledon size and trophoblast GTPase activity, whereas overexpression produced the opposite effects. These findings identify RASGRP3 as a key regulator of trophoblast function and placental development in goats through the RAP1-mediated PI3K/AKT/mTOR pathway, representing the first experimental evidence that elucidates the role and molecular mechanism of RASGRP3 in ruminant placental development and offering novel insights into the molecular basis of trophoblast regulation and placental morphogenesis.