Ifitm3 positively regulates homeostasis of long-term cultured oogonial stem cells derived from Paralichthys olivaceus
Yuqin Ren, Yucong Yang, Guixing Wang, Zengsheng Han, Nuan He, Xiyuan Wang, Yitong Zhang, Ziyang He, Xiaodong Cui, Zhongwei He, Yufeng Liu, Wei Cao, Xiaoyan Zhang, Yufen Wang, Xianjiang Kang, Jilun HouAbstract
Fish oogonial stem cells (OSCs) are vital for fish reproduction research and germplasm conservation, yet stable long-term in vitro culture of fish OSCs remains a major technical bottleneck. Herein, we established an efficient in vitro culture system (termed L15SP) for OSCs derived from Paralichthys olivaceus, consisting of L-15 medium supplemented with15% fetal bovine serum (FBS), 2 μg/L basic fibroblast growth factor (bFGF), 2 μg/L leukemia inhibitory factor (LIF), 50 μmol/L β-mercaptoethanol (β-ME), 1% fish serum, and 18 g/L embryo extract protein, cultured at 23 °C. This system enabled stable long-term passage of P. olivaceus OSCs. Further investigations revealed that interferon-induced transmembrane protein 3 (Ifitm3) was highly expressed on the membrane of P. olivaceus OSCs. Functional assays demonstrated that Ifitm3 significantly enhanced proliferation, migration, and stemness maintenance of long-term cultured OSCs. Ifitm3 positively modulated the expression of key PI3K-AKT signaling components, including pik3cb, pik3r1, and akt2, to regulate downstream OSC functional genes. Upon pathway activation, Ifitm3 facilitated PIP3 enrichment at the OSC plasma membrane, thereby amplifying PI3K-AKT signaling. In conclusion, Ifitm3 governed OSC biological functions by potentiating PI3K-AKT signal transduction. This study establishes a reliable long-term culture system for fish OSCs and identifies a novel functional marker for OSC identification, providing valuable support for fish germplasm preservation and genetic breeding.