TOX3 Promotes Ovarian Estrogen Synthesis: An RNA-Sequencing and Network Study

Front Endocrinol (Lausanne). 2021 Feb 24:11:615846. doi: 10.3389/fendo.2020.615846. eCollection 2020.


Background: Women who undergo chronic exposure to excessive estrogen are at a high risk of developing breast cancer. TOX3 has been reported to be highly expressed in breast tumors and is closely related to estrogen receptors. However, the effect of TOX3 on estrogen synthesis remains poorly understood.

Methods: Using lentiviruses as a vector, we stably overexpressed TOX3 in the ovarian granulosa cell line KGN, the cells where estradiol is primarily produced, to investigate its role in estrogen production as well as cell viability and apoptosis. RNA-Sequencing was applied to uncover the global gene expression upon TOX3 overexpression.

Results: We observed an increased level of cell viability and a reduced cell apoptosis rate after TOX3 overexpression, and the level of estradiol in the cell culture supernatant also increased significantly. Gene set enrichment analysis of the transcriptome showed that the ovarian steroidogenesis pathway was significantly enriched. Similarly, pathway mapping using the Kyoto Encyclopedia of Genes and Genomes and Gene Ontology analyses also showed that TOX3 overexpression affects the ovarian steroidogenesis pathway. Further experiments showed that upregulated FSHR, CYP19A1, and BMP6 accounted for the enhanced estrogen synthesis.

Conclusion: Our study demonstrated that TOX3 quantitatively and qualitatively stimulates estrogen synthesis by enhancing estrogen signaling pathway-related gene expression in ovarian granulosa cells. These findings suggest that TOX3 may play a vital role in the pathogenesis of breast cancer.

Keywords: GSEA; KGN; RNA-Seq; TOX3/TNRC9; breast cancer; estrogen.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Apoptosis Regulatory Proteins / biosynthesis*
  • Apoptosis Regulatory Proteins / genetics
  • Cell Line, Tumor
  • Estrogens / biosynthesis*
  • Estrogens / genetics
  • Female
  • Gene Regulatory Networks / physiology*
  • Granulosa Cells / metabolism*
  • Humans
  • Ovary / metabolism
  • Sequence Analysis, RNA / methods*
  • Trans-Activators / biosynthesis*
  • Trans-Activators / genetics


  • Apoptosis Regulatory Proteins
  • Estrogens
  • TOX3 protein, human
  • Trans-Activators