The transcriptional regulators GATA6 and TET1 regulate the TGF-β pathway in cancer-associated fibroblasts to promote breast cancer progression.

Cancer-associated fibroblasts (CAFs) are pivotal drivers of tumor progression, yet the molecular mechanisms underlying their activation remain incompletely understood. Here, we identified the TET1/SMAD4/GATA6 regulatory axis as a central mechanism governing CAF transformation and function in breast cancer. Through integrative in vitro and in vivo models, we demonstrated that TET1, an epigenetic modulator, demethylates the SMAD4 promoter, enhancing SMAD4 expression. SMAD4 transcriptionally upregulates GATA6, which amplifies TGF-β signaling by directly activating the TGF-β promoter, establishing a self-reinforcing feedforward loop critical for CAF identity and stromal-tumor crosstalk. GATA6 and TET1 were significantly upregulated in breast CAFs compared to normal fibroblasts (NFs) and TGF-β-induced CAFs. Loss- or gain-of-function experiments revealed that these regulators control CAF survival, marker expression, and secretion of pro-tumorigenic factors. Knockdown of GATA6 or TET1 reduced CAF-mediated migration and invasion of breast cancer cells in vitro, while their overexpression enhanced cancer cell aggressiveness. Mechanistically, TET1-mediated epigenetic remodeling and GATA6-driven transcriptional activation converge on the TGF-β/SMAD pathway, sustaining CAF activation. In vivo, tumors derived from GATA6- or TET1-depleted CAFs exhibited reduced growth, proliferation, and CAF engraftment, underscoring their role in tumor progression. These findings position GATA6 and TET1 as promising targets to disrupt CAF-driven tumorigenesis, offering novel strategies for breast cancer treatment. By unraveling the epigenetic-transcriptional interplay within the tumor microenvironment, this study advances our understanding of stromal reprogramming and its implications for precision oncology.