ZNF469 drives TGF-β1/SMAD3-mediated extracellular matrix regulation in pulmonary fibrosis

Pulmonary fibrosis is characterized by persistent activation of lung fibroblasts and excessive extracellular matrix (ECM) deposition; however, the transcriptional mechanisms sustaining this activated state remain poorly understood. Given the established profibrotic role of zinc finger protein 469 (ZNF469) in other mesenchymal lineages, we investigated its function in lung fibroblasts using an integrated functional and transcriptomic approach. We found that ZNF469 depletion attenuated key features of fibroblast activation, including proliferation, migration, and contractile capacity, concomitantly reducing collagen production and deposition. Mechanistically, combined transcriptomic profiling and Cleavage Under Targets and Release Using Nuclease (CUT&RUN) assays revealed that ZNF469 modulates ECM-associated transcriptional regulation, demonstrating preferential occupancy at the promoters of key collagen genes, such as collagen type I alpha 1 chain (COL1A1) and collagen type III alpha 1 chain (COL3A1). Furthermore, we demonstrated that transforming growth factor-β1 (TGF-β1) stimulated ZNF469 expression in a SMAD family member 3 (SMAD3)-dependent manner, with CUT&RUN further revealing SMAD3 occupancy at the ZNF469 promoter. Importantly, ZNF469 depletion blunted TGF-β1-induced collagen production, indicating that ZNF469 contributes to the profibrotic effects of the TGF-β1/SMAD3 axis. Finally, we underscored the clinical relevance of ZNF469 by analyzing bulk and single-cell RNA sequencing data from patients with idiopathic pulmonary fibrosis and systemic sclerosis. ZNF469 was found to be upregulated in fibrotic lung tissues, particularly within the activated fibroblast lineage, and correlated with key ECM gene signatures. Our findings collectively support a model wherein the TGF-β1/SMAD3/ZNF469 axis contributes significantly to ECM gene regulation and fibroblast activation, thereby positioning ZNF469 as a critical profibrotic factor and a promising therapeutic target in pulmonary fibrosis.