RSPO2-induced ferroptosis via PTBP1-mediated FSP1 mRNA decay suppresses breast cancer progression

Breast cancer is the most common malignant tumor worldwide, and its high heterogeneity limits the efficacy of current clinical therapies. R-spondin 2 (RSPO2) expression is closely associated with breast cancer subtypes, but the precise mechanisms by which it drives disease progression, particularly its relationship with ferroptosis, remain unclear.

RSPO2 expression was examined in primary breast cancer specimens and in MCF-7 and MDA-MB-231 cell lines. Functional assays, including proliferation, migration, and invasion tests, were performed following RSPO2 overexpression. In vivo tumor growth was assessed using a xenograft mouse model. Mechanistically, immunoprecipitation, ubiquitination assays, RNA sequencing (RNA-seq), RIP-qPCR, mRNA stability assays, and ferroptosis-related measurements (ROS, GSH, MDA, Fe²⁺) were employed to identify interacting proteins and downstream targets. Rescue experiments with FSP1 overexpression were conducted to validate pathway specificity.

RSPO2 was consistently downregulated in malignant breast cancer tissues. Restoring RSPO2 expression markedly suppressed cell proliferation, migration, and invasion in vitro and inhibited tumor growth in vivo. Mechanistically, RSPO2 recruited the E3 ubiquitin ligase TRIM21 to promote PTBP1 ubiquitination and proteasomal degradation. RNA-seq and validation assays revealed that PTBP1 bound the 3'UTR of ferroptosis suppressor protein 1 (FSP1) mRNA, enhancing its stability. RSPO2 overexpression reduced PTBP1 levels, leading to decreased FSP1 mRNA stability and expression. Consequently, RSPO2 overexpression triggered ferroptosis, as evidenced by ROS accumulation, GSH depletion, and elevated MDA and Fe²⁺ levels. These effects were effectively reversed by FSP1 overexpression, which also abrogated the RSPO2 induced inhibition of cell proliferation, migration, and invasion.

This study demonstrates that RSPO2 functions as a tumor suppressor in breast cancer by recruiting TRIM21 to degrade PTBP1, thereby reducing FSP1 mRNA stability and promoting ferroptosis. The RSPO2/PTBP1/FSP1 signaling axis is identified for the first time in breast cancer and represents a promising novel therapeutic target.