CSTF2 facilitates ATAD2 translation via m⁶A recognition to drive ER-positive breast cancer metabolic reprogramming and tumor progression

Abstract

Abstract Objective Breast cancer (BC) is the most common malignancy among women worldwide, accounting for 11.7% of female cancer cases. Cleavage stimulation factor subunit 2 ( CSTF2 ), a key RNA-binding protein, modulates tumor development by influencing N⁶-methyladenosine (m⁶A) modifications of mRNAs. This study aimed to clarify the precise molecular mechanism through which CSTF2 contributes to BC progression. Methods CSTF2 and ATAD2 expression levels were analyzed using public databases and in BC cell lines. After CSTF2 overexpression in MCF-7 cells, proliferation, migration, and invasion were assessed by CCK-8, EdU, and Transwell assays. Apoptosis was measured by flow cytometry. The effects of CSTF2 overexpression on mitochondrial membrane potential, glucose metabolism, and energy metabolism were also evaluated. Downstream targets of CSTF2 were predicted using ENCORI and SRAMP tools, and the CSTF2 - ATAD2 interaction was validated by RNA immunoprecipitation (RIP). Actinomycin D assays were performed to assess ATAD2 mRNA stability, and polysome profiling was used to evaluate translation efficiency. Rescue experiments and a xenograft mouse model further confirmed the role of the CSTF2/ATAD2 axis. Results CSTF2 was highly expressed in BC tissues and cell lines. Functional assays indicated that CSTF2 overexpression enhanced MCF-7 proliferation, migration, and invasion while reducing apoptosis. JC-1 staining revealed increased mitochondrial membrane potential (MMP) upon CSTF2 upregulation. Glucose and lactate assays demonstrated that CSTF2 overexpression promoted glucose uptake and lactate production in MCF-7 cells. Energy metabolism analysis demonstrated elevated ECAR and diminished OCR following CSTF2 overexpression. Western blot results indicated CSTF2 increased the expression of glycolysis-related proteins (PKM2, HK2) and decreased the levels of oxidative phosphorylation proteins (SDHB, COX IV). Mechanistically, CSTF2 bound to m⁶A-modified ATAD2 mRNA. This binding enhanced mRNA stability and translation efficiency, leading to increased ATAD2 expression. Rescue experiments demonstrated ATAD2 knockdown reversed CSTF2 -induced proliferation, migration, invasion, apoptosis resistance, and metabolic changes in MCF-7 cells. In vivo CSTF2 overexpression accelerated tumor growth and altered metabolic protein expression, effects that were abrogated by ATAD2 silencing. Conclusion In vitro, CSTF2 overexpression enhanced BC cell proliferation, migration, and invasion, promoted glucose uptake and lactate production, and induced glycolytic metabolic reprogramming. In vivo, these observations were confirmed, and CSTF2 overexpression accelerated tumor progression, upregulated HK2 and PKM2, and suppressed SDHB and COX IV in mouse tumor tissues.