Background <p>The reprogramming of energy metabolism, particularly glycolysis, upholds the malignancy of tumors. The relationship between CUE domain-containing protein-1 (CUEDC1) and glycolysis, along with its influence on the development of estrogen receptor-positive breast cancer (ER⁺ BRCA), is not well defined. This investigation explores the functional involvement of CUEDC1 in glycolysis regulation and uncovers a previously unidentified pathway contributing to the progression of ER⁺ BRCA.</p> Methods <p>Immunohistochemistry, western blotting and quantitative real-time polymerase chain reaction (qRT-PCR) were used to detect the expression of CUEDC1 in ER⁺ BRCA tissues and cell lines. A series of molecular analyses, including dual-luciferase reporter assays, RNA-seq, and chromatin immunoprecipitation (ChIP), were performed to elucidate the potential mechanisms underlying CUEDC1’s involvement in ER⁺ BRCA progression. Metabolic assays focusing on glycolysis were employed to investigate the functional roles of CUEDC1 and calcium voltage-gated channel auxiliary subunit gamma 4 (CACNG4). The Connectivity Map (CMap) database was utilized to screen CUEDC1 inhibitors.</p> Results <p>CUEDC1 is aberrantly upregulated in ER⁺ BRCA tissues and cells. Increased CUEDC1 can promote enhanced tumor growth and lead to poor clinical outcomes in patients with ER⁺ BRCA. The MYC associated zinc finger protein (MAZ) can upregulate CUEDC1 gene transcription in ER⁺ BRCA cells by directly binding to its promoter. We determined that CUEDC1 directly modulated CACNG4 to enhance phosphatidylinositol 3-kinase (PI3K)/protein kinase B (AKT) pathway activation, thereby facilitating tumor growth of ER⁺ BRCA. Clinical observations also revealed a positive correlation between CACNG4 expression and CUEDC1, with both factors being strongly associated with poor prognosis in patients with ER⁺ BRCA. Mechanistically, The CUEDC1/CACNG4/PI3K signal axis enhanced glycolysis through upregulating glucose transporter 1 (GLUT1), a crucial protein in glucose metabolism, thereby supporting tumor growth of ER⁺ BRCA. Furthermore, methotrexate was identified as a potential inhibitor of CUEDC1. Importantly, the combination of ipatasertib (a PI3K/AKT pathway inhibitor), with methotrexate effectively suppressed growth of ER⁺ BRCA in a mouse model.</p> Conclusions <p>Our research reveals that enhanced CUEDC1 plays an essential role to ER⁺ BRCA cell proliferation and tumor growth via the CACNG4/PI3K axis. CUEDC1 is a promising prognostic factor of ER⁺ BRCA, and the CUEDC1/CACNG4/PI3K axis can serve as a potential therapeutic target for ER⁺ BRCA treatment.</p>