AKT Phosphorylation of FOXK2 at Ser398 Impacts Breast Cancer Cell Growth and Viability

Abstract

ABSTRACT FOXK2 is a transcription factor known to mediate drug sensitivity in breast cancer. However, poor prognosis patients and drug‐resistant cells exhibit constitutively elevated levels of FOXK2 protein, which appears functionally inactive. Although emerging evidence suggests that FOXK2 is regulated by post‐translational modifications, the underlying mechanisms in breast cancer remain poorly understood. An in silico analysis identified AKT as a putative regulator of FOXK2, prompting the investigation of AKT‐mediated phosphorylation of FOXK2 in cancer. Our results show that elevated FOXK2 protein levels are associated with increased AKT phosphorylation and doxorubicin resistance. Modulation of AKT levels reduces FOXK2 at protein but not mRNA levels. Consistently, analysis of public breast tumor datasets revealed increased FOXK2 phosphorylation at different sites, including serine 398. Immunoblotting with an antibody against AKT substrates and lambda phosphatase treatment indicates that serine 398 in FOXK2 is an AKT phosphorylation site. Notably, the phosphodeficient FOXK2 mutant (S398A) impaired cell growth, metabolic cell viability, and clonogenic capacity, even in the presence of chemotherapeutic drugs. In summary, AKT regulates FOXK2 in drug‐resistant breast cancer cells, with serine 398 emerging as a novel phosphorylation site. Our findings reveal key insights into FOXK2 regulation and the contribution to understanding breast cancer cell biology.