Abstract

Histone lysine L-lactylation (hereafter referred to as histone Kla) is a chromatin modification induced by glycolytic metabolism, linking metabolic reprogramming and chromatin-mediated regulation. In this study, we uncover a transcriptional regulatory circuit involving AF9 and KLF2 that drives luminal breast cancer progression. AF9, identified as a reader of H3K9la, promotes KLF2 expression, while KLF2, functioning as a transcription factor for AF9, forms a positive feedback loop amplifying lactylation-dependent effects. This circuit activates tumor-associated pathways, including TGF-β1, glucose and lactate transporters, and metabolic enzymes essential for glycolysis and serine biosynthesis, driving tumorigenesis. Spatial and single-cell transcriptomics show AF9-positive tumor cells enriched in regions of active lactylation, correlating with immune evasion via M2 macrophage interactions. Together, AF9, H3K9la, and KLF2 integrate metabolism, chromatin regulation, and signaling to promote tumor progression, highlighting AF9’s central role as a histone lactylation reader and potential therapeutic target in breast cancer.