Histone H3.3K27M mutations are identified in diffuse midline glioma (DMG), resulting in a global reduction in H3K27me3. Analyzing the histone recognition mechanism at H3K27 position can enhance the understanding of this highly lethal mutation. Here we identify a novel histone reader CBFA2T2, which recognizes non-mutated or un-modified histone H3K27. Biochemical assays confirm this binding specificity, indicating that it is mediated by NHR2 domain of CBFA2T2. CBFA2T2 represses the transcription of metabolic genes involved in carbon metabolism, glycolysis/gluconeogenesis and the TCA cycle pathways through its binding to H3K27, thereby regulating metabolite levels. These alter the alpha-ketoglutarate/Succinate ratio and indirectly impacts H3K27me3 level, through affecting the H3K27me3 demethylases. Our results uncover a novel mechanism by which CBFA2T2 transcriptionally regulates metabolism and tumor growth in H3.3K27M cells. These findings suggest that CBFA2T2 and its targeted genes may serve as potential therapeutic targets for the treatment of H3.3K27M cancer.