Single-Cell Heterogeneity of Epigenetic Factor Regulation Deciphers Alteration of RNA Metabolism During Proliferative SHH-Medulloblastoma

<b>Background:</b> Medulloblastoma is an aggressive pediatric brain tumor characterized by marked molecular heterogeneity, which significantly impacts prognosis. The low frequency of genomic mutations in medulloblastoma suggests that alternative mechanisms, such as epigenetic regulation, may play a critical role in its pathogenesis. <b>Methods:</b> Using the EpiFactors database, we investigated the expression of epigenetic regulators in two independent RNA sequencing cohorts [Pediatric Brain Tumor Atlas (PBTA) and Williamson], stratified by molecular subgroups and clinical outcomes. We further analyzed expression heterogeneity at the single-cell level in malignant medulloblastoma cells using single-cell RNA sequencing. <b>Results:</b> Members of the SWI/SNF superfamily were dysregulated across all four molecular subtypes of medulloblastoma. Subtype-specific alterations were also observed: the acetyltransferase complex was shared between Group 3 (with SMARCD3 as a potential marker) and Group 4 (with RBM24 as a potential marker); SWR1, β-catenin/TCF, and protein-DNA complexes were specifically enriched in SHH-MB (with EYA1 and SATB2 as SHH markers); and RSC-type, PRC1, DNA polymerase complexes, and X-chromosome-related factors were enriched in WNT-MB (with FOXA1 and PIWIL4 as WNT markers). An epigenetic score (epi-score), linked to RNA metabolism and S-adenosyl-L-methionine pathways, was developed and identified as an independent adverse prognostic factor. High epi-scores were associated with proliferative, stem-like SHH malignant cells (characterized by G2/M phase, low pseudotime, and high entropy), exhibiting alterations in RNA splicing, DNA recombination, and nuclear division. <b>Conclusions:</b> Expression heterogeneity of epigenetic regulators is closely associated with molecular subgroups and clinical outcomes in medulloblastoma. These findings highlight the role of epigenetic dysregulation in RNA metabolism and tumor progression, particularly in SHH-driven proliferative cells.