Integrated multi-omics profiling identifies TTYH1 as an astrocyte-associated tumor suppressor in glioma

The complex cellular interactions and transcriptional networks that drive gliomas have not yet been fully elucidated. This study aimed to characterize the glioma microenvironment at single-cell resolution and identify the key molecular drivers of the pathogenesis and prognoses of glioma. We performed single-cell RNA sequencing (scRNA-seq) on 11 glioma samples and single-cell ATAC sequencing (scATAC-seq) on 1 glioma sample, followed by integrated multi-omics analysis. The cellular composition and interactions were determined using Seurat, CellChat and Monocle. Spatial transcriptomics of 2 high-grade glioma samples was used to validate the spatial localizations and interactions. The functional validation of TTYH1 was performed in vitro using glioma cell lines. scRNA-seq delineated nine major cell types, with astrocytes emerging as the central communication hubs. Integrated scRNA-seq and scATAC-seq identified 72 astrocyte-specific marker genes. Among these, TTYH1 was prioritized as a top prognostic candidate based on a random survival forest analysis. Elevated TTYH1 expression was significantly associated with a favorable prognosis, IDH mutations, and lower tumor grade in independent CGGA cohorts. Spatial transcriptomics confirmed the localization of key cell types and revealed interactions between astrocytes, tumor-associated fibroblasts, and endothelial cells. In vitro, TTYH1 overexpression suppressed the proliferation and migration of U251 and LN229 cells. In vivo experiments further validated that TTYH1 overexpression impeded tumor growth, while TTYH1 knockdown facilitated tumor expansion. Mechanistically, TTYH1 acted as a tumor suppressor through a process involving Notch signaling to inhibit cell proliferation and migration. Our study provides a comprehensive multi-omics landscape of the glioma microenvironment. We identified TTYH1 as a novel astrocyte-associated tumor suppressor gene that impedes glioma progression, supporting its role as a candidate prognostic biomarker and a potential therapeutic target that requires further validation.