Three-dimensional regulatory hubs support oncogenic programs in glioblastoma [scRNA-seq]
Enhancer-promoter communication in the context of the three-dimensional (3D) nucleus is critical for the regulation of transcriptional programs specific for cell identity. Here, we profile the 3D enhancer-promoter networks of primary patient-derived human glioma spheres (GSCs) in comparison with neuronal stem cells to identify candidate central nodes of GSC tumorigenic programs. We have identified highly interacting 3D enhancer-promoter “hubs” which strongly associate with robust and coordinated expression of tumor-associated genes, suggesting potentially central roles in the regulation of oncogenic programs. Integration of whole-genome sequencing (WGS) data from The Cancer Genome Atlas showed that 3D regulatory hubs are strongly enriched for recurrent point mutations in the larger cohort of glioblastoma patients. In a proof-of-concept experiment, epigenetic perturbation of a highly interacting and recurrent 3D enhancer hub in the GOLIM4 locus led to significant downregulation of multiple hub-connected genes and reduced clonogenicity along with a global shift in the transcriptional states. Finally, integration of published H3K27ac HiChIP from other cancer types enabled the identification of both “universal” and cancer type-specific 3D regulatory hubs which enrich for different oncogenic programs and potential regulators, and often associate with a worse prognosis. Overall, our study identifies complex 3D regulatory hubs in glioblastoma and provides computational and experimental support for their potential role in driving oncogenic programs and promoting tumorigenic properties. Overall design: Glioma stem cell lines expressing doxycycline-inducible dCas9-KRAB-GFP with guides targeting a negative control region and the GOLIM4 enhancer were co-cultured with 4.5 month old brain cerebral organoids (GLICO). After co-culture and doxycycline induction for 7 days, cells were harvested, sorted as single cells for GFP+/DAPI- cells and submitted for scRNA-seq (10X Genomics)