Nuclear cholesterol regulates nuclear size and DNA damage responses in the tumor hierarchy

Normal stem cells undergo morphologic, epigenetic, and metabolic changes during cellular differentiation. Here, we report that glioblastoma stem cells (GSCs) display smaller nuclear size than differentiated progeny. Leveraging differential gene expression and proteomics to discover potential mediators of nuclear size regulation, we found that GSCs are enriched for a nuclear lamina-localized sterol reductase, Lamin B Receptor (LBR). LBR bridges two functions, genome organization and cholesterol synthesis, of which nuclear cholesterol synthesis mediates LBR regulation of GSC nuclear size. Targeting LBR increased nuclear size and decreased GSC viability and tumor initiation. Mass spectrometry defined an LBR interactome in GSCs with mediators of DNA damage repair (DDR) and R-loop resolution, including DEAD-Box Helicase 5 (DDX5). Consistently, targeting LBR induced double-strand DNA breaks (DSBs) and P53-dependent DDR. Genetic LBR loss reduced DDX5 expression and increased R-loops; pharmacologic LBR sterol reductase inhibition phenocopied LBR loss, increasing R-loops and DSBs with synergy with irradiation to inhibit GSC survival. Together, we propose that LBR regulates nuclear cholesterol synthesis, which contributes to the small GSC nuclear sizes and promotes efficient DNA damage repair. Overall design: To understand mechanims underlying LBR requirements in GSCs, we profiled gene expression upon LBR loss in two independent GSC lines, each with two independent shRNAs targeting LBR