SMARCA4 regulates spatially restricted metabolic plasticity in 3D multicellular tissue

Abstract

SWI/SNF and related chromatin remodeling complexes act as tissue-specific tumor suppressors and are frequently inactivated in different cancers. Although many regulatory activities of SWI/SNF have been identified using 2D cell culture, the effects of SWI/SNF alterations in more complex 3D tissues have remained poorly understood. Here we employed 3D cell culture conditions that yield transcriptomic states mirroring primary lung adenocarcinoma (LUAD) specimens better than 2D culture. By analyzing spatial patterns of gene expression and DNA accessibility in 3D spheroids using single-cell RNA-seq and ATAC-seq, we find that the SWI/SNF ATPase SMARCA4 (BRG1) induces state-specific changes to DNA accessibility that influence spatially heterogeneous expression patterns and metabolism. In 3D conditions, SMARCA4 promotes accessibility for AP-1 transcription factors, including ATF3, a regulator of metabolism and repressor of NRF2 antioxidant signaling. These changes reduce expression of SLC7A11 in a distinct portion of cells, which sensitizes A549 spheroids to cell death via ferroptosis under oxidizing conditions. Consistent with these results, we find that SMARCA4 alterations are associated with derepression of NRF2 targets in human tumors independently of NRF2/KEAP1 status. Our work reveals new 3D-specific features and unanticipated spatial complexity associated with chromatin remodeling in multicellular tissues.

Pathological implications of metabolic reprogramming and its therapeutic potential in medulloblastoma

“…Other than exporting glutamate outside the cell membrane, SLC7A11 imports cysteine, which is essential for GSH biosynthesis. Indeed, SMARCA4-dependent negative regulation of this transporter leads to reduction in intracellular cysteine required for antioxidant defense ( Koppula et al, 2018 ; Song et al, 2020 ; Cermakova et al, 2021 ). Thus, loss-of-function mutations in the SMARCA4 gene could drive MB survival under oxidative stress conditions.…”

Breast cancer spheroids can model phenotypic heterogeneity of cancer cells to evaluate targeted therapies.

“…There are also limitations to the spheroid approach. Cells grown in spheroids show reduced intra-tumor heterogeneity compared to the tumors from patients [19,22], due to a smaller tumor size, shorter growth time (days, not years) limiting the accumulation of genetic diversity, and differences in selection pressure which biases cells toward thriving in 2D culture prior to establishing spheroids [42]. The present experiments focused on cancer cells and did not include stromal nor immune cells.…”

“…In low oxygen and nutrient regions, we nd instead slowly dividing cancer stem cells [14], cells with increased mitochondrial content, hypoxic and senescent cells [17]. The local adaptation of cells to their microenvironmental niche is being extensively studied at the molecular level in patient tumor samples, and the lessons learned are being applied in personalized oncology [18,19].…”

“…The contribution of factors such as innate immunity, adaptive immunity, and physicochemical constraints to how cancer cells adapt to their microenvironment can be isolated and studied individually [15,21]. Even though growth in vitro cannot exactly recapitulate the in vivo context, cancer cells grown as 3D spheroids are a more faithful representation of cancer cell behavior and patient tumors than cells grown in a monolayer [19,20,22].…”

“…The complexity of spheroids ranges from monoculture on low adhesion plates to co-culture embedded in extracellular matrices [23,24]. In these structures, cells acquire different phenotypes -depending on their location -similarly to the ones found in tumors, while carrying minimal genetic differences [19,22,25]. The architecture of a single spheroid can be divided into different regions based on the nutrient availability within the spheroid: a necrotic zone at the core caused by the limited distribution of oxygen, nutrients, and metabolites, an intermediate quiescent zone, and an external zone consisting of proliferating cells.…”