MDC filed the patent application EP18192715 based on the results of this study and GG is listed as an inventor. All other authors declare no competing interest.
Epithelial-mesenchymal transition (EMT) is a developmental process hijacked by cancer cells to modulate proliferation, migration, and stress response. Whereas kinase signaling is believed to be an EMT driver, the molecular mechanisms underlying epithelial-mesenchymal interconversion are incompletely understood. Here, we show that the impact of chromatin regulators on EMT interconversion is broader than that of kinases. By combining pharmacological modulation of EMT, synthetic genetic tracing, and CRISPR interference screens, we uncovered a minority of kinases and several chromatin remodelers, writers, and readers governing homeostatic EMT in lung cancer cells. Loss of ARID1A, DOT1L, BRD2, and ZMYND8 had nondeterministic and sometimes opposite consequences on epithelial-mesenchymal interconversion. Together with RNAPII and AP-1, these antagonistic gatekeepers control chromatin of active enhancers, including pan-cancer-EMT signature genes enabling supraclassification of anatomically diverse tumors. Thus, our data uncover general principles underlying transcriptional control of cancer cell plasticity and offer a platform to systematically explore chromatin regulators in tumor-state–specific therapy.
The molecular basis underlying glioblastoma (GBM) heterogeneity and plasticity is not fully understood. Using transcriptomic data of human patient-derived brain tumor stem cell lines (BTSCs), classified based on GBM-intrinsic signatures, we identify the AP-1 transcription factor FOSL1 as a key regulator of the mesenchymal (MES) subtype. We provide a mechanistic basis to the role of the neurofibromatosis type 1 gene (NF1), a negative regulator of the RAS/MAPK pathway, in GBM mesenchymal transformation through the modulation of FOSL1 expression. Depletion of FOSL1 in NF1-mutant human BTSCs and Kras-mutant mouse neural stem cells results in loss of the mesenchymal gene signature and reduction in stem cell properties and in vivo tumorigenic potential. Our data demonstrate that FOSL1 controls GBM plasticity and aggressiveness in response to NF1 alterations.
Glioblastoma is a lethal brain tumor with a high degree of heterogeneity and resistance to therapy. The lack of genetic tracing approaches to selectively identify tumor states and fate transitions limited our understanding of tumor homeostasis. By translating glioblastoma subtype signatures into synthetic genetic tracing cassettes, we investigated tumor heterogeneity at cellular and molecular level, in vitro and in vivo. Synthetic genetic tracing demonstrated that proneural glioblastoma is a hardwired identity, whereas the mesenchymal glioblastoma is an adaptive and metastable state downstream pro-inflammatory and differentiation cues, similarly wired in breast and lung tumors. Importantly, we discovered that innate immune cells divert glioblastoma cells into a proneural-to-mesenchymal transition causal to therapeutic resistance. Systematic phenotypic mapping using synthetic locus control regions (sLCRs) is a simple, automated, and scalable strategy for genetic tracing, generally applicable to study developmental and disease homeostasis. In glioblastoma, it uncovered causality between various (micro)environmental, genetic and pharmacological perturbations and mesenchymal commitment. Citation Format: Matthias Jürgen Schmitt, Carlos Company, Yuliia Dramaretska, Iros Barozzi, Andreas Göhrig, Sonia Kertalli, Melanie Großmann, Heike Naumann, Jikke Wierikx, Danielle Hulsman, Rainer Glass, Massimo Squatrito, Michela Serresi, Gaetano Gargiulo. Phenotypic mapping of pathological crosstalk between glioblastoma and innate immune cells by synthetic genetic tracing [abstract]. In: Proceedings of the AACR Virtual Special Conference on Tumor Heterogeneity: From Single Cells to Clinical Impact; 2020 Sep 17-18. Philadelphia (PA): AACR; Cancer Res 2020;80(21 Suppl):Abstract nr PO-104.
Descriptive data are rapidly expanding in biomedical research. Instead, functional validation methods with sufficient complexity remain underdeveloped. Transcriptional reporters allow experimental characterization and manipulation of developmental and disease cell states, but their design lacks flexibility. Here, we report logical design of synthetic cis-regulatory DNA(LSD), a computational framework leveraging phenotypic biomarkers and trans- regulatory networks as input to design reporters marking the activity of selected cellular states and pathways. LSD uses bulk or single-cell biomarkers and a reference genome or custom cis- regulatory DNA datasets with user-defined boundary regions. By benchmarking validated reporters, we integrated LSD with a computational classifier to rank phenotypic specificity of putative cis-regulatory DNA. Experimentally, LSD-designed reporters targeting a wide range of cell states are functional without minimal promoters. In silico, an LSD-unsupervised mesenchymal glioblastoma reporter outperformed previously validated ones. In genome- scale CRISPRa screens, it discovered known and novel bona fide cell-state-drivers. Thus, LSD captures core principles of cis-regulation and is broadly applicable to studying complex cell states and mechanisms of transcriptional regulation.
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