Project Details
Projekt
Searching for the signature and causes of muscular dystrophies associated with nuclear envelope dysfunction using single-cell multiomics
Applicants
Dr. Michael Robson; Professor Dr. Werner Stenzel
Subject Area
Human Genetics
Molecular and Cellular Neurology and Neuropathology
Cell Biology
Molecular and Cellular Neurology and Neuropathology
Cell Biology
Term
since 2023
Project identifier
Deutsche Forschungsgemeinschaft (DFG) - Project number 400728090
Mutations in nuclear envelope proteins cause diverse diseases ranging from muscular dystrophy to progeria but their underlying pathomechanisms remain unclear. Many of these disease-associated proteins mechanically attach chromatin to the nuclear envelope in so-called “lamina-associated domains” (LADs) that support cell type-specific gene silencing. Consequently, it has long been suggested that nuclear envelope linked pathologies are fundamentally gene mis-expression diseases driven by disrupted 3D genome organization. However, the inability to map LADs or directly relate their perturbation to altered gene regulation in single cells and diseased tissues has prevented examining this hypothesis. Here, we will address this question by generating two physiological models of the Emery-Dreifuss Muscular Dystrophy (EDMD) in iPSC-derived neuromuscular organoids. We will then examine these models with a new method that combines the protein-DNA interaction profiling of DNA adenine methyltransferase identification (DamID) with mRNA sequencing in the same cell (scDam&T) in vivo. scDam&T in vivo will allow us to simultaneously map LAD and transcriptional dynamics in single cells from any complex tissue. By combining scDam&T and single cell chromatin accessibility profiling (scATAC&T), we will comprehensively characterize the molecular events that drive cell-fate decisions during normal neuromuscular development. We will then describe the cellular and molecular signature of EDMD in mutant neuromuscular organoids using both histological studies, 2D and 3D electron microscopy, and single cell multimodality. We will map changes in cell composition, clarify which heterochromatin and LADs are disrupted, and determine its consequences for gene transcription. Finally, we will use electron and immunofluorescence-FISH microscopy to investigate whether these disruptions are actually detectable in muscle biopsy samples from EDMD patients. This study has the potential to establish new disease models of congenital muscular dystrophies, in this case EDMD. With these, we will thus reveal the first pathophysiologically important links between nuclear envelope-directed genome organization and possible gene dysregulation as a cause of disease.
DFG Programme
Research Units
