Project Details
Projekt
Dissecting regulatory programs of kidney disease with functional genomics
Subject Area
Nephrology
Term
since 2021
Project identifier
Deutsche Forschungsgemeinschaft (DFG) - Project number 445703531
In the previous funding period, we identified kidney disease relevant gene regulatory networks using mouse models and organoids via single-cell sequencing (RNA and ATAC) and developed novel computational approaches. In the first study, we used single-cell ATAC sequencing in a kidney disease model to identify transcription factors (TFs) controlling myofibroblast differentiation, which we validated in vitro models using CRISPR-gene knockouts. In a second study, we used single-cell multi-omics (RNA and ATAC) to characterize the cellular differentiation process in kidney organoids. This allowed us to infer the gene regulatory networks driving organoid differentiation, which included lineages producing proximal tubular cells and podocyte, as well as off-target lineages related to neuronal and muscle cells. We are currently perturbing predicted transcription factors, i.e., of off-target lineages, to improve organoid growth and cellular differentiation. These findings emphasize the potential to identify important TFs, which play a role in the different cell-types involved in kidney disease and fibrosis progression.Despite our progress in identifying TFs regulating disease phenotypes of kidney cells, spatial factors influencing the cellular processes associated with tubular regeneration and fibrosis initiation (e.g. clonal expansion) remain elusive at the tissue level. Based on our spatial sequencing experiments cell lineages and clonal relationships can however be inferred based on e.g. mitochondrial mutations or single-nucleotide variants, which can be used as unique “barcodes” of the cells. We hypothesize that the clonal architecture of tubular and fibroblast cells in disease progression will provide insights into molecular mechanisms of tubular regeneration and fibrosis. To tackle this, we will combine our complementary expertise in nephrology and single-cell sequencing (Kuppe) and bioinformatics (Costa) to generate spatial multiomics datasets of human kidney tissues. We will use newly established spatial profiling methods, which will allow us to reconstruct lineage relationships between cells within kidney tissue to uncover their relationship. In WP1, we will use our platform to perform cell lineage tracing using spatially resolved sequencing for natural barcodes in human tissues. In WP2 we will use a specific transgenic mouse model (DARLIN) to decode tubular regeneration in CKD models using CRISRP-Cas9 barcodes. In WP3, we will develop novel computational approaches that leverage information across distinct modalities (spatial, molecular, histology and clonotypes) over time to predict molecular mechanisms related to tubular regeneration and myofibroblast differentiation.In conclusion, this project will dissect the impact of spatial organization and clonal architecture into tubular regeneration and fibrosis of the human kidney. This will improve our understanding in regulatory mechanisms relevant for human kidney disease progression.
DFG Programme
Clinical Research Units
Subproject of
KFO 5011:
Integrating emerging methods to advance translational kidney research (InteraKD)
Co-Investigator
Professor Dr. Matthias Saar
