Mechanisms of active renal repair: Single-cell trajectories of chromatin accessibility and gene expression changes in renal tubule cells

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Acute kidney injury (AKI), an abrupt loss of kidney function, is an increasing global health burden associated with high morbidity and mortality, for which no targeted therapy exits to date. Common causes of AKI are renal hypoperfusion, sepsis and nephrotoxicity. They result in an acute tubular injury and loss of kidney function, while simultaneously triggering the intrinsic repair processes of the kidney. Tubule cells that survive the acute insult re-enter the cell cycle and proliferate to regenerate the epithelial cells that are essential for the recovery of kidney function. Yet, surprisingly, tubule cells can also become maladaptive and contribute to the development of chronic kidney disease by continuous profibrotic signaling. The development of therapeutics to prevent such maladaptive processes is currently hampered by an insufficient understanding of the underlying molecular mechanisms. Therefore, the aim of the proposed project is to obtain a comprehensive understanding of the molecular underpinnings of renal repair after AKI at a single-cell resolution and to identify key regulatory factors governing adaptive and maladaptive repair processes. We will characterize both chromatin accessibility and associated gene expression of all nephron cells at various time points after both mild and severe AKI in transgenic mice, using a novel combination of single-nucleus ATAC- and single-nucleus RNA-sequencing. In addition, we will acquire the same data from maladaptive tubule cells specifically by isolating their nuclei based on the expression of a marker gene. Together, these data sets will provide a comprehensive regulatory map of the nephron at various stages after mild and severe AKI. We will further analyze these data using state-of-the-art bioinformatic tools to reconstruct the single-cell trajectories that lead to both maladaptive and adaptive tubule cells. From these trajectories, we can identify the pivotal regulatory factors determining both reparative and maladaptive repair processes. We will employ additional methods like RNAScope in situ hybridization and immunostaining of the kidney to study these regulatory factors in even greater detail. Ultimately, the goal of this study is to achieve a detailed understanding of renal repair processes that could facilitate targeted drug development to treat AKI in the future.