Mechanical stress protection at the kidney filtration barrier

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Kidney function depends on the bulk filtration of large volumes of water and small solutes to clear potential toxins derived from cellular and gut microbial metabolism, and maintain salt and water and acid-base homeostasis in the organism. Remarkably, as much as 180 liters/day of glomerular filtration takes place in healthy adults. Glomerular filtration is driven by a hydrostatic pressure gradient of about 40 mmHg. This remarkable lumenal pressure exerts physical forces on the capillary wall that are counteracted by podocytes, specialized terminally differentiated epithelial cells. These cells enwrap the glomerular capillaries with interdigitating primary and secondary processes and are connected by a specialized membrane-like cell junction called the slit diaphragm. The complex podocyte architecture depends on a tightly regulated actin cytoskeletal machinery that enables them to adhere to the underlying glomerular basement membrane by cell-specific focal adhesion complexes and the slit diaphragm, which connects podocytes and contains mechanosensory proteins that sense mechanical forces exerted at the cell junctions. Mutations in either the slit diaphragm protein complex or the focal adhesion/cytoskeleton connection cause dysfunction of the filtration barrier and progressive renal disease in humans. In the first phase of this project, we used ultraresolution stimulated emission depletion (STED) microscopy in combination with mathematical modelling to understand the biophysical properties of kidney ultrafiltration (Butt et al., 2020). The model shows an active role of the podocyte in counteracting filtration pressure to sustain the filtration barrier. We demonstrated mechano-protective responses in podocytes to involve an adaptation of filamin expression and endocytic activity (Koehler et al., 2020) and proteolytic systems. Moreover, we showed that Bag3 and the chaperone-assisted selective autophagy (CASA) complex are enriched in podocytes. Bag3 localizes to the slit diaphragm suggesting that Bag3 and associated proteins may be involved in mechanoprotection in podocytes. Moreover, we could show that Bag3 interacts with regulators of the actin cytoskeleton including Rho A, Dynamin 2 and Arpc2. Perturbation of Bag3 induced a functional alteration of filtration and a late on-set proteinuria in the mouse model. In the next phase of the project we will now (1) study the role of regulators of chaperone machineries and degradation pathways mediating resistance to mechanical stress ,(2) characterize metabolic signalling pathways regulating mechanosensitivity and mechanical stress protection in podocytes, and (3) understand the relevance of stress protection systems for the maintenance of an intact renal filtration barrier in vivo.