The kidney regulates plasma protein levels by eliminating them from the circulation. Proteins filtered by glomeruli are endocytosed and degraded in the proximal tubule and defects in this process result in tubular proteinuria, an important clinical biomarker. However, the spatiotemporal organization of renal protein metabolism in vivo was previously unclear. Here, using functional probes and intravital microscopy, we track the fate of filtered proteins in real time in living mice, and map specialized processing to tubular structures with singular value decomposition analysis and three-dimensional electron microscopy. We reveal that degradation of proteins requires sequential, coordinated activity of distinct tubular sub-segments, each adapted to specific tasks. Moreover, we leverage this approach to pinpoint the nature of endo-lysosomal disorders in disease models, and show that compensatory uptake in later regions of the proximal tubule limits urinary protein loss. This means that measurement of proteinuria likely underestimates severity of endocytotic defects in patients.

Zinc is the second most prevalent metal element present in living organisms, and control of its concentration is pivotal to physiology. The amount of zinc available to the cell cytoplasm is regulated by the activity of members of the SLC39 family, the ZIP proteins. Selectivity of ZIP transporters has been the focus of earlier studies which provided a biochemical and structural basis for the selectivity for zinc over other metals such as copper, iron, and manganese. However, several previous studies have shown how certain ZIP proteins exhibit higher selectivity for metal elements other than zinc. Sequence similarities suggest an evolutionary basis for the elemental selectivity within the ZIP family. Here, by engineering HEK293 cells to overexpress ZIP proteins, we have studied the selectivity of two phylogenetic clades of ZIP proteins, that is ZIP8/ZIP14 (previously known to be iron and manganese transporters) and ZIP5/ZIP10. By incubating ZIP over-expressing cells in presence of several divalent metals, we found that ZIP5 and ZIP10 are high affinity copper transporters with greater selectivity over other elements, revealing a novel substrate signature for the ZIP5/ZIP10 clade.

The kidney proximal tubule is a highly specialized epithelium that transports metabolites and maintains body homeostasis. Cells lining this nephron segment are densely packed with lysosomes, but little is known about the dynamic activity of these organelles in situ. Here, using targeted sensors and live cell and intravital imaging we track acidified lysosomes along the mouse proximal tubule and uncover marked axial heterogeneity in their distribution, characteristics and organellar interactions. In the early part, cathepsin-rich lysosomes frequently contact with apical endosomes to receive and catabolize filtered plasma proteins. Conversely, in the later region, lipase-containing lysosomes traverse cells to mobilize and degrade mitochondria-associated lipid droplets and facilitate their extrusion into the tubular lumen. Acutely de-acidifying lysosomes dramatically alters their movement, causing major changes in tubular protein and lipid processing. Thus, lysosomes in proximal tubules are highly dynamic and adapted to perform distinct metabolic tasks within different specialized segments.