Modelling CubAm function and regulation in proximal tubular cells using iPSC-derived kidney organoids.

Idiopathic nephrotic syndrome (INS) associated to focal segmental glomeruloesclerosis or minimal change disease is characterized by the presence of heavy levels of proteinuria. Filtrated proteins are normally actively reabsorbed in the proximal tubule by the megalin-cubilin-amnionless complex, located at the apical membrane of the proximal tubule epithelial cells. Megalin has a transmembrane domain but cubilin needs to interact with amnionless to reach the cell membrane in a complex known as CubAm. While megalin has a wide variety of ligands only a few proteins are exclusively transported by binding CubAm; one of those is Apolipoprotein A-I (ApoA-I). The reabsorption of Apolipoprotein A-I (ApoA-I) seems to be impaired in INS but this aspect of INS is difficult to study due to the low expression of CubAm in cultured proximal tubular cells as well as in other in vitro tubular modelling approaches. Here we show that RPTEC/TERT-1 cells cultured in monolayer barely express cubilin and amnionless proteins. In contrast, proximal tubular cells of induced Pluripotent Stem Cells (iPSCs)-derived kidney organoids showed robust expression of CubAm. In addition, protein overload induced an increase of the number of proximal tubular cells expressing cubilin that is reversed when the stimuli is removed. Finally, exogenously added ApoA-I targets cubilin suggesting that the CubAm complex is functional in our iPSCs-derived kidney organoids. Thus, kidney organoids provide a valuable system for modelling specific aspects of INS, including the impact of protein overload on tubular cells and the CubAm-mediated endocytosis of ApoA-I, which is thought to be impaired in these conditions.