Distinct functions of megalin and cubilin receptors in recovery of normal and nephrotic levels of filtered albumin

Abstract

Proximal tubule (PT) cells express a single saturable albumin-binding site whose affinity matches the estimated tubular concentration of albumin; however, albumin uptake capacity is greatly increased under nephrotic conditions. Deciphering the individual contributions of megalin and cubilin to the uptake of normal and nephrotic levels of albumin is impossible in vivo, as knockout of megalin in mice globally disrupts PT endocytic uptake. We quantified concentration-dependent albumin uptake in an optimized opossum kidney cell culture model and fit the kinetic profiles to identify albumin-binding affinities and uptake capacities. Mathematical deconvolution fit best to a three-component model that included saturable high- and low-affinity uptake sites for albumin and underlying nonsaturable uptake consistent with passive uptake of albumin in the fluid phase. Knockdown of cubilin or its chaperone amnionless selectively reduced the binding capacity of the high-affinity site, whereas knockdown of megalin impacted the low-affinity site. Knockdown of disabled-2 decreased the capacities of both binding sites. Additionally, knockdown of megalin or disabled-2 profoundly inhibited the uptake of a fluid phase marker, with cubilin knockdown having a more modest effect. We propose a novel model for albumin retrieval along the PT in which cubilin and megalin receptors have different functions in recovering filtered albumin in proximal tubule cells. Cubilin binding to albumin is tuned to capture normally filtered levels of the protein. In contrast, megalin binding to albumin is of lower affinity, and its expression is also essential for enabling the recovery of high concentrations of albumin in the fluid phase.

Predicting the protein composition of human urine in normal and pathological states: Quantitative description based on Dent1 disease (CLCN5 mutation)

“…Differences in uptake kinetics could stem from dissimilar binding affinities to megalin and cubilin, from the differential expression of these receptors along the PT as observed in rodents (Schuh et al 2018;Clark et al 2019;Limbutara et al 2020) and/or from differences in trafficking kinetics. Recent observations in OK cells suggest that megalin and cubilin may function independently in different nephron segments, and that fluid phase endocytosis of albumin is megalin-dependent and may become significant under nephrotic conditions (Ren et al 2020). However, sufficient quantitative data are not available in humans to build a solid model based on these findings.…”

“…Recent observations in OK cells suggest that megalin and cubilin may function independently in different nephron segments, and that fluid phase endocytosis of albumin is megalin‐dependent and may become significant under nephrotic conditions (Ren et al . 2020). However, sufficient quantitative data are not available in humans to build a solid model based on these findings.…”

Making a Dent in Dent Disease

“…In addition to its global function in maintaining the PT apical endocytic pathway, recent studies suggest that megalin has distinct roles in regulating endocytic function in different PT nephron segments. 34 , 35 While loss of megalin clearly impairs endocytic uptake along the entire PT, a reduction in megalin expression may have different effects on endocytic pathway integrity within each subregion of this nephron segment (S1, S2, S3). Indeed, the evidence for megalin-dependent elaboration of the endocytic pathway is based on ultrastructural examination of a limited sample of undefined PT nephron segments.…”

“…Although most studies have monitored changes in megalin expression in DD, albumin and many other filtered ligands bind to cubilin rather than megalin. 35 , 36 Cubilin lacks a cytoplasmic domain and relies on association with the transmembrane protein amnionless for delivery to the plasma membrane. 37 Additionally, cubilin binds to megalin, and the two receptors have been generally assumed to traffic together as a complex.…”

Endocytic adaptation to functional demand by the kidney proximal tubule

“…Perhaps surprisingly, expression of megalin appears to be required for this reserve pathway to operate in the distal PT. Knockdown of megalin in OK cells reduces the capacity of the non-saturable component of albumin uptake (Ren et al 2020). Moreover, uptake of nephrotic concentrations of albumin in mice is dependent on megalin but not cubilin expression (Ren et al 2020).…”