Endocytosis provides a major alternative pathway for lysosomal biogenesis in kidney proximal tubular cells.
Recruitment of acid hydrolases to lysosomes generally occurs by intracellular sorting based on recognition of a common mannose 6-phosphate signal in the transGolgi network and selective transport to late endosomes/lysosomes. Here we provide evidence for an alternative, efficient secretion-recapture pathway mediated by megalin and exemplified by cathepsin B in kidney proximal convoluted tubules (PCT). We found that in mouse kidneys with defective megalin expression [megalin knockout (KO)] or apical PCT trafficking (ClC-5 KO), the (pro)cathepsin B mRNA level was essentially preserved, but the protein content was greatly decreased and the enzyme was excreted in the urine as mannose 6-phosphate-devoid species. In polarized PCT-derived cells, purified cathepsin B was avidly and selectively taken up at the apical membrane, and uptake was abolished by the megalin competitor, receptor-associated protein. Direct interaction of cathepsin B with megalin was demonstrated by surface plasmon resonance. Procathepsin B was detected in normal mouse serum. Purified cathepsin B injected into mice was efficiently taken up by kidneys (approximately 10% of injection) and targeted to lysosomes where it remained active, as shown by autoradiography and subcellular fractionation. A single cathepsin B injection into cathepsin B KO mice could reconstitute full lysosomal enzyme activity in the kidneys. These findings demonstrate a pathway whereby circulating lysosomal enzymes are continuously filtered in glomeruli, reabsorbed by megalin-mediated endocytosis, and transferred into lysosomes to exert their function, providing a major source of enzymes to PCT. These results also extend the significance of megalin in PCT and have several physiopathological and clinical implications.
Nielsen
Rikke
R
Cell Biology, Institute of Anatomy, University of Aarhus, 8000 Aarhus, Denmark.
Courtoy
Pierre J
PJ
Jacobsen
Christian
C
Dom
Geneviève
G
Lima
Wânia Rezende
WR
Jadot
Michel
M
Willnow
Thomas E
TE
Devuyst
Olivier
O
Christensen
Erik I
EI
D000818:Animals; D002401:Cathepsin B / Q000096:biosynthesis / Q000737:chemistry* / Q000494:pharmacology / Q000652:urine; D004705:Endocytosis; D004792:Enzyme Precursors / Q000737:chemistry*; D007668:Kidney / Q000166:cytology* / Q000378:metabolism; D007687:Kidney Tubules, Proximal / Q000166:cytology; D008024:Ligands; D026561:Low Density Lipoprotein Receptor-Related Protein-2 / Q000096:biosynthesis; D008247:Lysosomes / Q000378:metabolism*; D008360:Mannosephosphates / Q000737:chemistry; D051379:Mice; D018345:Mice, Knockout; D021381:Protein Transport; D015398:Signal Transduction; D020349:Surface Plasmon Resonance
D004792:Enzyme Precursors; D008024:Ligands; D026561:Low Density Lipoprotein Receptor-Related Protein-2; D008360:Mannosephosphates; C027693:mannose-6-phosphate; C057832:procathepsin B; D002401:Cathepsin B