title: Role of Arf1 in endosomal trafficking of protein-metal complexes and cadmium-metallothionein-1 toxicity in kidney proximal tubule cells.
abstract: Cadmium (Cd) is nephrotoxic. Circulating Cd-metallothionein complexes (CdMT) are filtered by the kidney, reabsorbed by proximal tubule cells (PTC) via receptor-mediated endocytosis, and trafficked to lysosomes which results in apoptosis. ADP-ribosylation factors (Arfs) regulate vesicular trafficking. Arf1 is traditionally associated with the secretory pathway, but has been recently found involved in endocytotic trafficking in PTC. Hence, the role of Arf1 was investigated in MT-1 and transferrin (Tf) endocytosis, and in CdMT-1-induced cell death in a PTC line by overexpressing Arf1-wildtype (WT) or dominant-negative mutant Arf1-T31N. Endogenous Arf1 distribution in PTC was punctate throughout the cytosol, but was not detected in the plasma membrane. Arf1 colocalized with markers for sorting to late endosomes (Rab7, CLC6). Arf1 weakly overlapped with the late endosomal/lysosomal marker CLC7, but not with markers for early (Rab5, CLC5) and recycling endosomes (Rab11). Arf1-T31N significantly attenuated CdMT-1 toxicity by ∼60% when compared to Arf1-WT. However, overexpression of Arf1-T31N did not prevent internalization of Alexa Fluor 546-coupled Tf or MT-1 which accumulated in an EEA1-positive early endocytotic compartment, but not in Arf1-WT overexpressing cells. We conclude that Arf1 is involved in trafficking of protein-metal complexes, including CdMT, to late endosomes/lysosomes in renal PTC.
authors.affiliation: Institute of Physiology & Pathophysiology, University of Witten/Herdecke, Stockumer Strasse 12, D-58453 Witten, Germany.
authors.forename: Natascha A
authors.initials: NA
authors.lastname: Wolff
authors.forename: Wing-Kee
authors.initials: WK
authors.lastname: Lee
authors.forename: Frank
authors.initials: F
authors.lastname: Thévenod
mesh_terms: D020823:ADP-Ribosylation Factor 1 / Q000032:analysis / Q000502:physiology*; D000818:Animals; D001692:Biological Transport; D002478:Cells, Cultured; D004705:Endocytosis; D011992:Endosomes / Q000378:metabolism*; D007687:Kidney Tubules, Proximal / Q000187:drug effects*; D008668:Metallothionein / Q000378:metabolism / Q000633:toxicity*; D021381:Protein Transport; D051381:Rats; D014168:Transferrin / Q000378:metabolism; D033921:Vesicular Transport Proteins / Q000032:analysis; D020691:rab GTP-Binding Proteins / Q000032:analysis; D000090802:rab7 GTP-Binding Proteins
chemical_list: C491055:Mt2A protein, rat; D014168:Transferrin; D033921:Vesicular Transport Proteins; C116509:cadmium-metallothionein complex; C093846:early endosome antigen 1; D000090802:rab7 GTP-Binding Proteins; C000716466:rab7 GTP-binding proteins, rat; D008668:Metallothionein; D020823:ADP-Ribosylation Factor 1; D020691:rab GTP-Binding Proteins