Abstract
The ATP6ap2 (Pro)renin receptor protein associates with H+-ATPases which regulate organellar, cellular, and systemic acid–base homeostasis. In the kidney, ATP6ap2 colocalizes with H+-ATPases in various cell types including the cells of the proximal tubule. There, H+-ATPases are involved in receptor-mediated endocytosis of low molecular weight proteins via the megalin/cubilin receptors. To study ATP6ap2 function in the proximal tubule, we used an inducible shRNA Atp6ap2 knockdown rat model (Kd) and an inducible kidney-specific Atp6ap2 knockout mouse model. Both animal lines showed higher proteinuria with elevated albumin, vitamin D binding protein, and procathepsin B in urine. Endocytosis of an injected fluid-phase marker (FITC- dextran, 10 kDa) was normal whereas processing of recombinant transferrin, a marker for receptor-mediated endocytosis, to lysosomes was delayed. While megalin and cubilin expression was unchanged, abundance of several subunits of the H+-ATPase involved in receptor-mediated endocytosis was reduced. Lysosomal integrity and H+-ATPase function are associated with mTOR signaling. In ATP6ap2, KO mice mTOR and phospho-mTOR appeared normal but increased abundance of the LC3-B subunit of the autophagosome was observed suggesting a more generalized impairment of lysosomal function in the absence of ATP6ap2. Hence, our data suggests a role for ATP6ap2 for proximal tubule function in the kidney with a defect in receptor-mediated endocytosis in mice and rats.
Highlights
Therenin receptor ((P)RR) was identified as the receptor forrenin and renin [45, 67]
Transgenic rats (Sh rats) expressing an shRNA against Atp6ap2 were generated by inserting a transgene under the control of tetracycline repressor protein (TetR) allowing the controlled activation of the shRNA by doxycycline treatment (Fig. 1A)
RT-qPCR for Atp6ap2 mRNA in the whole kidney showed a decrease by approx. 90% of the transcript in Sh rats when compared with control rats (Wt) (Fig. 1D)
Summary
The (pro)renin receptor ((P)RR) was identified as the receptor for (pro)renin and renin [45, 67]. The importance of H+-ATPases in endocytosis has been further confirmed in studies in mice lacking the V0 domain H+-ATPase a4 subunit in the kidney [22, 48] These animals presented a generalized impairment of proximal tubule functions with glycosuria, phosphaturia, and lysosomal defects. Deletion of ATP6ap impaired Wnt signaling by reducing endosomal recycling of frizzled-related receptors, and reduced fluid-phase endocytosis, both functions linked to reduced H +-ATPase activity [25]. Efficient acidification of the endocytic pathway is critical for recycling and processing of proteins, activation of lysosomal enzymes and transporters, and lysosomal signaling via the mammalian target of rapamycin (mTOR) pathway [40, 65, 68] Several of these processes have been shown to be H+-ATPase dependent [34, 37]. We report that ATP6ap plays a role in cargo reabsorption and processing via receptor-mediated endocytosis, contributing to kidney proximal tubule function
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