Hyperphosphatemia is a common complication in patients with advanced chronic kidney disease (CKD) as well as an increased risk of cardiovascular mortality; however, the molecular mechanisms of phosphate-mediated kidney injury are largely unknown. Autophagy is a lysosomal degradation system, which plays protective roles against kidney diseases. Here, we studied the role of autophagy in kidney proximal tubular cells (PTECs) during phosphate overload. Temporal cessation of autophagy in drug-induced PTEC-specific autophagy-deficient mice that were fed high phosphate diet induced mild cytosolic swelling and an accumulation of SQSTM1/p62-and ubiquitin-positive protein aggregates in PTECs, indicating that phosphate overload requires enhanced autophagic activity for the degradation of increasing substrate. Morphological and biochemical analysis demonstrated that high phosphate activates mitophagy in PTECs in response to oxidative stress. PTEC-specific autophagy-deficient mice receiving heminephrectomy and autophagy-deficient cultured PTECs exhibited mitochondrial dysfunction, increased reactive oxygen species production, and reduced ATP production in response to phosphate overload, suggesting that high phosphate-induced autophagy counteracts mitochondrial injury and maintains cellular bioenergetics in PTECs. Thus, potentiating autophagic activity could be a therapeutic option for suppressing CKD progression during phosphate overload. (C) 2020 Elsevier Inc. All rights reserved.