Heptahelical protein PQLC2 is a lysosomal cationic amino acid exporter underlying the action of cysteamine in cystinosis therapy

Abstract

Cystinosin, the lysosomal cystine exporter defective in cystinosis, is the founding member of a family of heptahelical membrane proteins related to bacteriorhodopsin and characterized by a duplicated motif termed the PQ loop. PQ-loop proteins are more frequent in eukaryotes than in prokaryotes; except for cystinosin, their molecular function remains elusive. In this study, we report that three yeast PQ-loop proteins of unknown function, Ypq1, Ypq2, and Ypq3, localize to the vacuolar membrane and are involved in homeostasis of cationic amino acids (CAAs). We also show that PQLC2, a mammalian PQ-loop protein closely related to yeast Ypq proteins, localizes to lysosomes and catalyzes a robust, electrogenic transport that is selective for CAAs and strongly activated at low extracytosolic pH. Heterologous expression of PQLC2 at the yeast vacuole rescues the resistance phenotype of an ypq2 mutant to canavanine, a toxic analog of arginine efficiently transported by PQLC2. Finally, PQLC2 transports a lysine-like mixed disulfide that serves as a chemical intermediate in cysteamine therapy of cystinosis, and PQLC2 gene silencing trapped this intermediate in cystinotic cells. We conclude that PQLC2 and Ypq1–3 proteins are lysosomal/vacuolar exporters of CAAs and suggest that small-molecule transport is a conserved feature of the PQ-loop protein family, in agreement with its distant similarity to SWEET sugar transporters and to the mitochondrial pyruvate carrier. The elucidation of PQLC2 function may help improve cysteamine therapy. It may also clarify the origin of CAA abnormalities in Batten disease.

Synthesis of diacylated γ-glutamyl-cysteamine prodrugs, and in vitro evaluation of their cytotoxicity and intracellular delivery of cysteamine

“…Evidence for the involvement of this transporter in the cysteamine-driven depletion of cystine was obtained by its genetic inactivation or silencing [19,20]. Molecular modeling has shown the structural and electronic similarities of the L-cysteine-cysteamine mixed disulfide to L-lysine, facilitating its use of this transporter [21].…”

Vacuolar amino acid transporters upregulated by exogenous proline and involved in cellular localization of proline in <i>Saccharomyces cerevisiae</i>

“…Atg22 mediates the efflux of leucine and other amino acids resulting from autophagic degradation, and also redundantly functions with Avt3 and Avt4 (Yang et al, 2006). In the homeostasis of cationic amino acids, the vacuolar basic amino acid transporter (VBA)-family member proteins Vba1, Vba2, and Vba3 (Shimazu et al, 2005) and the PQ-loop proteins Ypq1, Ypq2, and Ypq3 (Jézégou et al, 2012) are specifically involved in the import and export of basic amino acids, respectively. Since proline is one of the nonpreferred nitrogen sources, the analysis of intracellular localization of proline has been difficult under typical nutrient conditions.…”

Loss of ATP-dependent lysine uptake in the vacuolar membrane vesicles of Saccharomyces cerevisiae ypq1∆ mutant

“…Jézégou et al 5) described in the previous paper that Ypq1p as well as Ypq2p is involved in export of basic amino acids from vacuoles, because ypq1Δ mutant acquired resistance to canavanine although it was weaker than ypq2Δ mutant. Accumulation of the drug into vacuoles might be accelerated in ypq1Δ mutant, resulting in decrease of its cytosolic level.…”

“…1(A)) although these functions were not investigated. Very recently, Jézégou et al 5) reported a new closest homolog YPQ2 (YDR352W) in S. cerevisiae ( Fig. 1(A)).…”

“…Ypq1p-GFP, Ypq2p-GFP, and Ypq3p-GFP were located at the vacuolar membrane when expressed in the sigma1278b strain background. 5) They found that ypq2Δ mutant acquired the resistance to canavanine (L-arginine analog). ypq1Δ mutant also displayed resistance to canavanine but to a lesser extent than ypq2Δ mutant.…”