Abstract
Backgroundy+LAT1, encoded by SCL7A7, is the protein mutated in Lysinuric Protein Intolerance (LPI), a rare metabolic disease caused by a defective cationic amino acid (CAA, arginine, lysine, ornithine) transport at the basolateral membrane of intestinal and renal tubular cells. The disease is characterized by protein-rich food intolerance with secondary urea cycle disorder, but symptoms are heterogeneous with lung and immunological complications that are not explainable by the CAA transport defect. With the exception of the Finnish founder mutation (c.895-2A > T, LPIFin), LPI-causative mutations are heterogeneous and genotype-phenotype correlations have not been found. Here we addressed system y+L-mediated arginine uptake in monocytes from three LPI Italian patients and in lymphoblasts carrying the same mutations; in parallel, the genetic defects carried by the patients were reproduced as eGFP-tagged y+LAT1 mutants in transfected CHO cells to define the function and localization protein.ResultsSystem y+L activity is impaired in monocytes isolated from all LPI patients, and in CHO cells transfected with the three eGFP-y+LAT1 mutants, but not in lymphoblasts bearing the same mutations. The analysis of protein localization with confocal microscopy revealed that the eGFP-tagged mutants were retained inside the cytosol, with a pattern of expression quite heterogeneous among the mutants.ConclusionsThe three mutations studied of y+LAT1 transporter result in a defective arginine transport both in ex vivo (monocytes) and in vitro (CHO transfected cells) models, likely caused by the retention of the mutated proteins in the cytosol. The different effect of y+LAT1 mutation on arginine transport in monocytes and lymphoblasts is supposed to be due to the different expression of SLC7A7 mRNA in the two models, supporting the hypothesis that the impact of LPI defect largely depends on the relative abundance of LPI target gene in each cell type.
Highlights
Lysinuric protein intolerance (LPI; MIM 222700) is a recessively inherited autosomal disease caused by a defective cationic amino acid (CAA: arginine, lysine, ornithine) transport at the basolateral membrane of intestinal and renal tubular cells [1].From a clinical point of view, the disease manifests with protein-rich food intolerance and secondary urea cycle disorders; symptoms are heterogeneous, including failure to thrive, recurrent vomiting, hepatosplenomegaly, osteoporosis, lung involvement, kidney failure, The gene causing LPI, identified at the end of the nineties in SLC7A7, encodes for y+LAT1 protein [3, 4]
This feature of the transporter gains particular relevance when addressing the physiopathology of LPI and, of its classical hallmarks: normal-to-low plasma levels and leakage of cationic amino acids in the urine are, directly referable to both a defective intestinal absorption and an increased loss of the amino acids in the kidney consequent to SLC7A7/y+LAT1 mutation; clinical signs observed upon protein ingestion, such as nausea, vomiting, and, episodes of hyperammonemia can be explained by an impairment of urea cycle due to the deficiency of intermediates, mainly arginine, in the blood
System y+L activity in LPI monocytes and lymphoblasts Arginine transport through system y+L was measured in monocytes isolated from the three LPI patients enrolled and in three normal subjects age- and sex-matched
Summary
From a clinical point of view, the disease manifests with protein-rich food intolerance and secondary urea cycle disorders; symptoms are heterogeneous, including failure to thrive, recurrent vomiting, hepatosplenomegaly, osteoporosis, lung involvement, kidney failure, The gene causing LPI, identified at the end of the nineties in SLC7A7 (solute carrier family 7A member 7; MIM #603593), encodes for y+LAT1 protein [3, 4]. Little is known about the molecular and cellular mechanisms responsible for the life-threatening extra-renal complications of the disease, such as those affecting lung and immune system, we recently provided evidences sustaining a central role for the mononuclear phagocyte system [8,9,10]
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