Acidosis-sensing glutamine pump SNAT2 determines amino acid levels and mammalian target of rapamycin signalling to protein synthesis in L6 muscle cells

Wasting of lean tissue as a consequence of metabolic acidosis is a serious problem in patients with chronic renal failure. A possible contributor is inhibition by low pH of the System A (SNAT2) transporter, which carries the amino acid L-glutamine (L-Gln) into muscle cells. The aim of this study was to determine the effect of selective SNAT2 inhibition on intracellular amino acid profiles and amino acid–dependent signaling through mammalian target of rapamycin in L6 skeletal muscle cells. Inhibition of SNAT2 with the selective competitive substrate methylaminoisobutyrate, metabolic acidosis (pH 7.1), or silencing SNAT2 expression with small interfering RNA all depleted intracellular L-Gln. SNAT2 inhibition also indirectly depleted other amino acids whose intracellular concentrations are maintained by the L-Gln gradient across the plasma membrane, notably the anabolic amino acid L-leucine. Consequently, SNAT2 inhibition strongly impaired signaling through mammalian target of rapamycin to ribosomal protein S6 kinase, ribosomal protein S6, and 4E-BP1, leading to impairment of protein synthesis comparable with that induced by rapamycin. It is concluded that even though SNAT2 is only one of several L-Gln transporters in muscle, it may determine intracellular anabolic amino acid levels, regulating the amino acid signaling that affects protein mass, nucleotide/nucleic acid metabolism, and cell growth. Cachexia, the wasting of soft tissue, particularly skeletal muscle, is a frequent occurrence in patients with ESRD and is particularly severe in patients with diabetic nephropathy (1). It is a serious clinical problem because of its strong association with morbidity and mortality. An important cause is uremic metabolic acidosis (2), and there is good evidence that correction of acidosis decreases both weight loss and morbidity (3,4). Depletion of intramuscular free amino acids is thought to be an important early step in muscle wasting in uremia (5), and depletion is reversed