The C-terminal domain of the neutral amino acid transporter SNAT2 regulates transport activity through voltage-dependent processes

Abstract

Sodium-coupled neutral amino acid transporter 2 (SNAT21) belongs to the SLC38 family of solute transporters. Transport of 1 amino acid molecule into the cell is driven by the co-transport of 1 Na+ ion. The functional significance of the C-terminus of SNAT2, which is predicted to be located in the extracellular space, is currently unknown. Here, we removed 13 amino acid residues from the SNAT2 C-terminus and studied the effect of the deletion on transporter function. The truncation abolished amino acid transport currents at negative membrane potentials (< 0 mV), as well as substrate uptake. However, transport currents were observed at positive membrane potentials, demonstrating that transport was accelerated while the driving force decreased. Membrane expression levels were normal in the truncated transporter. SNAT2Del C-ter showed 3-fold higher apparent affinity for alanine, and 2-fold higher Na+ affinity compared to SNAT2WT, suggesting that the C-terminus is not required for high-affinity substrate and Na+ interaction with SNAT2. pH sensitivity of amino acid transport was partially retained after the truncation. In contrast to the truncation after the final trans-membrane domain, TM11, deletion of TM11 resulted in an inactive transporter, most likely due to a defect in cell surface expression. Together, the results demonstrate that the C-terminal domain of SNAT2 is an important voltage regulator that is required for a normal amino acid translocation process at physiological membrane potentials. However, the C-terminus appears not to be involved in regulation of membrane expression.

Membrane Topological Structure of Neutral System N/A Amino Acid Transporter 4 (SNAT4) Protein

“…The extracellular orientation of C termini of SNAT family proteins have previously been determined both experimentally (35) and by computer simulation (9,17,36), which is consistent with our results. However, based primarily on theoretical prediction, but not on experimental data, previous published studies, including those from our laboratory, suggest an intracellular orientation of N termini for SNAT family proteins (10,11,17,36,37).…”

“…However, based primarily on theoretical prediction, but not on experimental data, previous published studies, including those from our laboratory, suggest an intracellular orientation of N termini for SNAT family proteins (10,11,17,36,37). Based on the prediction by HHpred, we noticed that SNAT family proteins share a certain degree of sequential homologies with plant auxin permease, auxin1, and bacterial transporters, AdiC and Mhp1.…”

An Arginine Deprivation Response Pathway Is Induced in Leishmania during Macrophage Invasion

“…SNAT2 is a mammalian transceptor that transports neutral amino acid and senses amino acid availability [ 36 ]. The long soluble N-terminus of SNAT2 senses intracellular amino acid content thereby stabilizes the amino acid pool, while the short extracellular C-terminus regulates transport activity [ 42 ]. In contrast to the yeast and mammalian transceptors, which have low specificity and respond to amino acid availability, the Leishmania arginine sensor responds only to arginine availability.…”

Combined walking exercise and alkali therapy in patients with CKD4–5 regulates intramuscular free amino acid pools and ubiquitin E3 ligase expression

“…2004), is strongly inhibited by acidosis (Baird et al 2006;Evans et al 2007;Evans et al 2008) and inhibited in CKD (Asola et al 2001). No change was seen in SNAT2 expression (Table 6), but studies in vitro have shown that acidosis directly inhibits this transporter protein (Baird et al 2006;Zhang et al 2011) independent of gene expression (Evans et al 2007;Evans et al 2008). However this inhibition by acid has previously only been studied acutely, and the corresponding chronic effects of acid on this transporter in the context of exercise and mechanical stress merit further investigation.…”