SN2 and SAT2 (ATA2) are isoforms of structurally related amino acid (AA) transporters effecting System N and A transport activity, respectively (Yao et al. 2000; Nakanishi et al. 2001). Such Na⁺-dependent transporters classically mediate net concentrative uptake of AA substrate into the cells. We are investigating the suggestion (Chaudhry et al. 1999) that System N transporters may also be involved in cellular release of substrates (notably glutamine; Gln).

Rat SN2 and SAT2 transporters were studied by overexpression in Xenopus laevis oocytes. Transporter cRNAs (50 ng) were injected into oocytes and experiments were performed 2-3 days later. [³H] Gln uptake studies demonstrated similar levels of functional expression for SN2 and SAT2; both have Gln transport K_m values of the order of 1-2 mM. For efflux assays, 50 nl (1.85 kBq) of [³H] Gln were injected into oocytes. After a 30 min recovery period, tracer appearance from oocytes was monitored at 10 min intervals.

The rate constant for efflux (K_e) of [³H] Gln was 6.4 ± 2.2 times higher for SN2-expressing oocytes than for both SAT2-expressing cells and water-injected controls (control value was 0.0018 ± 0.0006 min^-1; n = 6 oocyte batches); see Fig. 1. Efflux of AA tracer substrate through SN2 was saturable, being suppressed by co-injection of excess unlabelled substrate. Reversal of the transmembrane Na⁺ gradient by incubating cells in Na⁺-free buffer increased K_e by 66 % in SN2-expressing oocytes, but had no effect on AA efflux from SAT2-expressing or control oocytes.

The results reveal that SN2 produces saturable, bidirectional movements of Gln across the plasma membrane. Gln efflux through SN2 is influenced by changes in electrochemical gradient of Na⁺ co-substrate. In contrast, SAT2 does not exhibit measurable glutamine efflux under our experimental conditions, although Gln influx characteristics are similar to those of SN2. Such differences in transport properties may have important consequences for intercellular Gln movements within tissues such as brain and liver.

This work was supported by the MRC.