Arginine transport in human erythroid cells: discrimination of CAT1 and 4F2hc/y+LAT2 roles

Abstract

Since arginine metabolites, such as nitric oxide and polyamines, influence the expression of genes involved in erythroid differentiation, the transport of the cationic amino acid may play an important role in erythroid cells. However, available data only concern the presence in these cells of CAT1 transporter (system y(+)), while no information exists on the role of the heterodimeric transporters of system y(+)L (4F2hc/y(+)LAT1 and 4F2hc/y(+)LAT2) which operates transmembrane arginine fluxes cis-inhibited by neutral amino acids in the presence of sodium. Using erythroleukemia K562 cells and normal erythroid precursors, we demonstrate here that arginine transport in human erythroid cells is due to the additive contributions of a leucine-sensitive and leucine-insensitive component. In both cell types, leucine inhibition of arginine influx is much less evident in the absence of sodium, a hallmark of system y(+)L. In K562 cells, N-ethylmaleimide, a known inhibitor of CAT transporters (system y(+)), suppresses only a fraction of arginine influx corresponding to leucine-insensitive uptake. Moreover, in Xenopus oocytes coexpressing 4F2hc and y(+)LAT2, leucine exerts a marked inhibition of arginine transport, partially dependent on sodium, while no inhibition is seen in oocytes expressing CAT1. Lastly, silencing of SLC7A6, the gene for y(+)LAT2, lowers arginine transport and doubles the intracellular content of the cationic amino acid in K562 cells. We conclude that arginine transport in human erythroid cells is due to both system y(+) (CAT1 transporter) and system y(+)L (4F2hc/y(+)LAT2 isoform), which mainly contribute, respectively, to the influx and to the efflux of the cationic amino acid.

Maturing reticulocytes internalize plasma membrane in glycophorin A–containing vesicles that fuse with autophagosomes before exocytosis

“…Arginine import through the CAT1 and CD98hc transport systems is required for proliferation and differentiation of erythroid cells. 21,22 In contrast, some CD147 is retained in the mature erythrocyte, possibly to facilitate lactate transport through MCT1. We observe a gross reduction in expression of tubulin and loss of cytosolic actin as the human reticulocyte matured, as described previously in murine reticulocytes.…”

Arginine transport in human monocytic leukemia THP-1 cells during macrophage differentiation

“…The characterization of arginine transport was performed, according to the strategy already used in other cell models [9,18,28], either in undifferentiated (Fig. 1A) or in PMA-differentiated THP-1 cells (Fig.…”

“…l-arginine uptake in undifferentiated cultures of THP-1 was assessed as described previously for cells grown in suspension [9]. For this assay, cells were centrifuged 5 min at 1100 rpm, and the pellet was suspended in an EBSS, buffered at pH 7.4, with 26 mM Tris-HCl.…”

“…After 1 min, the incubation was ended with a rapid dilution of the cell suspension in ice-cold urea (2 ml, 300 mM), followed by a centrifugation at 12,000 g. The pellet was then washed with urea and extracted with 200 l 5% sodium deoxycholate in 1 N NaOH. After 1 h, 100 l of the cell extracts were used for radioactivity determination with a Wallac Microbeta Trilux 2 liquid scintillation spectrometer (Perkin Elmer, Wellesley, MA, USA), and the remaining aliquot was used for protein determination through a modified Lowry procedure [9].…”

“…System y ϩ L operates as an antiport that couples the flux of arginine with that of neutral amino acids and sodium [7]. While the activity of system y ϩ is mainly responsible for the intracellular accumulation of arginine, system y ϩ L likely acts as an efflux route for the amino acid [8,9]. Interestingly, SLC7A7 is the gene mutated in patients affected by LPI (MIM 222700) [10,11], a rare genetic disorder characterized by impaired, dibasic amino acid transport.…”