Sustained Nitric Oxide Production in Macrophages Requires the Arginine Transporter CAT2

Abstract

The aberrant production of nitric oxide (NO) contributes to the pathogenesis of diseases as diverse as cancer and arthritis. Sustained NO production via the inducible enzyme, nitric-oxide synthase 2 (NOS2), requires extracellular arginine uptake. Three closely related cationic amino acid transporter genes (Cat1-3) encode the transporters that mediate most arginine uptake in mammalian cells. Because CAT2 is induced coordinately with NOS2 in numerous cell types, we investigated a possible role for CAT2-mediated arginine transport in regulating NO production. The complexity of arginine transport systems and their biochemically similar transport properties called for a genetic approach to determine the role of CAT2. CAT2-deficient mice were generated and found to be healthy and fertile in contrast to Cat1 animals. Analysis of cytokine-activated macrophages from Cat2؊/؊ mice revealed a 92% reduction in NO production and a 95% reduction in L-Arg uptake. The reduction in NO production was not due to differences in NOS2 protein expression, NOS2 activity, or intracellular L-arginine content. In conclusion, our results show that sustained abundant NO synthesis by macrophages requires arginine transport via the CAT2 transporter.The production and release of nitric oxide (NO) 1 are involved in numerous cellular processes. Overproduction of NO by inflammatory cells is implicated in the pathogenesis of diseases as diverse as cancer, endotoxic shock, atherosclerosis, and arthritis (1-7). NO is synthesized from arginine by three related enzymes. Two of these enzymes, the Ca 2ϩ -dependent neuronal nitric-oxide synthase and endothelial NOS, generate small amounts of NO over short periods of time (7). The Ca 2ϩ -independent inducible NOS (NOS2) produces large amounts of NO over sustained periods of time (1, 2). Activated macrophages produce copious quantities of NO via NOS2 over extended periods of time that contribute to tissue injury (2). Extracellular L-arginine is not required for endothelial NO synthasemediated NO production in human endothelial cells (8). In contrast, extracellular arginine is required for sustained NO production via NOS2 in macrophages (8,9). In addition, increased arginine transport is known to accompany NO production via NOS2 (1, 2, 9, 10).Among the several transport systems that mediate L-arginine uptake (y ϩ , B 0ϩ

Creation and implications of a phenome-genome network

“…For example, mice that are homozygous null for Slc7a2 show a reduction in nitric oxide production in macrophages and fibroblasts, important in immune response and wound healing. 30,31 Mice homozygous null for Mgat2 show early post-natal lethality and a number of other defects. 32 Decreases in Tnnt1 and increases in Synj1 have been found to be markers for stem cell differentiation.…”

Lipopolysaccharide-induced Apoptosis of Macrophages Determines the Up-regulation of Concentrative Nucleoside Transporters Cnt1 and Cnt2 through Tumor Necrosis Factor-α-dependent and -independent Mechanisms

“…The TNF-␣ receptor I knock-out mice were generated and characterized as reported (33). Functional inducible NO knock-out mice generated by disrupting the arginine transporter CAT2 gene (34) were also used to complement the data obtained with the iNOS knock-out animals.…”

Molecular basis of substrate-induced permeation by an amino acid antiporter

“…The APC superfamily includes two families of eukaryotic transporters that have an impact on human health: Cationic Amino acid Transporters (CATs) and L-Amino acid Transporters (LATs). CATs play key roles in macrophage activation (6,7) and proliferation (8). Within the LAT members, mutations in b 0;þ AT and y þ LAT1 cause the primary inherited aminoacidurias cystinuria B and lysinuric protein intolerance, respectively (9)(10)(11), and xCT (system x c -transporter) is involved in cell redox balance (12), Kaposi's sarcoma-associated herpesvirus infection (13,14), and cocaine relapse (15).…”