Cystinuria (MIM 220100) is a common recessive disorder of renal reabsorption of cystine and dibasic amino acids. Mutations in SLC3A1, encoding rBAT, cause cystinuria type I (ref. 1), but not other types of cystinuria (ref. 2). A gene whose mutation causes non-type I cystinuria has been mapped by linkage analysis to 19q12-13.1 (Refs 3,4). We have identified a new transcript, encoding a protein (bo, +AT, for bo,+ amino acid transporter) belonging to a family of light subunits of amino acid transporters, expressed in kidney, liver, small intestine and placenta, and localized its gene (SLC7A9) to the non-type I cystinuria 19q locus. Co-transfection of bo,+AT and rBAT brings the latter to the plasma membrane, and results in the uptake of L-arginine in COS cells. We have found SLC7A9 mutations in Libyan-Jews, North American, Italian and Spanish non-type I cystinuria patients. The Libyan Jewish patients are homozygous for a founder missense mutation (V170M) that abolishes b o,+AT amino-acid uptake activity when co-transfected with rBAT in COS cells. We identified four missense mutations (G105R, A182T, G195R and G295R) and two frameshift (520insT and 596delTG) mutations in other patients. Our data establish that mutations in SLC7A9 cause non-type I cystinuria, and suggest that bo,+AT is the light subunit of rBAT.

Cystinuria (OMIM 220100) is a common recessive disorder of renal reabsorption of cystine and dibasic amino acids that results in nephrolithiasis of cystine. Mutations in SLC3A1, which encodes rBAT, cause Type I cystinuria, and mutations in SLC7A9, which encodes a putative subunit of rBAT (b(o,+)AT), cause non-Type I cystinuria. Here we describe the genomic structure of SLC7A9 (13 exons) and 28 new mutations in this gene that, together with the seven previously reported, explain 79% of the alleles in 61 non-Type I cystinuria patients. These data demonstrate that SLC7A9 is the main non-Type I cystinuria gene. Mutations G105R, V170M, A182T and R333W are the most frequent SLC7A9 missense mutations found. Among heterozygotes carrying these mutations, A182T heterozygotes showed the lowest urinary excretion values of cystine and dibasic amino acids. Functional analysis of mutation A182T after co-expression with rBAT in HeLa cells revealed significant residual transport activity. In contrast, mutations G105R, V170M and R333W are associated to a complete or almost complete loss of transport activity, leading to a more severe urinary phenotype in heterozygotes. SLC7A9 mutations located in the putative transmembrane domains of b(o,+)AT and affecting conserved amino acid residues with a small side chain generate a severe phenotype, while mutations in non-conserved residues give rise to a mild phenotype. These data provide the first genotype-phenotype correlation in non-Type I cystinuria, and show that a mild urinary phenotype in heterozygotes may associate with mutations with significant residual transport activity.

The heteromeric amino acid transporters are composed of a type II glycoprotein and a non-glycosylated polytopic membrane protein. System b o,+ exchanges dibasic for neutral amino acids. It is composed of rBAT and b o,+ AT, the latter being the polytopic membrane subunit. Mutations in either of them cause malfunction of the system, leading to cystinuria. IntroductionThe heteromeric amino acid transporters (HATs) are the only known eukaryotic transporters of amino acids composed of two different subunits bound by a disul®de bridge: (i) a type II glycoprotein or heavy subunit (rBAT and 4F2hc), and (ii) a non-glycosylated light subunit with 12 putative transmembrane domains (LAT-1, LAT-2, asc1, y + LAT-1, y + LAT-2, xCT and b o,+ AT) (for reviews see Verrey et al., 1999Verrey et al., , 2000Deve Âs and Boyd, 2000;Chillaro Ân et al., 2001;Wagner et al., 2001). The heavy subunit has an ectodomain that is homologous to bacterial glucosidases (reviewed in Chillaro Ân et al., 2001). The light subunit is suspected to be the catalytic part of the holotransporter because it is considered to be a polytopic membrane protein and heterodimers of 4F2hc with different light subunits yield different amino acid transport systems: (i) with LAT-1 or LAT-2, variants of system L ( Kanai et al., 1998;Mastroberardino et al., 1998;Pineda et al., 1999;Rossier et al., 1999;Segawa et al., 1999) The light subunit of system b o,+ is fully functional in the absence of the heavy subunit