Our research is focused on an unusual group of membrane proteins which catalyze amino acid transport in mammalian cells. This group includes genes involved in two amino acid transport disorders: cystinuria and lysinuric protein intolerance (LPI). In cystinuria, excessive amounts of cystine and basic amino acids (BAA's) are excreted in urine. It is a common genetic disorder (overall incidence of 1 in 7,000). Low solubility of cystine results in formation of kidney stones which can lead to renal failure. Symptoms of LPI include failure to thrive, muscle hypotonia, hepatosplenomegaly, and episodic hyperammonemia. The defect in cystinuria involves a transporter for cystine and BAA's located in the apical membranes of renal and intestinal epithelial cells, and in LPI, a BAA transporter in the basolateral membranes of these cells. Both these transporters are hetero-oligomeric complexes, each composed of at least 2 subunits, one a glycoprotein anchored to the plasma membrane via 1 to 4 membrane-spanning domains (MSD's) and the other, a 12-MSD, highly hydrophobic protein. In cystinurics, mutations have been noted in both the glycoprotein subunit (termed NBAT or rBAT) and in the 12-MSD subunit (termed bAT). In LPI, mutations have been detected only in the 12-MSD subunit (termed yLAT) of the BAA transporter (the glycoprotein subunit of this transporter is called 4F2hc, found in many tissues). In cell expression systems, eg., the Xenopus oocytes, the NBAT/bAT complex expresses a high-affinity, Na-independent transport system for cystine, BAA's, and certain neutral amino acids (NAA's) (system b0,+), whereas the 4F2hc/yLAT complex expresses Na-independent transport for BAA's and Na-dependent transport for certain of the NAA's (system y+L). The 12-MSD subunit (either bAT or yLAT), when expressed alone in the oocytes, fails to reach the cell surface; coexpression with the respective glycoprotein subunit is necessary for the complex to be expressed on the cell surface. The glycoproteins, NBAT and 4F2hc, display significant sequence homology. yLAT belongs to a family of highly homologous 12-MSD proteins, each of which interact with 4F2hc producing complexes which exhibit different transport phenotypes (thus, 4F2hc/yLAT complex expresses, as noted above, system y+L; 4F2hc complexes containing other LAT family members express either system L, a major Na-independent NAA transport system, or a cystine-glutamate exchange transporter). Furthermore, bAT and yLAT exhibit about 40% sequence identity; however, bAT does not form a functional complex with 4F2hc, neither does yLAT form a functional complex with NBAT.

Our current studies focus on: (1) the effect of the various cystinuria-specific and LPI-specific mutations on the structure and transport functions of the cystine and BAA transporters; (2) use of site-specific and chimeric mutants to map and identify sites which are the determinants in expression of the different transport phenotypes; and (3) elucidate the targeting information in the glycoprotein subunits which are responsible for the intracellular transport and targeting of the transporter complexes.