Human Cationic Amino Acid Transporters hCAT-1, hCAT-2A, and hCAT-2B:  Three Related Carriers with Distinct Transport Properties^,

Abstract

In this study, we aimed at analyzing the human homologues of the murine cationic amino acid transporters mCAT-1, mCAT-2A, and mCAT-2B. cDNAs encoding hCAT-1 had been previously reported by two independent groups [Albritton, L.M., et al. (1993) Genomics 12, 430; Yoshimoto, T., et al. (1991) Virology 185, 10]. We isolated cDNAs encoding hCAT-2A and hCAT-2B from a human liver cDNA library and from cDNA derived from the human hepatoma cell line HepG2, respectively. Analyses of the deduced amino acid sequences of both carriers demonstrated 90.9% identity with the respective murine proteins. In their functional domains (42 amino acids), both hCAT-2A and hCAT-2B differ only by one residue from the respective mouse proteins. Thus, CAT-2 proteins demonstrate a higher interspecies conservation than CAT-1 proteins that are overall 86.5% identical between mouse and human and differ by seven residues in the functional domain. The high degree of sequence conservation was reflected by the functional similarity of the human carriers with their mouse homologues. When expressed in Xenopus oocytes, hCAT-1 and hCAT-2B demonstrated transport properties consistent with y+. Unlike the mouse CAT-1 and CAT-2B, whose transport properties could hardly be distinguished, the transport properties of the human CAT-1 and CAT-2B isoforms showed clear differences: hCAT-1 had a 3-fold higher substrate affinity and was more sensitive to trans-stimulation than hCAT-2B. In contrast to the y+ carriers, hCAT-2A exhibited a 10-30-fold lower substrate affinity, a greater maximal velocity, and was much less sensitive to trans-stimulation at physiological substrate concentrations.

Increased Cat3-mediated Cationic Amino Acid Transport Functionally Compensates in Cat1 Knockout Cell Lines

“…As Cat1 is the only member of the Cat family reported to be trans-stimulated by greater than 3-fold (14) it is likely that Cat1 is responsible for the bulk of the observed transstimulation. The 2.3 Ϯ 0.2-fold increase in the initial rate of L-arginine transport in Cat1 Ϫ/Ϫ cells following trans-stimulation is in agreement with the reported values for Cat2 (14) and Cat3 (12). Furthermore, the finding that trans-stimulation in Cat1…”

“…Whereas these transporters share only about 61% amino acid sequence identity they were found to be kinetically indistinguishable with very similar K m values for L-arginine in oocytes (13,14). Cat1 1 appears to have a greater capacity than Cat2 for trans-stimulation (14), whereas an alternately spliced transcript of Cat2 encodes a kinetically distinct low affinity form of Cat2a (15). Cat3 was cloned from mouse (12) and rat (16).…”

In Vivo and In Vitro Evidence for Impaired Arginine Transport in Human Heart Failure

“…36 -38 Cloning studies have identified 4 CAT proteins. 39,40 CAT-1 and CAT-2B are both high-affinity, low-capacity transporters, thus resembling system y ϩ . In contrast, CAT-2A is a low-affinity, high-capacity transporter with limited distribution.…”

“…In contrast, CAT-2A is a low-affinity, high-capacity transporter with limited distribution. 40 CAT-3, a recently identified high-affinity transporter, has limited distribution, confined largely to the brain. 41 To identify the mechanism responsible for the observed reduction in both the V max for transport in isolated PBMCs and forearm clearance of [ 3 H]L-arginine, we examined the mRNA expression of CAT-1 and CAT-2B in PBMCs.…”

“…Although the y ϩ system accounts for the major proportion of arginine transport, it is possible that changes in expression of other arginine transporting systems, including the b o,ϩ system, could explain our findings. 40 However, this would seem unlikely given the fact that heart failure was associated with up to an 80% reduction in arginine transport, suggesting the involvement of the predominant arginine transport system. In summary, for the first time, we demonstrate an impairment of L-arginine transport in patients with CHF.…”