The amino acid transporter asc-1 is not involved in cystinuria.
BACKGROUND
The human amino acid transporter asc-1 (SLC7A10) exhibits substrate selectivity for small neutral amino acids, including cysteine, is expressed in kidney, is located close to the cystinuria B gene and presents sequence variants (e.g., E112D) in some cystinuria patients. We have cloned human asc-1, assessed its transport characteristics, localized its expression in kidney, searched for mutations in cystinuria patients, and tested the transport function of variant E112D.


METHODS
We used an EST-based homology cloning strategy. Transport characteristics of asc-1 were assessed by coexpression with 4F2hc in Xenopus oocytes and HeLa cells. Localization of asc-1 mRNA in kidney was assessed by in situ hybridization. Exons and intron-exon boundaries were polymerase chain reaction (PCR)-amplified from blood cell DNA and mutational screening was performed by single-stranded conformational polymorphism (SSCP).


RESULTS
Asc-1 reaches the plasma membrane in HeLa cells, unlike in oocytes, most probably by interaction with endogenous 4F2hc and presents similar transport characteristics to those in oocytes coexpressing asc-1/4F2hc. Asc-1 mediates a substantial efflux of alanine in a facilitated diffusion mode of transport. Expression of asc-1 mRNA localized to Henle's loop, distal tubules, and collecting ducts. Finally, SLC7A10 polymorphisms were identified in cystinuria probands and the SLC7A10 sequence variant E112D showed full transport activity.


CONCLUSIONS
The lack of expression of asc-1 in the proximal tubule indicates that it plays no role in the bulk of renal reabsorption of amino acids. No mutations causing cystinuria have been found in SLC7A10. The facilitated diffusion mode of transport and the expression in distal nephron suggest a role for asc-1 in osmotic adaptation.
Department of Biochemistry and Molecular Biology, Faculty of Biology, University of Barcelona, Spain.
Marta
M
Pineda
Mariona
M
Font
Maria Teresa
MT
Bassi
Marta
M
Manzoni
Giuseppe
G
Borsani
Valeria
V
Marigo
Esperanza
E
Fernández
Rafael Martín del
RM
Río
Jesús
J
Purroy
Antonio
A
Zorzano
Virginia
V
Nunes
Manuel
M
Palacín
D027182:Amino Acid Transport System y+ / Q000235:genetics* / Q000378:metabolism; D000596:Amino Acids / Q000378:metabolism; D000818:Animals; D003555:Cystinuria / Q000235:genetics* / Q000503:physiopathology*; D015870:Gene Expression; D006367:HeLa Cells; D006801:Humans; D007687:Kidney Tubules, Proximal / Q000502:physiology*; D009154:Mutation; D009865:Oocytes / Q000502:physiology; D012333:RNA, Messenger / Q000032:analysis; D014882:Water-Electrolyte Balance / Q000502:physiology; D014981:Xenopus
D027182:Amino Acid Transport System y+; D000596:Amino Acids; D012333:RNA, Messenger; C498799:SLC7A10 protein, human