Expression cloning of a human renal cDNA that induces high affinity transport of L-cystine shared with dibasic amino acids in Xenopus oocytes

Joan Bertran, Andreas Werner, Josep Chillarón, Virginia Nunes, Jürg Biber, Xavier Testar, Antonio Zorzano, Xavier P. Estivill, Heini Murer, Manuel Palacín

Research output: Contribution to journalArticle

111 Citations (Scopus)

Abstract

A renal cDNA clone (rBAT) that induces system bo,+-like amino acid transport activity in Xenopus oocytes has recently been isolated (Bertran, J., Werner, A., Moore, M. L., Stange, G., Markovich, D., Biber, J., Tester, X., Zorzano, A., Palacín, and Murer, H. (1992) Proc. Natl. Acad. Sci. U. S. A. 89, 5601-5605). Here we show the isolation of a cDNA clone by screening a human kidney cortex cDNA library for expression of sodium-independent transport of L-[3H]arginine in Xenopus oocytes. The cRNA of this clone induces in oocytes, in addition to the uptake of L-arginine, that of L-[35S]cystine and L-[3H]leucine. Expressed uptake of these amino acids is mutually cis-inhibitable by the other 2 amino acids. Expressed uptake of L-cystine is saturable and shows an apparent Km in the micromolar range. All these characteristics resemble induction of system bo,+ related to rBAT in the oocytes. Human rBAT mRNA (∼2.5 kilobases) is found in kidney, small intestine (i.e., jejunum), pancreas, and liver. Human kidney poly(A)+ RNA (mRNA) induces sodium-independent uptake of L-cystine, L-arginine, and L-leucine in Xenopus oocytes. Hybrid depletion with an antisense oligonucleotide of the isolated clone greatly prevents (80-97%) human kidney mRNA-dependent induction of the uptake of these amino acids (i.e. L-cystine, L-arginine, and L-leucine). The isolated clone (2304 base pairs in length) contains a poly(A) tail and encodes a predicted 78.8-kDa protein which is 85 and 80% identical to the rabbit and rat rBAT, respectively. This predicted protein corresponds to a membrane glycoprotein, and contains six potential N-glycosylation sites which might be functional in the oocyte: [35S] methionine labeling of oocytes shows a specific band of 94 kDa in crude membranes of these human cRNA-injected oocytes; treatment of these oocytes with tunicamycin shifts the cRNA-specific translation product to approximately 72 kDa. We conclude that we have isolated a functional cDNA corresponding to human rBAT. The isolation of this human cDNA would lead to the study of the possible involvement of rBAT in human hyperaminoacidurias.

Original languageEnglish
Pages (from-to)14842-14849
Number of pages8
JournalJournal of Biological Chemistry
Volume268
Issue number20
Publication statusPublished - 15 Jul 1993
Externally publishedYes

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Diamino Amino Acids
Cystine
Cloning
Xenopus
Oocytes
Organism Cloning
Complementary DNA
Complementary RNA
Arginine
Kidney
Messenger RNA
Leucine
Amino Acids
Clone Cells
Sodium
Glycosylation
Tunicamycin
Antisense Oligonucleotides
Membrane Glycoproteins
Gene Library

ASJC Scopus subject areas

  • Biochemistry

Cite this

Bertran, J., Werner, A., Chillarón, J., Nunes, V., Biber, J., Testar, X., ... Palacín, M. (1993). Expression cloning of a human renal cDNA that induces high affinity transport of L-cystine shared with dibasic amino acids in Xenopus oocytes. Journal of Biological Chemistry, 268(20), 14842-14849.

Expression cloning of a human renal cDNA that induces high affinity transport of L-cystine shared with dibasic amino acids in Xenopus oocytes. / Bertran, Joan; Werner, Andreas; Chillarón, Josep; Nunes, Virginia; Biber, Jürg; Testar, Xavier; Zorzano, Antonio; Estivill, Xavier P.; Murer, Heini; Palacín, Manuel.

In: Journal of Biological Chemistry, Vol. 268, No. 20, 15.07.1993, p. 14842-14849.

Research output: Contribution to journalArticle

Bertran, J, Werner, A, Chillarón, J, Nunes, V, Biber, J, Testar, X, Zorzano, A, Estivill, XP, Murer, H & Palacín, M 1993, 'Expression cloning of a human renal cDNA that induces high affinity transport of L-cystine shared with dibasic amino acids in Xenopus oocytes', Journal of Biological Chemistry, vol. 268, no. 20, pp. 14842-14849.
Bertran, Joan ; Werner, Andreas ; Chillarón, Josep ; Nunes, Virginia ; Biber, Jürg ; Testar, Xavier ; Zorzano, Antonio ; Estivill, Xavier P. ; Murer, Heini ; Palacín, Manuel. / Expression cloning of a human renal cDNA that induces high affinity transport of L-cystine shared with dibasic amino acids in Xenopus oocytes. In: Journal of Biological Chemistry. 1993 ; Vol. 268, No. 20. pp. 14842-14849.
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