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J. Biol. Chem., Vol. 283, Issue 30, 20761-20769, July 25, 2008

This Article Full Text Full Text (PDF) All Versions of this Article: 283/30/20761    most recent M801024200v1 Alert me when this article is cited Alert me if a correction is posted Citation Map Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Google Scholar Articles by López-Contreras, A. J. Articles by Peñafiel, R. PubMed PubMed Citation Articles by López-Contreras, A. J. Articles by Peñafiel, R. Social Bookmarking                      What's this?

Antizyme Inhibitor 2 (AZIN2/ODCp) Stimulates Polyamine Uptake in Mammalian Cells^*

Andrés J. López-Contreras¹, Bruno Ramos-Molina², Asunción Cremades, and Rafael Peñafiel³

From the Department of Biochemistry and Molecular Biology B and Immunology and Department of Pharmacology, Faculty of Medicine, University of Murcia, 30100 Murcia, Spain

One of the processes that regulate intracellular levels of polyamines in mammalian cells is polyamine uptake. We have measured polyamine uptake in COS7 cells for putrescine, spermidine, and spermine, obtaining K_m values of 4.5, 1.0, and 0.8 µM, respectively. Treatment of nonconfluent cells with cycloheximide stimulated polyamine uptake and prevented the inhibitory effect found in cells preloaded with polyamines, suggesting the existence of a feedback repression mechanism mediated by antizymes. Transient transfected cells with mutated antizyme forms of AZ1, AZ2, and AZ3, which do not require frameshifting, showed a total blockade of polyamine uptake. Transfection of COS7 cells with mouse or human AZIN2, a novel member of the antizyme inhibitor family, recently characterized by our group, markedly stimulated polyamine uptake and counteracted the action of any of the three antizymes in co-transfected cells. The stimulatory effect of AZIN2 on polyamine uptake was abrogated when the putative antizyme binding sequence, formed by residues 117–140 in AZIN2, was deleted. Real time reverse transcription-PCR analysis of antizyme inhibitor transcripts revealed that in brain and testes AZIN2 is more expressed than AZIN1, especially in the testes where the relative expression was about 25-fold higher. Collectively, our results clearly indicate that AZIN2 affects polyamine homeostasis not only by increasing ornithine decarboxylase activity but also by stimulating polyamine uptake, through negating the inhibitory effect of the antizymes. This finding may have physiological relevance, mostly in testes where AZ3 and AZIN2 are mainly expressed.

Received for publication, February 7, 2008 , and in revised form, May 1, 2008.

^* This work was supported in part by Grants 00466/PI/04 from The Seneca Foundation (Autonomous Community of Murcia) and BFU2005-09378-C02 from the Spanish Ministry of Education. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

¹ Recipient of a fellowship from The Seneca Foundation.

² Supported by the University of Murcia, 30100 Murcia, Spain.

³ To whom correspondence should be addressed: Dept. of Biochemistry and Molecular Biology B and Immunology, Faculty of Medicine, University of Murcia, Campus de Espinardo 30100 Murcia, Spain. Tel.: 34-968367174; E-mail: rapegar{at}um.es.

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