Regulation of all members of the antizyme family by antizyme inhibitor

• Published in: Biochemical journal Vol. 385; no. Pt 1; pp. 21 - 28
• Main Authors: Mangold, Ursula, Leberer, Ekkehard
• Format: Journal Article
• Language: English
• Published: England Portland Press Ltd 01.01.2005
• Subjects: Carrier Proteins - genetics; Carrier Proteins - metabolism; Cell Line; Enzyme Inhibitors - metabolism; Genetic Complementation Test; Humans; Immunoprecipitation; Multiprotein Complexes - chemistry; Multiprotein Complexes - genetics; Multiprotein Complexes - metabolism; Organ Specificity; Ornithine Decarboxylase - genetics; Ornithine Decarboxylase - metabolism; Ornithine Decarboxylase Inhibitors; Protein Binding; Proteins - antagonists & inhibitors; Proteins - classification; Proteins - genetics; Proteins - metabolism; Putrescine - pharmacology; Saccharomyces cerevisiae - enzymology; Saccharomyces cerevisiae - genetics; Saccharomyces cerevisiae - metabolism; Two-Hybrid System Techniques
• ISSN: 0264-6021; 1470-8728
• DOI: 10.1042/BJ20040547

ODC (ornithine decarboxylase) is the rate-limiting enzyme in polyamine biosynthesis. Polyamines are essential for cellular growth and differentiation but enhanced ODC activity is associated with cell transformation. Post-translationally, ODC is negatively regulated through members of the antizyme family. Antizymes inhibit ODC activity, promote ODC degradation through the 26 S proteasome and regulate polyamine transport. Besides the ubiquitously expressed antizymes 1 and 2, there is the tissue-specific antizyme 3 and an yet uncharacterized antizyme 4. Antizyme 1 has been shown to be negatively regulated through the AZI (antizyme inhibitor) that binds antizyme 1 with higher affinity compared with ODC. In the present study, we show by yeast two- and three-hybrid protein-protein interaction studies that AZI interacts with all members of the antizyme family and is capable of disrupting the interaction between each antizyme and ODC. In a yeast-based ODC complementation assay, we show that human ODC is able to complement fully the function of the yeast homologue of ODC. Co-expression of antizymes resulted in ODC inhibition and cessation of yeast growth. The antizyme-induced growth inhibition could be reversed by addition of putrescine or by the co-expression of AZI. The protein interactions could be confirmed by immunoprecipitation of the human ODC-antizyme 2-AZI complexes. In summary, we conclude that human AZI is capable of acting as a general inhibitor for all members of the antizyme family and that the previously not yet characterized antizyme 4 is capable of binding ODC and inhibiting its enzymic activity similar to the other members of the antizyme family.