Subcellular localization of antizyme inhibitor 2 in mammalian cells: Influence of intrinsic sequences and interaction with antizymes

López-Contreras, Andrés J.; Sánchez-Laorden, Berta; Ramos‐Molina, Bruno; Morena, Maria; Cremades, Adolfo Díaz-Bautista; Peñafiel, Rafael

doi:10.1002/jcb.22168

Cited by 22 publications

(26 citation statements)

References 43 publications

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“…2b). In accordance with previous reports, both ODC 47 and SPR (proteinatlas.org) primarily reside in the cytoplasm. In the MYCN2 NB cell line used in this study, ODC and SPR resided in punctuate sites within the cytoplasm.…”

Section: Resultssupporting

confidence: 92%

Novel Interaction of Ornithine Decarboxylase with Sepiapterin Reductase Regulates Neuroblastoma Cell Proliferation

Lange

Geerts

Feith

et al. 2014

Journal of Molecular Biology

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Ornithine decarboxylase (ODC) is the sentinel enzyme in polyamine biosynthesis. Both ODC and polyamines regulate cell division, proliferation, and apoptosis. Sepiapterin reductase (SPR) catalyzes the last step in the biosynthesis of tetrahydrobiopterin (BH4), an essential cofactor of nitric oxide synthase (NOS), and has been implicated in neurological diseases but not yet in cancer. In this study, we present compelling evidence that native ODC and SPR physically interact, and we defined the individual amino acid residues involved in both enzymes using in silico protein-protein docking simulations. The resulting heterocomplex is a surprisingly compact structure, featuring two energetically and structurally equivalent binding modes both in monomer and dimer conformations. The novel interaction between ODC and SPR proteins was confirmed under physiological conditions by co-immunoprecipitation and co-localization in neuroblastoma (NB) cells. Importantly, we showed that siRNA-mediated knock-down of SPR expression significantly reduced endogenous ODC enzyme activity in NB cells, thus demonstrating the biological relevance of the ODC-SPR interaction. Finally, in a cohort of 88 human NB tumors we found that high SPR mRNA expression correlated significantly with poor survival prognosis using a Kaplan-Meier analysis (Logrank test P = 5 • 10−4), suggesting an oncogenic role for SPR in NB tumorigenesis. In conclusion, we showed that ODC binds SPR and thus propose a new concept in which two well-characterized biochemical pathways converge via the interaction of two enzymes. We identified SPR as a novel regulator of ODC enzyme activity, and based on clinical evidence present a model in which SPR drives ODC-mediated malignant progression in NB.

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Section: Resultssupporting

confidence: 92%

Novel Interaction of Ornithine Decarboxylase with Sepiapterin Reductase Regulates Neuroblastoma Cell Proliferation

Lange

Geerts

Feith

et al. 2014

Journal of Molecular Biology

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“…This new set of AZIN2 expressing tissues have in common to possess either endocrine or paracrine functions, and therefore, to be endowed with a prominent secretory activity. This fact, together with the reported subcellular localization in the ERGIC [23] and in the trans-Golgi network [24], key components of the secretory pathway, suggests a relevant role for AZIN2 in this process. This contention is also supported by recent findings showing that polyamines may influence the secretory activity in various systems.…”

Section: Discussionmentioning

confidence: 74%

“…Azin2 βGeo/+ mice did not display any evident phenotype. It is important to note that the gene product from the recombinant allele comprises the N-terminal 92 amino acid residues of native AZIN2, the critical sequence that drives AZIN2 to its subcellular localization in the Endoplasmic Reticulum-Golgi Intermediate Compartment (ERGIC) [23], and therefore it can be considered that the pattern of β-D-galactosidase staining might be informative of the native localization of AZIN2. Amidst those tissues, we detected histochemical signal in testis, pancreas, epididymis, brain and adrenal gland.…”

Section: Resultsmentioning

confidence: 99%

“…Since we had found that in HEK 293T cells transfected with Azin2, the AZIN2 protein was mainly found in the particulate fraction of the cells [23], in the transgenic mice we analyzed the distribution of β–galactosidase fused to the N-terminal fragment of AZIN2in the soluble and particulate fractions of tissue extracts. For that purpose, tissues were homogenized in an isotonic buffer containing 25 mM TRIS-HCl pH7.4, 0.25 M sucrose and 0.2 mM EDTA, and the homogenates were centrifuged at 12,000×g for 20 min.…”

Section: Methodsmentioning

confidence: 99%

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Antizyme Inhibitor 2 Hypomorphic Mice. New Patterns of Expression in Pancreas and Adrenal Glands Suggest a Role in Secretory Processes

et al. 2013

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The intracellular levels of polyamines, polycations implicated in proliferation, differentiation and cell survival, are regulated by controlling their biosynthesis, catabolism and transport. Antizymes and antizyme inhibitors are key regulatory proteins of polyamine levels by affecting ornithine decarboxylase, the rate-limiting biosynthetic enzyme, and polyamine uptake. We recently described the molecular function of a novel antizyme inhibitor (AZIN2). However, the physiological function of AZIN2 in mammals is mostly unknown. To gain insight on the tissue expression profile of AZIN2 and to find its possible physiological role, we have generated, transgenic mice with severe Azin2 hypomorphism. This mouse model expresses transgenic bacterial β-D-galactosidase as a reporter gene, under the control of the Azin2 endogenous promoter, what allows a very sensitive and specific detection of the expression of the gene in the different tissues of transgenic mice. The biochemical and histochemical analyses of β-D-galactosidase together with the quantification of Azin2 mRNA levels, corroborated that AZIN2 is mainly expressed in testis and brain, and showed for the first time that AZIN2 is also expressed in the adrenal glands and pancreas. In these tissues, AZIN2 was not expressed in all type of cells, but rather in specific type of cells. Thus, AZIN2 was mainly found in the haploid germinal cells of the testis and in different brain regions such as hippocampus and cerebellum, particularly in specific type of neurons. In the adrenal glands and pancreas, the expression was restricted to the adrenal medulla and to the Langerhans islets, respectively. Interestingly, plasma insulin levels were significantly reduced in the transgenic mice. These results support the idea that AZIN2 may have a role in the modulation of reproductory and secretory functions and that this mouse model might be an interesting tool for the progress of our understanding on the role of AZIN2 and polyamines in specific mammalian cells.

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“…These are paralogs of ODC that lack enzymatic activity and, due to their structural resemblance to ODC, inhibit AZ1‐3 by binding to them, thus lowering the access of antizymes to ODC and polyamine uptake‐related binding sites. This action enhances ODC‐mediated synthesis of polyamines and polyamine uptake, increasing cytoplasmic polyamine levels (López‐Contreras et al, ; López‐Contreras et al, ; López‐Contreras et al, ; Ramos‐Molina et al, ). It appears that polyamines indirectly promote cell growth.…”

Section: Introductionmentioning

confidence: 99%

An exercise in brain genoarchitectonics: Analysis of AZIN2‐Lacz expressing neuronal populations in the mouse hindbrain

Martı́nez-de-la-Torre

Lambertos

Peñafiel

et al. 2017

J of Neuroscience Research

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We examined in detail the distribution of AZIN2 (antizyme inhibitor 2) expression in the adult mouse hindbrain and neighboring spinal cord. AZIN2, similar to previously known AZIN1, is a recently-discovered, a functional paralog of ornithine decarboxylase (ODC). Due to their structural similarity to ODC, both AZIN1 and AZIN2 counteract the inhibitory action of 3 known antizymes (AZ1-3) on the ODC synthesis of polyamines, thus increasing intracytoplasmic levels of polyamines. AZIN2 is strongly, but heterogeneously, expressed in the brain. Our study uses a mouse line carrying an AZIN2-LacZ construct, and, in our topographic analysis of AZIN2-positive structures, we intend to share new knowledge about the rhombomeric segmentation of the hindbrain (a function of Hox paralogs and other genes). The observed labeled cell populations predominantly coincide with known cholinergic and glutamatergic cells, but occasionally also correspond to GABAergic, and possibly glycinergic cells. Some imperfectly known hindbrain populations stood out in unprecedented detail, and some axonal tracts were also differentially stained. © 2017 Wiley Periodicals, Inc.

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Subcellular localization of antizyme inhibitor 2 in mammalian cells: Influence of intrinsic sequences and interaction with antizymes

Cited by 22 publications

References 43 publications

Novel Interaction of Ornithine Decarboxylase with Sepiapterin Reductase Regulates Neuroblastoma Cell Proliferation

Novel Interaction of Ornithine Decarboxylase with Sepiapterin Reductase Regulates Neuroblastoma Cell Proliferation

Antizyme Inhibitor 2 Hypomorphic Mice. New Patterns of Expression in Pancreas and Adrenal Glands Suggest a Role in Secretory Processes

An exercise in brain genoarchitectonics: Analysis of AZIN2‐Lacz expressing neuronal populations in the mouse hindbrain

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