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Interactions of the natural product (+)-avrainvillamide with nucleophosmin and exportin-1 Mediate the cellular localization of nucleophosmin and its AML-associated mutants.

ABSTRACT: Nucleophosmin (NPM1) is a multifunctional phosphoprotein localized predominantly within the nucleoli of eukaryotic cells. Mutations within its C-terminal domain are frequently observed in patients with acute myeloid leukemia (AML), are thought to play a key role in the initiation of the disease, and result in aberrant, cytoplasmic localization of the mutant protein. We have previously shown that the electrophilic antiproliferative natural product (+)-avrainvillamide (1) binds to proteins, including nucleophosmin, by S-alkylation of cysteine residues. Here, we report that avrainvillamide restores nucleolar localization of certain AML-associated mutant forms of NPM1 and provide evidence that this relocalization is mediated by interactions of avrainvillamide with mutant NPM1 and exportin-1 (Crm1). Immunofluorescence and mass spectrometric experiments employing a series of different NPM1 constructs suggest that a specific interaction between avrainvillamide and Cys275 of certain NPM1 mutants mediates the relocalization of these proteins to the nucleolus. Avrainvillamide treatment is also shown to inhibit nuclear export of Crm1 cargo proteins, including AML-associated NPM1 mutants. We also observe that avrainvillamide treatment displaces Thr199-phosphorylated NPM1 from duplicated centrosomes, leads to an accumulation of supernumerary centrosomes, and inhibits dephosphorylation of Thr199-phosphorylated NPM1 by protein phosphatase 1. Avrainvillamide is the first small molecule reported to relocalize specific cytoplasmic AML-associated NPM1 mutants to the nucleolus, providing an important demonstration of principle that small molecule induction of a wild-type NPM1 localization phenotype is feasible in certain human cancer cells.

SUBMITTER: Mukherjee H

PROVIDER: S-EPMC4652655 | biostudies-literature | 2015 Mar

REPOSITORIES: biostudies-literature

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Interactions of the natural product (+)-avrainvillamide with nucleophosmin and exportin-1 Mediate the cellular localization of nucleophosmin and its AML-associated mutants.

Mukherjee Herschel H Chan Kok-Ping KP Andresen Vibeke V Hanley Mariah L ML Gjertsen Bjørn Tore BT Myers Andrew G AG

ACS chemical biology 20150105 3

Nucleophosmin (NPM1) is a multifunctional phosphoprotein localized predominantly within the nucleoli of eukaryotic cells. Mutations within its C-terminal domain are frequently observed in patients with acute myeloid leukemia (AML), are thought to play a key role in the initiation of the disease, and result in aberrant, cytoplasmic localization of the mutant protein. We have previously shown that the electrophilic antiproliferative natural product (+)-avrainvillamide (1) binds to proteins, includ ...[more]

PMID: 25531824

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Project description:BACKGROUND:Programmed ribosomal frameshifting (PRF) is a gene expression mechanism which enables the translation of two N-terminally coincident, C-terminally distinct protein products from a single mRNA. Many viruses utilize PRF to control or regulate gene expression, but very few phylogenetically conserved examples are known in vertebrate genes. Additional sex combs-like (ASXL) genes 1 and 2 encode important epigenetic and transcriptional regulatory proteins that control the expression of homeotic genes during key developmental stages. Here we describe an ~150-codon overlapping ORF (termed TF) in ASXL1 and ASXL2 that, with few exceptions, is conserved throughout vertebrates. RESULTS:Conservation of the TF ORF, strong suppression of synonymous site variation in the overlap region, and the completely conserved presence of an EH[N/S]Y motif (a known binding site for Host Cell Factor-1, HCF-1, an epigenetic regulatory factor), all indicate that TF is a protein-coding sequence. A highly conserved UCC_UUU_CGU sequence (identical to the known site of +1 ribosomal frameshifting for influenza virus PA-X expression) occurs at the 5' end of the region of enhanced synonymous site conservation in ASXL1. Similarly, a highly conserved RG_GUC_UCU sequence (identical to a known site of -2 ribosomal frameshifting for arterivirus nsp2TF expression) occurs at the 5' end of the region of enhanced synonymous site conservation in ASXL2. CONCLUSIONS:Due to a lack of appropriate splice forms, or initiation sites, the most plausible mechanism for translation of the ASXL1 and 2 TF regions is ribosomal frameshifting, resulting in a transframe fusion of the N-terminal half of ASXL1 or 2 to the TF product, termed ASXL-TF. Truncation or frameshift mutants of ASXL are linked to myeloid malignancies and genetic diseases, such as Bohring-Opitz syndrome, likely at least in part as a result of gain-of-function or dominant-negative effects. Our hypothesis now indicates that these disease-associated mutant forms represent overexpressed defective versions of ASXL-TF. REVIEWERS:This article was reviewed by Laurence Hurst and Eugene Koonin.

Dataset Information

Interactions of the natural product (+)-avrainvillamide with nucleophosmin and exportin-1 Mediate the cellular localization of nucleophosmin and its AML-associated mutants.

Publications

Interactions of the natural product (+)-avrainvillamide with nucleophosmin and exportin-1 Mediate the cellular localization of nucleophosmin and its AML-associated mutants.

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