Destabilisation, aggregation, toxicity and cytosolic mislocalisation of nucleophosmin regions associated with acute myeloid leukemia

Oncotarget. 2016 Sep 13;7(37):59129-59143. doi: 10.18632/oncotarget.10991.

Abstract

Nucleophosmin (NPM1) is a multifunctional protein that is implicated in the pathogenesis of several human malignancies. To gain insight into the role of isolated fragments of NPM1 in its biological activities, we dissected the C-terminal domain (CTD) into its helical fragments. Here we focus the attention on the third helix of the NPM1-CTD in its wild-type (H3 wt) and AML-mutated (H3 mutA and H3 mutE) sequences. Conformational studies, by means of CD and NMR spectroscopies, showed that the H3 wt peptide was partially endowed with an α-helical structure, but the AML-sequences exhibited a lower content of this conformation, particularly the H3 mutA peptide. Thioflavin T assays showed that the H3 mutE and the H3 mutA peptides displayed a significant aggregation propensity that was confirmed by CD and DLS assays. In addition, we found that the H3 mutE and H3 mutA peptides, unlike the H3 wt, were moderately and highly toxic, respectively, when exposed to human neuroblastoma cells. Cellular localization experiments confirmed that the mutated sequences hamper their nucleolar accumulation, and more importantly, that the helical conformation of the H3 region is crucial for such a localization.

Keywords: AML; CD spectroscopy; aggregation phenomena; helical peptides.

MeSH terms

  • Apoptosis
  • Cell Line, Tumor
  • Cell Nucleus / metabolism*
  • Cytosol / metabolism*
  • Humans
  • Leukemia, Myeloid, Acute / metabolism*
  • Nuclear Proteins / metabolism*
  • Nucleophosmin
  • Peptide Fragments
  • Protein Aggregation, Pathological
  • Protein Conformation
  • Protein Stability
  • Protein Transport

Substances

  • NPM1 protein, human
  • Nuclear Proteins
  • Peptide Fragments
  • Nucleophosmin