Synergic Role of Nucleophosmin Three-helix Bundle and a Flanking Unstructured Tail in the Interaction with G-quadruplex DNA

Arcovito, Alessandro; Chiarella, Sara; Longa, Stefano Della; Matteo, Adele Di; Sterzo, Claudio Lo; Scaglione, Giovanni Luca; Federici, L.

doi:10.1074/jbc.m114.565010

Cited by 20 publications

(12 citation statements)

References 35 publications

(61 reference statements)

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“…The CTD of NPM1 is an essential element for the activities of this multifunction protein. It is crucial for shuttling and for its interactions with DNA (6, 23, 31), and its functionality relies on the specific fold that it adopts: a 3‐helix bundle. Investigations have indicated that the 3 helical regions of the domain are endowed with a high intrinsic propensity for this structural element (9, 12).…”

Section: Discussionmentioning

confidence: 99%

Nucleophosmin contains amyloidogenic regions that are able to form toxic aggregates under physiological conditions

et al. 2015

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Nucleophosmin (NPM)-1 is a multifunctional protein involved in a variety of biologic processes and has been implicated in the pathogenesis of several human malignancies. To gain insight into the role of isolated fragments in NPM1 activities, we dissected the C-terminal domain (CTD) into its helical fragments. In this study, we observed the unexpected structural behavior of the peptide fragment corresponding to helix (H)2 (residues 264-277). This peptide has a strong tendency to form amyloidlike assemblies endowed with fibrillar morphology and β-sheet structure, under physiologic conditions, as shown by circular dichroism, thioflavin T, and Congo red binding assays; dynamic light scattering; and atomic force microscopy. The aggregates are also toxic to neuroblastoma cells, as determined using 3-(4;5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide reduction and Ca(2+) influx assays. We also found that the extension of the H2 sequence beyond its N terminus, comprising the connecting loop with H1, delayed aggregation and its associated cytotoxicity, suggesting that contiguous regions of H2 have a protective role in preventing aggregation. Our findings and those in the literature suggest that the helical structures present in the CTD are important in preventing harmful aggregation. These findings could elucidate the pathogenesis of acute myeloid leukemia (AML) caused by NPM1 mutants. Because the CTD is not properly folded in these mutants, we hypothesize that the aggregation propensity of this NPM1 region is involved in the pathogenesis of AML. Preliminary assays on NPM1-Cter-MutA, the most frequent AML-CTD mutation, revealed its significant propensity for aggregation. Thus, the aggregation phenomena should be seriously considered in studies aimed at unveiling the molecular mechanisms of this pathology.

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Section: Discussionmentioning

confidence: 99%

Nucleophosmin contains amyloidogenic regions that are able to form toxic aggregates under physiological conditions

et al. 2015

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“…It was shown that the three-helix bundle engages the G-quadruplex phosphate scaffold with a positively charged groove located between helices H1 and H2 (Figure 1C). Structural studies also revealed that the terminal part of the central domain, which is unstructured and markedly positively charged, is also necessary for high affinity binding, through both long range electrostatic effects and transient interactions with the G-quadruplex [44, 45]. NPM1 loses its nucleolar localization following lysine acetylation played by p300 [46] and, consistently with structural studies, both lysine residues located in the three-helix bundle at the G-quadruplex interface (Lys250, Lys257 and Lys267) [43] and lysine residues located in the flanking unstructured tail (Lys229 and Lys230) are acetylated by p300 [46].…”

Section: Npm1 Structurementioning

confidence: 99%

Molecules that target nucleophosmin for cancer treatment: an update

et al. 2016

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Nucleophosmin is a highly and ubiquitously expressed protein, mainly localized in nucleoli but able to shuttle between nucleus and cytoplasm. Nucleophosmin plays crucial roles in ribosome maturation and export, centrosome duplication, cell cycle progression, histone assembly and response to a variety of stress stimuli. Much interest in this protein has arisen in the past ten years, since the discovery of heterozygous mutations in the terminal exon of the NPM1 gene, which are the most frequent genetic alteration in acute myeloid leukemia. Nucleophosmin is also frequently overexpressed in solid tumours and, in many cases, its overexpression correlates with mitotic index and metastatization. Therefore it is considered as a promising target for the treatment of both haematologic and solid malignancies. NPM1 targeting molecules may suppress different functions of the protein, interfere with its subcellular localization, with its oligomerization properties or drive its degradation. In the recent years, several such molecules have been described and here we review what is currently known about them, their interaction with nucleophosmin and the mechanistic basis of their toxicity. Collectively, these molecules exemplify a number of different strategies that can be adopted to target nucleophosmin and we summarize them at the end of the review.

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“…Several investigations pointed out that the CTD of NPM1 is crucial to specifically recognize G-quadruplex DNA motifs [ 74 , 75 , 76 , 77 , 78 , 79 ]. Initial contrasting NMR analyses established that G-quadruplex recognition by NPM1 was primarily due to residues belonging to the helices H1 and H2 of the CTD [ 75 ], but more recently molecular dynamics simulations and SPR data indicated that the unstructured region plays a primary role in the mechanism.…”

Section: Dna Binding Proteinsmentioning

confidence: 99%

“…Initial contrasting NMR analyses established that G-quadruplex recognition by NPM1 was primarily due to residues belonging to the helices H1 and H2 of the CTD [ 75 ], but more recently molecular dynamics simulations and SPR data indicated that the unstructured region plays a primary role in the mechanism. Besides, facilitating the formation of the DNA-complex through long range electrostatic interactions, it directly contacts the G-quadruplex scaffold through multiple and transient electrostatic interactions significantly enlarging the contact surface [ 78 ].…”

Section: Dna Binding Proteinsmentioning

confidence: 99%

Identification of Inhibitors of Biological Interactions Involving Intrinsically Disordered Proteins

Marasco

Scognamiglio

2015

IJMS

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Protein–protein interactions involving disordered partners have unique features and represent prominent targets in drug discovery processes. Intrinsically Disordered Proteins (IDPs) are involved in cellular regulation, signaling and control: they bind to multiple partners and these high-specificity/low-affinity interactions play crucial roles in many human diseases. Disordered regions, terminal tails and flexible linkers are particularly abundant in DNA-binding proteins and play crucial roles in the affinity and specificity of DNA recognizing processes. Protein complexes involving IDPs are short-lived and typically involve short amino acid stretches bearing few “hot spots”, thus the identification of molecules able to modulate them can produce important lead compounds: in this scenario peptides and/or peptidomimetics, deriving from structure-based, combinatorial or protein dissection approaches, can play a key role as hit compounds. Here, we propose a panoramic review of the structural features of IDPs and how they regulate molecular recognition mechanisms focusing attention on recently reported drug-design strategies in the field of IDPs.

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Synergic Role of Nucleophosmin Three-helix Bundle and a Flanking Unstructured Tail in the Interaction with G-quadruplex DNA

Cited by 20 publications

References 35 publications

Nucleophosmin contains amyloidogenic regions that are able to form toxic aggregates under physiological conditions

Nucleophosmin contains amyloidogenic regions that are able to form toxic aggregates under physiological conditions

Molecules that target nucleophosmin for cancer treatment: an update

Identification of Inhibitors of Biological Interactions Involving Intrinsically Disordered Proteins

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