CNR Institute of Molecular Biology and Pathology


CNR Institute of Molecular Biology and Pathology

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Giri R.,CNR Institute of Molecular Biology and Pathology | Morrone A.,CNR Institute of Molecular Biology and Pathology | Toto A.,CNR Institute of Molecular Biology and Pathology | Brunori M.,CNR Institute of Molecular Biology and Pathology | Gianni S.,CNR Institute of Molecular Biology and Pathology
Proceedings of the National Academy of Sciences of the United States of America | Year: 2013

A classical dogma of molecular biology dictates that the 3D structure of a protein is necessary for its function. However, a considerable fraction of the human proteome, although functional, does not adopt a defined folded state under physiological conditions. These intrinsically disordered proteins tend to fold upon binding to their partners with a molecular mechanism that is elusive to experimental characterization. Indeed, although many hypotheses have been put forward, the functional role (if any) of disorder in these intrinsically denatured systems is still shrouded in mystery. Here, we characterize the structure of the transition state of the binding-induced folding in the reaction between the KIX domain of the CREB-binding protein and the transactivation domain of c-Myb. The analysis, based on the characterization of a series of conservative site-directed mutants, reveals a very high content of native-like structure in the transition state and indicates that the recognition between KIX and c-Myb is geometrically precise. The implications of our results in the light of previous work on intrinsically unstructured systems are discussed.

Giri R.,CNR Institute of Molecular Biology and Pathology | Morrone A.,CNR Institute of Molecular Biology and Pathology | Travaglini-Allocatelli C.,CNR Institute of Molecular Biology and Pathology | Jemth P.,Uppsala University | And 2 more authors.
Proceedings of the National Academy of Sciences of the United States of America | Year: 2012

Much experimental work has been devoted in comparing the folding behavior of proteins sharing the same fold but different sequence. The recent design of proteins displaying very high sequence identities but different 3D structure allows the unique opportunity to address the protein-folding problem from a complementary perspective. Here we explored by ℙ-value analysis the pathways of folding of three different heteromorphic pairs, displaying increasingly high-sequence identity (namely, 30%, 77%, and 88%), but different structures called GA (a 3-α helix fold) and GB (an α/β fold). The analysis, based on 132 site-directed mutants, is fully consistent with the idea that protein topology is committed very early along the pathway of folding. Furthermore, data reveals that when folding approaches a perfect two-state scenario, as in the case of the GA domains, the structural features of the transition state appear very robust to changes in sequence composition. On the other hand, when folding is more complex and multistate, as for the GBs, there are alternative nuclei or accessible pathways that can be alternatively stabilized by altering the primary structure. The implications of our results in the light of previous work on the folding of different members belonging to the same protein family are discussed.

Kostiainen M.A.,Aalto University | Hiekkataipale P.,Aalto University | Laiho A.,Aalto University | Lemieux V.,St Jean Photochimie SJPC | And 3 more authors.
Nature Nanotechnology | Year: 2013

Binary nanoparticle superlattices are periodic nanostructures with lattice constants much shorter than the wavelength of light1,2 and could be used to prepare multifunctional metamaterials3,4. Such superlattices are typically made from synthetic nanoparticles5-8, and although biohybrid structures have been developed9-15, incorporating biological building blocks into binary nanoparticle superlattices remains challenging16-18. Protein-based nanocages provide a complex yet monodisperse and geometrically well-defined hollow cage that can be used to encapsulate different materials19,20. Such protein cages have been used to program the self-assembly of encapsulated materials to form free-standing crystals21,22 and superlattices at interfaces 21,23 or in solution24,25. Here, we show that electrostatically patchy protein cages-cowpea chlorotic mottle virus and ferritin cages-can be used to direct the self-assembly of three-dimensional binary superlattices. The negatively charged cages can encapsulate RNA or superparamagnetic iron oxide nanoparticles, and the superlattices are formed through tunable electrostatic interactions with positively charged gold nanoparticles. Gold nanoparticles and viruses form an AB8 fcc crystal structure that is not isostructural with any known atomic or molecular crystal structure and has previously been observed only with large colloidal polymer particles26. Gold nanoparticles and empty or nanoparticle-loaded ferritin cages form an interpenetrating simple cubic AB structure (isostructural with CsCl). We also show that these magnetic assemblies provide contrast enhancement in magnetic resonance imaging. © 2013 Macmillan Publishers Limited. All rights reserved.

D'Avino P.P.,University of Cambridge | Giansanti M.G.,CNR Institute of Molecular Biology and Pathology | Petronczki M.,Cancer Research UK Research Institute
Cold Spring Harbor Perspectives in Biology | Year: 2015

Cell division ends with the physical separation of the two daughter cells, a process known as cytokinesis. This final event ensures that nuclear and cytoplasmic contents are accurately partitioned between the two nascent cells. Cytokinesis is one of the most dramatic changes in cell shape and requires an extensive reorganization of the cell’s cytoskeleton. Here, we describe the cytoskeletal structures, factors, and signaling pathways that orchestrate this robust and yet highly dynamic process in animal cells. Finally, we discuss possible future directions in this growing area of cell division research and its implications in human diseases, including cancer. © 2015 Cold Spring Harbor Laboratory Press; all rights reserved.

Caffarelli E.,CNR Institute of Molecular Biology and Pathology | Filetici P.,CNR Institute of Molecular Biology and Pathology
Frontiers in Bioscience | Year: 2011

From an operational definition of epigenetic, we move to provide the reader a general but comprehensive description of epigenetic phenomena that often lead to cell transformation. The last decade has, in fact, seen novel players involved in the regulation of gene expression. Not only protein factors but also a number of chromatin modifiers and remodelling proteins, which regulate the level of compaction of the genome through a variety of post-translational modifications deposed on histone tails or on DNA itself. Meanwhile, the discovery of tiny RNAs, of only 21-23 nucleotides in length, has brought to the attention their role as key regulators in the cell, being able to direct differentiation programs and function as oncogenes or oncosuppressors. In this general compendium, we aim to describe main cellular functions that through an epigenetic or epigenetic associated mechanism have been found to be directly implicated in cancerogenesis.

Di Domenico F.,University of Rome La Sapienza | Foppoli C.,CNR Institute of Molecular Biology and Pathology | Coccia R.,University of Rome La Sapienza | Perluigi M.,University of Rome La Sapienza
Biochimica et Biophysica Acta - Molecular Basis of Disease | Year: 2012

Cervical cancer lesions are a major threat to the health of women, representing the second most common cancer worldwide. The unanimously recognized etiological factor in the causation of cervical cancer is the infection with human papilloma virus (HPV). HPV infection, although necessary, is not per se sufficient to induce cancer. Other factors have to be involved in the progression of infected cells to the full neoplastic phenotype. Oxidative stress represents an interesting and under-explored candidate as a promoting factor in HPV-initiated carcinogenesis. Oxidative stress is known to perturb the cellular redox status thus leading to alteration of gene expression responses through the activation of several redox-sensitive transcription factors. This signaling cascade affects both cell growth and cell death. The ability of naturally occurring antioxidants to modulate cellular signal transduction pathways, through the activation/repression of multiple redox-sensitive transcription factors, has been claimed for their potential therapeutic use as chemopreventive agents. Among these compounds, polyphenols have been found to be promising agents toward cervical cancer. In addition to acting as antioxidants, polyphenols display a wide variety of biological function including induction of apoptosis, growth arrest, inhibition of DNA synthesis and modulation of signal transduction pathways. They can interfere with each stage of carcinogenesis initiation, promotion and progression to prevent cancer development. The present review discusses current knowledge of the major molecular pathways, which are involved in HPV-driven cancerogenesis, and the ability of polyphenols to modulate these pathways. By acting at specific steps of viral transformation cascade, polyphenols have been demonstrated to selectively inhibit tumor cell growth and may be a promising therapeutic tool for treatment of cervical cancer. In addition, recent results obtained in clinical trials using polyphenols are also discussed. This article is part of a Special Issue entitled: Antioxidants and Antioxidant Treatment in Disease. © 2011 Elsevier B.V.

Giansanti M.G.,CNR Institute of Molecular Biology and Pathology | Fuller M.T.,Stanford University
Cytoskeleton | Year: 2012

Cytokinesis separates the genomic material and organelles of a dividing cell equitably into two physically distinct daughter cells at the end of cell division. This highly choreographed process involves coordinated reorganization and regulated action of the actin and microtubule cytoskeletal systems, an assortment of motor proteins, and membrane trafficking components. Due to their large size, the ease with which exquisite cytological analysis may be performed on them, and the availability of numerous mutants and other genetic tools, Drosophila spermatocytes have provided an excellent system for exploring the mechanistic basis for the temporally programmed and precise spatially localized events of cytokinesis. Mutants defective in male meiotic cytokinesis can be easily identified in forward genetic screens by the production of multinucleate spermatids. In addition, the weak spindle assembly checkpoint in spermatocytes, which causes only a small delay of anaphase onset in the presence of unattached chromosomes, allows investigation of whether gene products required for spindle assembly and chromosome segregation are also involved in cytokinesis. Perhaps due to the large size of spermatocytes and the requirement for two rapid-fire rounds of division without intervening S or growth phases during meiosis, male meiotic mutants have also revealed much about molecular mechanisms underlying new membrane addition during cytokinesis. Finally, cell type-specific differences in the events that set up and complete cytokinesis are emerging from comparison of spermatocytes with cells undergoing mitosis either elsewhere in the organism or in tissue culture. © 2012 Wiley Periodicals, Inc.

Dogan J.,Uppsala University | Gianni S.,CNR Institute of Molecular Biology and Pathology | Gianni S.,University of Cambridge | Jemth P.,Uppsala University
Physical Chemistry Chemical Physics | Year: 2014

Intrinsically disordered proteins (IDPs) and intrinsically disordered regions (IDRs) of proteins are very common and instrumental for cellular signaling. Recently, a number of studies have investigated the kinetic binding mechanisms of IDPs and IDRs. These results allow us to draw conclusions about the energy landscape for the coupled binding and folding of disordered proteins. The association rate constants of IDPs cover a wide range (10 5-109 M-1 s-1) and are largely governed by long-range charge-charge interactions, similarly to interactions between well-folded proteins. Off-rate constants also differ significantly among IDPs (with half-lives of up to several minutes) but are usually around 0.1-1000 s-1, allowing for rapid dissociation of complexes. Likewise, affinities span from pM to μM suggesting that the low-affinity high-specificity concept for IDPs is not straightforward. Overall, it appears that binding precedes global folding although secondary structure elements such as helices may form before the protein-protein interaction. Short IDPs bind in apparent two-state reactions whereas larger IDPs often display complex multi-step binding reactions. While the two extreme cases of two-step binding (conformational selection and induced fit) or their combination into a square mechanism is an attractive model in theory, it is too simplistic in practice. Experiment and simulation suggest a more complex energy landscape in which IDPs bind targets through a combination of conformational selection before binding (e.g., secondary structure formation) and induced fit after binding (global folding and formation of short-range intermolecular interactions). © 2014 the Partner Organisations.

Raffa G.D.,CNR Institute of Molecular Biology and Pathology
Nucleus (Austin, Tex.) | Year: 2011

In most organisms, telomeres are extended by telomerase and contain GC-rich repeats. Drosophila telomeres are elongated by occasional transposition of specialized retroelements rather than telomerase activity, and are assembled independently of the sequence of the DNA termini. Recent work has shown that Drosophila telomeres are capped by a complex, we call terminin, which includes HOAP, HipHop, Moi and Ver; these are fast-evolving proteins that prevent telomere fusion, directly interact with each other, and appear to localize and function only at telomeres. With the possible exception of Ver that contains an OB fold domain structurally similar to the Stn1 OB fold, none of the terminin proteins is evolutionarily conserved outside the Drosophila species. Human telomeres are protected by the shelterin complex, which comprises six proteins that bind chromosome ends in a sequence-dependent manner. Shelterin subunits are not fast-evolving proteins and are not conserved in flies, but localize and function only at telomeres like the terminin components. Based on these findings, we propose that concomitant with telomerase loss Drosophila rapidly evolved terminin to bind chromosome ends in a sequence-independent fashion, and that terminin is functionally analogous to shelterin.

Lavia P.,CNR Institute of Molecular Biology and Pathology
Chromosome Research | Year: 2016

Growing lines of evidence implicate the small GTPase RAN, its regulators and effectors—predominantly, nuclear transport receptors—in practically all aspects of centrosome biology in mammalian cells. These include duplication licensing, cohesion, positioning, and microtubule-nucleation capacity. RAN cooperates with the protein nuclear export vector exportin 1/CRM1 to recruit scaffolding proteins containing nuclear export sequences that play roles in the structural organization of centrosomes. Together, they also limit centrosome reduplication by regulating the localization of key “licensing” proteins during the centrosome duplication cycle. In parallel, RAN also regulates the capacity of centrosomes to nucleate and organize functional microtubules, and this predominanlty involves importin vectors: many factors regulating microtubule nucleation or function harbor nuclear localization sequences that interact with importin molecules and such interaction inhibits their activity. Active RANGTP binding to importin molecules removes the inhibition and releases microtubule regulatory factors in the free productive form. A dynamic scenario emerges, in which RAN is pivotal in linking spatiotemporal control of centrosome regulators to the cell cycle machinery. © 2015, Springer Science+Business Media Dordrecht.

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