Publications (7)18.15 Total impact
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Article: Novel cell death pathways induced by N-(4-hydroxyphenyl)retinamide: therapeutic implications.
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ABSTRACT: We previously reported that N-(4-hydroxyphenyl)retinamide (4HPR) inhibits retinoblastoma tumor growth in a murine model in vivo and kills Y79 retinoblastoma cells in vitro. In this work, we assayed different cell death-related parameters, including mitochondrial damage and caspase activation, in Y79 cells exposed to 4HPR. 4HPR induced cytochrome c release from mitochondria, caspase-3 activation, and oligonucleosomal DNA fragmentation. However, pharmacologic inactivation of caspases by the pan-caspase inhibitor BOC-D-fmk, or specific caspase-3 inhibition by Z-DEVD-fmk, was not sufficient to prevent cell death, as assessed by loss of 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide reduction, lactate dehydrogenase release, disruption of mitochondrial transmembrane potential (Deltapsi(m)), and ATP depletion. We found that 4HPR causes lysosomal membrane permeabilization and cytosolic relocation of cathepsin D. Pepstatin A partially rescued cell viability and reduced DNA fragmentation and cytosolic cytochrome c. The antioxidant N-acetylcysteine attenuated cathepsin D relocation into the cytosol, suggesting that lysosomal destabilization is dependent on elevation of reactive oxygen species and precedes mitochondrial dysfunction. Activation of AKT, which regulates energy level in the cell, by the retinal survival facto]r insulin-like growth factor I was impaired and insulin-like growth factor I was ineffective against ATP and Deltapsi(m) loss in the presence of 4HPR. Lysosomal destabilization, associated with mitochondrial dysfunction, was induced by 4HPR also in other cancer cell lines, including PC3 prostate adenocarcinoma and the vascular tumor Kaposi sarcoma KS-Imm cells. The novel finding of a lysosome-mediated cell death pathway activated by 4HPR could have implications at clinical level for the development of combination chemoprevention and therapy of cancer.Molecular Cancer Therapeutics 02/2007; 6(1):286-98. · 5.23 Impact Factor -
Article: SCN- binding to the charged lysines of histones end domains mimics acetylation and shows the major histone-DNA interactions involved in eu and heterochromatin stabilization.
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ABSTRACT: SCN- binds to the charged amino group of lysines, inducing local changes in the electrostatic free energy of histones. We exploited this property to selectively perturb the histone-DNA interactions involved in the stabilization of eu and heterochromatin. Differential scanning calorimetry (DSC) was used as leading technique in combination with trypsin digestion that selectively cleaves the histone end domains. Euchromatin undergoes progressive destabilization with increasing KSCN concentration from 0 to 0.3 M. Trypsin digestion in the presence of 0.2 M KSCN show that the stability of the linker decreases as a consequence of the competitive binding of SCN- to the amino groups located in the C and N-terminal domain of H1 and H3, respectively; likewise, the release of the N-terminal domain of H4 induces an appreciable depression in both the temperature and enthalpy of melting of core particle DNA. Unfolding of heterochromatin requires, in addition to further cleavage of H4, extensive digestion of H2A and H2B, strongly suggesting that these histones stabilize the higher order structure by forming a protein network which extends throughout the heterochromatin domain.Journal of Cellular Biochemistry 04/2006; 97(4):869-81. · 2.87 Impact Factor -
Article: Anti-keratin monoclonal antibodies for identifying animal hair fibers
Textile Research Journal 01/2004; 74(5):458-464. · 1.12 Impact Factor -
Article: DNA supercoiling in apoptotic chromatin.
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ABSTRACT: In a previous paper, we have reported that in rat thymocyte apoptosis chromatin undergoes a specific structural change as well as an appreciable increase in the unacetylated forms of histones H3 and H4. Here, we show that H3 and H4 deacetylation bears no relation to chromatin condensation, and present new ultrastructural and topological observations that largely clarify the organization of the condensed state. The texture of the latter corresponds to a closely woven network of negatively supercoiled 11 nm fibers, as shown by both ultrastructural observations and relaxation experiments using ethidium bromide. Circularly closed chromatin loops undergoing apoptotic condensation, clearly showing nucleosome compact dimers or higher oligomers, as well as long stretches of supercoiled DNA, have also been detected. All of these modifications are strongly reminiscent of the alterations induced in nucleosome bearing plasmids by the chromatin remodeling factors SWI/SNF and RSC.Biochemical and Biophysical Research Communications 10/2003; 309(3):540-6. · 2.48 Impact Factor -
Article: An intranuclear frame for chromatin compartmentalization and higher-order folding.
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ABSTRACT: Recent ultrastructural, immunoelectron, and confocal microscopy observations done in our laboratory [Barboro et al. [2002] Exp. Cell. Res. 279:202-218] have confirmed that lamins and the nuclear mitotic apparatus protein (NuMA) are localized inside the interphase nucleus in a polymerized form. This provided evidence of the existence of a RNA stabilized lamin/NuMA frame, consisting of a web of thin ( approximately 3 and approximately 5 nm) lamin filaments to which NuMA is anchored mainly in the form of discrete islands, which might correspond to the minilattices described by Harborth et al. [1999] (EMBO. J. 18:1689-1700). In this article we propose that this scaffold is involved in the compartmentalization of both chromatin and functional domains and further determines the higher-order nuclear organization. This hypothesis is strongly supported by the scrutiny of different structural transitions which occur inside the nucleus, such as chromatin displacement and rearrangements, the collapse of the internal nuclear matrix after RNA digestion and the disruption of chromosome territories induced by RNase A and high salt treatment. All of these destructive events directly depend on the loss of the stabilizing effect exerted on the different levels of structural organization by the interaction of RNA with lamins and/or NuMA. Therefore, the integrity of nuclear RNA must be safeguarded as far as possible to isolate the matrix in the native form. This material will allow for the first time the unambiguous ultrastructural localization inside the INM of the components of the functional domains, so opening new avenues of investigation on the mechanisms of gene expression in eukaryotes.Journal of Cellular Biochemistry 02/2003; 88(1):113-20. · 2.87 Impact Factor -
Article: Unraveling the organization of the internal nuclear matrix: RNA-dependent anchoring of NuMA to a lamin scaffold.
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ABSTRACT: Using quantitative immunoelectron microscopy we show here that when the nuclear matrix is isolated from rat hepatocytes in the presence of an inhibitor of RNase activity both lamins and the nuclear mitotic apparatus protein (NuMA) preferentially localize within the electron-dense domains of the internal nuclear matrix (INM). After RNA digestion NuMA undergoes a sharp depletion, while labeling by an antibody against lamins A and C within the electron-transparent regions increases, suggesting that a subset of lamin epitopes is masked by the interaction with RNA. We were able to explain this result by visualizing for the first time a thin web of lamin protofibrils which connects the electron-dense regions. Confirmation of these changes has been obtained by immunoblot analysis and confocal microscopy. As RNA digestion results both in the release of NuMA and in the collapse of the INM, we propose that a fraction of nuclear RNA brings about the association of NuMA islands with a lamin scaffold and that this interaction is required to maintain the latter in a state of high molecular dispersion.Experimental Cell Research 11/2002; 279(2):202-18. · 3.58 Impact Factor -
Article: DNA supercoiling in apoptotic chromatin
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ABSTRACT: In a previous paper, we have reported that in rat thymocyte apoptosis chromatin undergoes a specific structural change as well as an appreciable increase in the unacetylated forms of histones H3 and H4. Here, we show that H3 and H4 deacetylation bears no relation to chromatin condensation, and present new ultrastructural and topological observations that largely clarify the organization of the condensed state. The texture of the latter corresponds to a closely woven network of negatively supercoiled 11 nm fibers, as shown by both ultrastructural observations and relaxation experiments using ethidium bromide. Circularly closed chromatin loops undergoing apoptotic condensation, clearly showing nucleosome compact dimers or higher oligomers, as well as long stretches of supercoiled DNA, have also been detected. All of these modifications are strongly reminiscent of the alterations induced in nucleosome bearing plasmids by the chromatin remodeling factors SWI/SNF and RSC.Biochemical and Biophysical Research Communications.
Top co-authors
Top Journals
Institutions
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2003–2006
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Istituto per lo Studio delle Macromolecole
Biella, Piedmont, Italy
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2002–2003
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National Institute for Research on Cancer
Genova, Liguria, Italy
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