Simona Paladino

Researcher of Applied Biology

Name Simona
Surname Paladino
Institution University of Naples – Federico II
Address Department of Molecular Medicine and Medical Biotechnology, University of Naples Federico II, Naples, Italy
Simona Paladino


  • Golgi sorting regulates organization and activity of GPI proteins at apical membranes.

    Publication Date: 01/05/2014 on Nature chemical biology
    by Paladino S, Lebreton S, Tivodar S, Formiggini F, Ossato G, Gratton E, Tramier M, Coppey-Moisan M, Zurzolo C
    DOI: 10.1038/nchembio.1495

    Here we combined classical biochemistry with new biophysical approaches to study the organization of glycosylphosphatidylinositol (GPI)-anchored proteins (GPI-APs) with high spatial and temporal resolution at the plasma membrane of polarized epithelial cells. We show that in polarized MDCK cells, after sorting in the Golgi, each GPI-AP reaches the apical surface in homoclusters. Golgi-derived homoclusters are required for their subsequent plasma membrane organization into cholesterol-dependent heteroclusters. By contrast, in nonpolarized MDCK cells, GPI-APs are delivered to the surface as monomers in an unpolarized manner and are not able to form heteroclusters. We further demonstrate that this GPI-AP organization is regulated by the content of cholesterol in the Golgi apparatus and is required to maintain the functional state of the protein at the apical membrane. Thus, in contrast to fibroblasts, in polarized epithelial cells, a selective cholesterol-dependent sorting mechanism in the Golgi regulates both the organization and function of GPI-APs at the apical surface.

  • Resveratrol couples apoptosis with autophagy in UVB-irradiated HaCaT cells.

    Publication Date: 19/11/2013 on PloS one
    by Vitale N, Kisslinger A, Paladino S, Procaccini C, Matarese G, Pierantoni GM, Mancini FP, Tramontano D
    DOI: 10.1371/journal.pone.0080728

    UVB radiation causes about 90% of non-melanoma skin cancers by damaging DNA either directly or indirectly by increasing levels of reactive oxygen species (ROS). Skin, chronically exposed to both endogenous and environmental pro-oxidant agents, contains a well-organised system of chemical and enzymatic antioxidants. However, increased or prolonged free radical action can overwhelm ROS defence mechanisms, contributing to the development of cutaneous diseases. Thus, new strategies for skin protection comprise the use of food antioxidants to counteract oxidative stress. Resveratrol, a phytoalexin from grape, has gained a great interest for its ability to influence several biological mechanisms like redox balance, cell proliferation, signal transduction pathways, immune and inflammatory response. Therefore, the potential of resveratrol to modify skin cell response to UVB exposure could turn out to be a useful option to protect skin from sunlight-induced degenerative diseases. To investigate into this matter, HaCaT cells, a largely used model for human skin keratinocytes, were treated with 25 or 100 µM resveratrol for 2 and 24 hours prior to UVB irradiation (10 to 100 mJ/cm(2)). Cell viability and molecular markers of proliferation, oxidative stress, apoptosis, and autophagy were analyzed. In HaCaT cells resveratrol pretreatment: reduces UVB-induced ROS formation, enhances the detrimental effect of UVB on HaCaT cell vitality, increases UVB-induced caspase 8, PARP cleavage, and induces autophagy. These findings suggest that resveratrol could exert photochemopreventive effects by enhancing UVB-induced apoptosis and by inducing autophagy, thus reducing the odds that damaged cells could escape programmed cell death and initiate malignant transformation.

  • Translational control in the stress adaptive response of cancer cells: a novel role for the heat shock protein TRAP1.

    Publication Date: 10/10/2013 on Cell death & disease
    by Matassa DS, Amoroso MR, Agliarulo I, Maddalena F, Sisinni L, Paladino S, Romano S, Romano MF, Sagar V, Loreni F, Landriscina M, Esposito F
    DOI: 10.1038/cddis.2013.379

    TNF receptor-associated protein 1 (TRAP1), the main mitochondrial member of the heat shock protein (HSP) 90 family, is induced in most tumor types and is involved in the regulation of proteostasis in the mitochondria of tumor cells through the control of folding and stability of selective proteins, such as Cyclophilin D and Sorcin. Notably, we have recently demonstrated that TRAP1 also interacts with the regulatory protein particle TBP7 in the endoplasmic reticulum (ER), where it is involved in a further extra-mitochondrial quality control of nuclear-encoded mitochondrial proteins through the regulation of their ubiquitination/degradation. Here we show that TRAP1 is involved in the translational control of cancer cells through an attenuation of global protein synthesis, as evidenced by an inverse correlation between TRAP1 expression and ubiquitination/degradation of nascent stress-protective client proteins. This study demonstrates for the first time that TRAP1 is associated with ribosomes and with several translation factors in colon carcinoma cells and, remarkably, is found co-upregulated with some components of the translational apparatus (eIF4A, eIF4E, eEF1A and eEF1G) in human colorectal cancers, with potential new opportunities for therapeutic intervention in humans. Moreover, TRAP1 regulates the rate of protein synthesis through the eIF2α pathway either under basal conditions or under stress, favoring the activation of GCN2 and PERK kinases, with consequent phosphorylation of eIF2α and attenuation of cap-dependent translation. This enhances the synthesis of selective stress-responsive proteins, such as the transcription factor ATF4 and its downstream effectors BiP/Grp78, and the cystine antiporter system xCT, thereby providing protection against ER stress, oxidative damage and nutrient deprivation. Accordingly, TRAP1 silencing sensitizes cells to apoptosis induced by novel antitumoral drugs that inhibit cap-dependent translation, such as ribavirin or 4EGI-1, and reduces the ability of cells to migrate through the pores of transwell filters. These new findings target the TRAP1 network in the development of novel anti-cancer strategies.

  • TRAP1 and the proteasome regulatory particle TBP7/Rpt3 interact in the endoplasmic reticulum and control cellular ubiquitination of specific mitochondrial proteins.

    Publication Date: 01/04/2012 on Cell death and differentiation
    by Amoroso MR, Matassa DS, Laudiero G, Egorova AV, Polishchuk RS, Maddalena F, Piscazzi A, Paladino S, Sarnataro D, Garbi C, Landriscina M, Esposito F
    DOI: 10.1038/cdd.2011.128

    Tumor necrosis factor receptor-associated protein-1 (TRAP1) is a mitochondrial (MITO) antiapoptotic heat-shock protein. The information available on the TRAP1 pathway describes just a few well-characterized functions of this protein in mitochondria. However, our group's use of mass-spectrometric analysis identified TBP7, an AAA-ATPase of the 19S proteasomal subunit, as a putative TRAP1-interacting protein. Surprisingly, TRAP1 and TBP7 colocalize in the endoplasmic reticulum (ER), as demonstrated by biochemical and confocal/electron microscopic analyses, and interact directly, as confirmed by fluorescence resonance energy transfer analysis. This is the first demonstration of TRAP1's presence in this cellular compartment. TRAP1 silencing by short-hairpin RNAs, in cells exposed to thapsigargin-induced ER stress, correlates with upregulation of BiP/Grp78, thus suggesting a role of TRAP1 in the refolding of damaged proteins and in ER stress protection. Consistently, TRAP1 and/or TBP7 interference enhanced stress-induced cell death and increased intracellular protein ubiquitination. These experiments led us to hypothesize an involvement of TRAP1 in protein quality control for mistargeted/misfolded mitochondria-destined proteins, through interaction with the regulatory proteasome protein TBP7. Remarkably, expression of specific MITO proteins decreased upon TRAP1 interference as a consequence of increased ubiquitination. The proposed TRAP1 network has an impact in vivo, as it is conserved in human colorectal cancers, is controlled by ER-localized TRAP1 interacting with TBP7 and provides a novel model of the ER-mitochondria crosstalk.

  • Identification of sumoylation sites in CCDC6, the first identified RET partner gene in papillary thyroid carcinoma, uncovers a mode of regulating CCDC6 function on CREB1 transcriptional activity.

    Publication Date: 01/01/2012 on PloS one
    by Luise C, Merolla F, Leone V, Paladino S, Sarnataro D, Fusco A, Celetti A
    DOI: 10.1371/journal.pone.0049298

    CCDC6 was originally identified in chimeric genes as caused by chromosomal translocation involving the RET protooncogene in some thyroid tumors. Recognised as a 65 kDa pro-apoptotic phosphoprotein, CCDC6 has been enrolled as an ATM substrate that contribute to protect genome integrity by modulating PP4c activity in response to genotoxic stress. Recently, CCDC6 has been identified as a repressor of CREB1-dependent transcription. Sumoylation has emerged as an important mechanism in transcriptional control. Here, we report the identification and characterization of three sites of sumoylation in CCDC6 (K74, K266 and K424) which are highly conserved in vertebrates. We demonstrate that the post-translational modifications by SUMO2 constrain most of the CCDC6 protein in the cytosol and affect its functional interaction with CREB1 with a decrease of CCDC6 repressive function on CREB1 transcriptional activity. Indeed, the impairment of functional outcome of sumoylated CCDC6 is obtained knocking down all three the sumoylation sites. Interestingly, in thyroid cells the SUMO2-mediated CCDC6 post-translational modifications are induced by Forskolin, a cAMP analog. Signal transduction via the cAMP pathway is known to be ubiquitous and represents a major line of communication between many organisms and their environment. We believe that CCDC6 could be an important player in the dynamics of cAMP signaling by fine regulating CREB1 transcriptional activity in normal and transformed thyroid cells.

  • N-Glycosylation instead of cholesterol mediates oligomerization and apical sorting of GPI-APs in FRT cells.

    Publication Date: 01/12/2011 on Molecular biology of the cell
    by Imjeti NS, Lebreton S, Paladino S, de la Fuente E, Gonzalez A, Zurzolo C
    DOI: 10.1091/mbc.E11-04-0320

    Sorting of glycosylphosphatidyl-inositol--anchored proteins (GPI-APs) in polarized epithelial cells is not fully understood. Oligomerization in the Golgi complex has emerged as the crucial event driving apical segregation of GPI-APs in two different kind of epithelial cells, Madin-Darby canine kidney (MDCK) and Fisher rat thyroid (FRT) cells, but whether the mechanism is conserved is unknown. In MDCK cells cholesterol promotes GPI-AP oligomerization, as well as apical sorting of GPI-APs. Here we show that FRT cells lack this cholesterol-driven oligomerization as apical sorting mechanism. In these cells both apical and basolateral GPI-APs display restricted diffusion in the Golgi likely due to a cholesterol-enriched membrane environment. It is striking that N-glycosylation is the critical event for oligomerization and apical sorting of GPI-APs in FRT cells but not in MDCK cells. Our data indicate that at least two mechanisms exist to determine oligomerization in the Golgi leading to apical sorting of GPI-APs. One depends on cholesterol, and the other depends on N-glycosylation and is insensitive to cholesterol addition or depletion.

  • Endoplasmic reticulum stress reduces the export from the ER and alters the architecture of post-ER compartments.

    Publication Date: 01/12/2009 on The international journal of biochemistry & cell biology
    by Amodio G, Renna M, Paladino S, Venturi C, Tacchetti C, Moltedo O, Franceschelli S, Mallardo M, Bonatti S, Remondelli P
    DOI: 10.1016/j.biocel.2009.08.006

    In eukaryotic cells several physiologic and pathologic conditions generate the accumulation of unfolded proteins in the endoplasmic reticulum (ER), leading to ER stress. To restore normal function, some ER transmembrane proteins sense the ER stress and activate coordinated signalling pathways collectively called the Unfolded Protein Response (UPR). Little is known on how the UPR relates to post-ER compartments and to the export from the ER of newly synthesized proteins. Here, we report that the ER stress response induced by either thapsigargin or nitric oxide modifies the dynamics of the intracellular distribution of ERGIC-53 and GM130, two markers of the ER Golgi Intermediate Compartment and of the cis-Golgi, respectively. In addition, induction of ER stress alters the morphology of the ERGIC and the Golgi complex and interferes with the reformation of both compartments. Moreover, ER stress rapidly reduces the transport to the Golgi complex of the temperature sensitive mutant of the Vesicular Stomatitis Virus G Glycoprotein (VSV-G) fused with the Green Fluorescent Protein (ts045G), without apparently decreasing the amount of the protein competent for export. Interestingly, a parallel rapid reduction of the number of Sec31 labelled fluorescent puncta on the ER membranes does occur, thus suggesting that the ER stress alters the ER export and the dynamic of post-ER compartments by rapidly targeting the formation of COPII-coated transport intermediates.

  • Lipid rafts and clathrin cooperate in the internalization of PrP in epithelial FRT cells.

    Publication Date: 08/06/2009 on PloS one
    by Sarnataro D, Caputo A, Casanova P, Puri C, Paladino S, Tivodar SS, Campana V, Tacchetti C, Zurzolo C
    DOI: 10.1371/journal.pone.0005829

    The cellular prion protein (PrP(C)) plays a key role in the pathogenesis of Transmissible Spongiform Encephalopathies in which the protein undergoes post-translational conversion to the infectious form (PrP(Sc)). Although endocytosis appears to be required for this conversion, the mechanism of PrP(C) internalization is still debated, as caveolae/raft- and clathrin-dependent processes have all been reported to be involved.

  • Different GPI-attachment signals affect the oligomerisation of GPI-anchored proteins and their apical sorting.

    Publication Date: 15/12/2008 on Journal of cell science
    by Paladino S, Lebreton S, Tivodar S, Campana V, Tempre R, Zurzolo C
    DOI: 10.1242/jcs.036038

    To understand the mechanism involved in the apical sorting of glycosylphosphatidylinositol (GPI)-anchored proteins (GPI-APs) we fused to the C-terminus of GFP the GPI-anchor-attachment signal of the folate receptor (FR) or of the prion protein (PrP), two native GPI-anchored proteins that are sorted apically or basolaterally, respectively, in MDCK cells. We investigated the behaviour of the resulting fusion proteins GFP-FR and GFP-PrP by analysing three parameters: their association with DRMs, their oligomerisation and their apical sorting. Strikingly, we found that different GPI-attachment signals differently modulate the ability of the resulting GFP-fusion protein to oligomerise and to be apically sorted. This is probably owing to differences in the GPI anchor and/or in the surrounding lipid microenvironment. Accordingly, we show that addition of cholesterol to the cells is necessary and sufficient to drive the oligomerisation and consequent apical sorting of GFP-PrP, which under control conditions does not oligomerise and is basolaterally sorted.

  • N- and O-glycans are not directly involved in the oligomerization and apical sorting of GPI proteins.

    Publication Date: 01/12/2008 on Traffic (Copenhagen, Denmark)
    by Catino MA, Paladino S, Tivodar S, Pocard T, Zurzolo C
    DOI: 10.1111/j.1600-0854.2008.00826.x

    Oligomerization of glycosyl-phosphatidylinositol-anchored proteins (GPI-APs) into high molecular weight complexes is an essential step for their apical sorting in polarized epithelial cells. However, the mechanism by which apical GPI-APs oligomerize is still unclear. To investigate the possible role of N- and O-glycosylation, we have analysed the behaviour of two glycosylated GPI-anchored apical proteins, p75GPI and placental alkaline phosphatase (PLAP), and their glycosylation mutants. We found that both the N- and O-glycosylation mutants are apically sorted, associate to detergent-resistant microdomains and are able to oligomerize, like the wild-type proteins, suggesting that glycosylation does not have a direct role in GPI-AP oligomerization and apical sorting. Interestingly, when cells are depleted of cholesterol and treated with tunicamycin, treatments that by themselves do not affect PLAP sorting, PLAP is not able to oligomerize and is missorted to the basolateral surface, thus supporting an indirect role of N-glycosylation, possibly mediated by a raft-associated glycosylated interactor.