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Kay Huebner - One of the best experts on this subject based on the ideXlab platform.
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loss of fragile histidine triad FHIT protein expression alters the translation of cancer associated mrnas
BMC Research Notes, 2018Co-Authors: Daniel L. Kiss, Kay Huebner, Ralf Bundschuh, William D Baez, Daniel R. SchoenbergAbstract:In > 50% of cancers tumor development involves the early loss of FHIT (fragile histidine triad) protein expression, yet the mechanistic pathway(s) by which FHIT mediates its tumor suppressor functions are not fully understood. Earlier attempts to identify a FHIT-deficient gene expression profile relied on total cellular RNA and microarray analysis. The data here used RNA sequencing (RNA-Seq) of FHIT-negative and FHIT-positive cells as proof of principle for the impact of FHIT on specific mRNAs, and to lay the foundation for a study using ribosome profiling to identify mRNAs whose translation is affected by FHIT loss. RNA-Seq was performed on RNA from lines of FHIT-expressing and FHIT-deficient lung cancer cells. This identified changes in the levels of mRNAs for a number of cell survival and cell cycle progression genes. Polysome profile analysis performed on cytoplasmic extracts from FHIT-negative and FHIT-positive cells showed changes in the sedimentation of select mRNAs consistent with changes in translation efficiency. The impact of differential FHIT expression on the turnover of selected cancer-linked mRNAs was determined by RT-qPCR of cytoplasmic RNA isolated at intervals after treating cells with a transcription inhibitor.
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Loss of fragile histidine triad (FHIT) protein expression alters the translation of cancer-associated mRNAs
BMC, 2018Co-Authors: Daniel L. Kiss, Kay Huebner, Ralf Bundschuh, William D Baez, Daniel R. SchoenbergAbstract:Abstract Objectives In > 50% of cancers tumor development involves the early loss of FHIT (fragile histidine triad) protein expression, yet the mechanistic pathway(s) by which FHIT mediates its tumor suppressor functions are not fully understood. Earlier attempts to identify a FHIT-deficient gene expression profile relied on total cellular RNA and microarray analysis. The data here used RNA sequencing (RNA-Seq) of FHIT-negative and FHIT-positive cells as proof of principle for the impact of FHIT on specific mRNAs, and to lay the foundation for a study using ribosome profiling to identify mRNAs whose translation is affected by FHIT loss. Data description RNA-Seq was performed on RNA from lines of FHIT-expressing and FHIT-deficient lung cancer cells. This identified changes in the levels of mRNAs for a number of cell survival and cell cycle progression genes. Polysome profile analysis performed on cytoplasmic extracts from FHIT-negative and FHIT-positive cells showed changes in the sedimentation of select mRNAs consistent with changes in translation efficiency. The impact of differential FHIT expression on the turnover of selected cancer-linked mRNAs was determined by RT-qPCR of cytoplasmic RNA isolated at intervals after treating cells with a transcription inhibitor
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exome wide single base substitutions in tissues and derived cell lines of the constitutive FHIT knockout mouse
Cancer Science, 2016Co-Authors: Carolyn Paisie, Teresa Druck, Catherine E. Waters, Iman M. Ouda, Joshua C. Saldivar, Jenna R. Karras, Satoshi Miuma, Morgan S Schrock, Jie Zhang, Kay HuebnerAbstract:Loss of expression of FHIT, a tumor suppressor and genome caretaker, occurs in preneoplastic lesions during development of many human cancers. Furthermore, FHIT‐deficient mouse models are exquisitely susceptible to carcinogen induction of cancers of the lung and forestomach. Due to absence of FHIT genome caretaker function, cultured cells and tissues of the constitutive FHIT knockout strain develop chromosome aneuploidy and allele copy number gains and losses and we hypothesized that FHIT‐deficient cells would also develop point mutations. On analysis of whole exome sequences of FHIT‐deficient tissues and cultured cells, we found 300 to >1000 single‐base substitutions associated with FHIT loss in the 2% of the genome included in exomes, relative to the C57Bl6 reference genome. The mutation signature is characterized by increased C>T and T>C mutations, similar to the “age at diagnosis” signature identified in human cancers. The FHIT‐deficiency mutation signature also resembles a C>T and T>C mutation signature reported for human papillary kidney cancers and a similar signature recently reported for esophageal and bladder cancers, cancers that are frequently FHIT deficient. The increase in T>C mutations in −/− exomes may be due to dNTP imbalance, particularly in thymidine triphosphate, resulting from decreased expression of thymidine kinase 1 in FHIT‐deficient cells. FHIT‐deficient kidney cells that survived in vitro dimethylbenz(a)anthracene treatment additionally showed increased T>A mutations, a signature generated by treatment with this carcinogen, suggesting that these T>A transversions may be evidence of carcinogen‐induced preneoplastic changes.
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The FHIT gene product: tumor suppressor and genome “caretaker”
Cellular and Molecular Life Sciences, 2014Co-Authors: Catherine E. Waters, Joshua C. Saldivar, Seyed Ali Hosseini, Kay HuebnerAbstract:The FHIT gene at FRA3B is one of the earliest and most frequently altered genes in the majority of human cancers. It was recently discovered that the FHIT gene is not the most fragile locus in epithelial cells, the cell of origin for most FHIT-negative cancers, eroding support for past claims that deletions at this locus are simply passenger events that are carried along in expanding cancer clones, due to extreme vulnerability to DNA damage rather than to loss of FHIT function. Indeed, recent reports have reconfirmed FHIT as a tumor suppressor gene with roles in apoptosis and prevention of the epithelial–mesenchymal transition. Other recent works have identified a novel role for the FHIT gene product, FHIT, as a genome “caretaker.” Loss of this caretaker function leads to nucleotide imbalance, spontaneous replication stress, and DNA breaks. Because FHIT loss-induced DNA damage is “checkpoint blind,” cells accumulate further DNA damage during subsequent cell cycles, accruing global genome instability that could facilitate oncogenic mutation acquisition and expedite clonal expansion. Loss of FHIT activity therefore induces a mutator phenotype. Evidence for FHIT as a mutator gene is discussed in light of these recent investigations of FHIT loss and subsequent genome instability.
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Characterization of the role of FHIT in suppression of DNA damage.
Advances in biological regulation, 2012Co-Authors: Joshua C. Saldivar, Seyed Ali Hosseini, Jessica Bene, Satoshi Miuma, Susan Horton, Nyla A. Heerema, Kay HuebnerAbstract:The fragile histidine triad protein, FHIT, has a number of reported tumor suppressive functions which include signaling of apoptosis in cancer cells in vitro and in vivo, modulation of the DNA damage response, down-regulation of target oncogene expression, suppression of tumor growth in vivo, and suppression of cancer cell invasion and metastasis. Most of these functions of FHIT have been observed on exogenous re-expression of FHIT in FHIT-negative cancer cells. However, little is known about the tumorigenic changes that occur in normal or precancerous cells following loss of FHIT expression. Recently, we have shown that shortly after loss of FHIT expression, cells exhibit signs of DNA replication stress-induced DNA damage and develop genomic instability. Here, we extend these findings through investigation of different factors that affect FHIT function to prevent DNA damage. We found that FHIT activity is dependent upon a functional HIT domain and the tyrosine-114 residue, previously shown to be required for tumor suppression by FHIT. Furthermore, FHIT function was shown to be independent of exogenous and endogenous sources of oxidative stress. Finally, FHIT function was shown to be dependent upon Chk1 kinase activity, but independent of Atr or Atm kinases. Evidence suggests that FHIT and Chk1 kinase cooperate to prevent replication stress-induced DNA damage. These findings provide important and unexpected insights into the mechanism whereby loss of FHIT expression contributes to cell transformation.
Carlo M. Croce - One of the best experts on this subject based on the ideXlab platform.
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prevention of urinary bladder cancer in the FHIT knock out mouse with rofecoxib a cox 2 inhibitor
Urologic Oncology-seminars and Original Investigations, 2010Co-Authors: Nicola Zanesi, Domenico Darca, James Lenoir, Bernadette Wildemore, Fedra Gottardo, Emma Bragantini, Dolores Shuppbyrne, Matteo Fassan, Carlo M. CroceAbstract:Objectives: Aberrant or increased expression of cyclooxygenase-2 (COX-2) has been implicated in the pathogenesis of many diseases, including cancer. However, the exact mechanism by which COX-2 may influence tumorigenesis has yet to be described. To investigate the chemopreventive role of a COX-2 inhibitor, rofecoxib, in the development of urinary bladder cancer, we studied the effect of this drug in heterozygous and nullizygous fragile histidine triad (FHIT) gene-deficient mice in a chemically induced carcinogenesis model. Materials and methods: Two-hundred eight mice consisting of 50 FHIT / ,6 3FHIT /– and 95 FHIT –/–, were divided into five treatment groups and followed up for 15 weeks. Mice were treated with freshly prepared solution of 0.1% or 0.01% N-butyl-N-(-4hydroxybutyl)-nitrosamine (BBN) in their drinking water and rofecoxib was administered in mouse chow at 150 parts per million concentration. Mice were sacrificed, and accurate histological analysis of the bladder was performed. Results: Rofecoxib treatment significantly reduced the incidence of preneoplastic lesions/bladder tumors (P 0.016). Comparing the incidence of neoplastic lesions in mice treated with rofecoxib and BBN (22/56, 39.3%) and mice treated only with BBN (32/57, 56.1%), a protective role of rofecoxib on the BBN tumor induction has been observed (P 0.024). A similar result (P 0.002) has been reached observing the incidence of mild and moderate dysplasia in mice treated with a lower concentration of BBN (8/16, 50.0% vs. 20/24, 83.3%).Moreover, as previously observed, a significant increase in neoplastic lesions in the FHIT /– and FHIT –/– vs. FHIT / mice after BBN treatment has been observed (P 0.003). Conclusions: These findings suggest that rofecoxib provides a therapeutic defense against bladder carcinogenesis in our model and confirmed that the FHIT knock-out mouse is a suitable system to study in vivo bladder carcinogenesis. © 2010 Elsevier Inc. All rights reserved.
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fragile gene product FHIT in oxidative and replicative stress responses
Cancer Science, 2009Co-Authors: Hiroshi Okumura, Carlo M. Croce, Masaki Mori, Flavia Pichiorri, Hideshi Ishii, Kay HuebnerAbstract:Though the fragile histidine triad gene product, FHIT, was discovered and characterized as a tumor suppressor 13 years ago, its sequence, structure, and cellular location did not provide clues to aid discovery of its mechanisms of suppression. Recently, using chemical cross-linkers and immunoprecipitation, a FHIT protein complex was identified that includes Hsp60 and Hsp10 which may mediate FHIT stability and mitochondrial localization, where FHIT binds and stabilizes ferredoxin reductase (Fdxr); when Fdxr is overexpressed, it can lead to production of reactive oxygen species (ROS) that induce apoptosis. Cancer cells expressing endogenous or exogenous FHIT, when exposed to H2O2, an oxidative stress, produce higher levels of apoptosis-inducing ROS than matched, FHIT-negative cells; the FHIT-negative cancer cells survive, carrying DNA damage. In addition to this mitochondrial function, FHIT-overexpression in cancer cells exposed to replicative stress-inducing agents leads to enhanced caspase 3 activation and apoptosis, due to defective Chk1 activation. Thus, damage to the fragile FHIT locus leads to reduced expression of FHIT protein, and makes a two-pronged contribution to development of preneoplastic clonal expansion: (1) absence or reduction of FHIT leads to reduced expression of Fdxr and reduced ROS-induced apoptosis; (2) cells that escape ROS- or replicative stress-induced apoptosis can carry misrepaired DNA damage. The aberrant DNA damage response checkpoint in FHIT-deficient preneoplasias and cancers may make these lesions targets for inhibitors of proteins such as Parp1 and Chk1 with important roles in checkpoint responses, as observed for BRCA1-deficient cancer cells that also exhibit DNA damage repair deficiencies. (Cancer Sci 2009; 100: 1145–1150)
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correlation of fragile histidine triad FHIT protein structural features with effector interactions and biological functions
Journal of Biological Chemistry, 2009Co-Authors: Flavia Pichiorri, Teresa Druck, Sung-suk Suh, Hiroshi Okumura, Larry D. Barnes, Tatsuya Nakamura, Preston N Garrison, Pierluigi Gasparini, Kelly A Mccorkell, Carlo M. CroceAbstract:We have previously shown that FHIT tumor suppressor protein interacts with Hsp60 chaperone machinery and ferredoxin reductase (Fdxr) protein. FHIT-effector interactions are associated with a FHIT-dependent increase in Fdxr stability, followed by generation of reactive oxygen species and apoptosis induction under conditions of oxidative stress. To define FHIT structural features that affect interactions, downstream signaling, and biological outcomes, we used cancer cells expressing FHIT mutants with amino acid substitutions that alter enzymatic activity, enzyme substrate binding, or phosphorylation at tyrosine 114. Gastric cancer cell clones stably expressing mutants that do not bind substrate or cannot be phosphorylated showed decreased binding to Hsp60 and Fdxr and reduced mitochondrial localization. Expression of FHIT or mutants that bind interactor proteins results in oxidative damage and accumulation of cells in G2/M or sub-G1 fractions after peroxide treatment; noninteracting mutants are defective in these biological effects. Gastric cancer clones expressing noncomplexing FHIT mutants show reduction of FHIT tumor suppressor activity, confirming that substrate binding, interaction with heat shock proteins, mitochondrial localization, and interaction with Fdxr are important for FHIT tumor suppressor function.
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FHIT tumor suppressor: guardian of the preneoplastic genome
Future oncology (London England), 2008Co-Authors: Flavia Pichiorri, Kay Huebner, Tiziana Palumbo, Sung-suk Suh, Hiroshi Okamura, Francesco Trapasso, Hideshi Ishii, Carlo M. CroceAbstract:Environmental agents induce intragenic alterations in the FRA3B/FHIT chromosome fragile site, resulting in fragile FHIT allele loss early in cancer development. FHIT knockout mice are predisposed to tumor development and FHIT gene therapy reduces tumor burden. Repair-deficient cancers are likely to be FHIT-deficient and FHIT-deficient cells show enhanced resistance to ultraviolet C, mitomycin C, camptothecin and oxidative stress-induced cell killing. Loss of FHIT leads to alterations in the DNA damage response checkpoint and contributes to DNA instability. Hsp60/Hsp10 are FHIT interactors, suggesting a direct role for FHIT in stress responses. FHIT also interacts with and stabilizes ferrodoxin reductase (Fdxr), a mitochondrial flavoprotein that transfers electrons from NADPH to cytochrome P450, suggesting a role for FHIT in the modulation of reactive oxygen species production and of genomic damage.
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FHIT-proteasome degradation caused by mitogenic stimulation of the EGF receptor family in cancer cells.
Proceedings of the National Academy of Sciences of the United States of America, 2006Co-Authors: Francesca Bianchi, Carlo M. Croce, Yuri Pekarsky, Alessandra Magnifico, Clelia Olgiati, Nicola Zanesi, Elda Tagliabue, Sylvie Ménard, Manuela CampiglioAbstract:The tumor suppressor gene FHIT is inactivated by genetic and epigenetic changes in the majority of common human cancers. The human FHIT protein undergoes phosphorylation on tyrosine residue 114 by Src and related kinases both in vitro and in vivo. Src is a key cytoplasmic tyrosine kinase downstream to several growth factor receptors, including those of the EGF receptor family, which are overexpressed and activated in about one-third of human breast and ovarian carcinomas. However, the biological significance of FHIT phosphorylation by Src has remained elusive. In the present study, we demonstrate that FHIT acts as a checkpoint in cell proliferation mediated by activated tyrosine kinase receptors that recruit Src. Activation of EGF receptor family members induced FHIT phosphorylation by Src and the subsequent proteasome degradation of the phosphorylated FHIT protein. Indeed, the use of the FHIT mutant Y114F, which carries a phenylalanine instead of a tyrosine at position 114, unable to be phosphorylated on tyrosine 114 by Src, prevents FHIT degradation. Moreover, FHIT protein reduction is transient and occurs in a specific temporal window. During the signaling pathway of activated tyrosine kinase receptors, the phosphorylation of FHIT induces its degradation and the subsequent reduction in FHIT protein levels allows the transmission of the mitogenic signal; immediately thereafter, FHIT protein levels are restored. Such a scenario would suggest a key role for FHIT in the balance of proliferation/survival/apoptosis signals.
Hiroshi Okumura - One of the best experts on this subject based on the ideXlab platform.
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FHIT–Fdxr interaction in the mitochondria: modulation of reactive oxygen species generation and apoptosis in cancer cells
Cell Death & Disease, 2019Co-Authors: Teresa Druck, Flavia Pichiorri, Tiziana Palumbo, Francesco Trapasso, Marco Gaspari, Rami I Aqeilan, Eugenio Gaudio, Dongju Park, Douglas G. Cheung, Hiroshi OkumuraAbstract:FHIT protein is lost in cancers of most, perhaps all, cancer types; when restored, it can induce apoptosis and suppress tumorigenicity, as shown in vitro and in mouse tumor models in vivo. Following protein cross-linking and proteomics analyses, we characterized a FHIT protein complex involved in triggering FHIT-mediated apoptosis. The complex includes the heat-shock chaperonin pair, HSP60/10, which is likely involved in importing FHIT into the mitochondria, where it interacts with ferredoxin reductase, responsible for transferring electrons from NADPH to cytochrome P450 via ferredoxin, in electron transport chain complex III. Overexpression of FHIT protein in FHIT-deficient cancer cells modulates the production of intracellular reactive oxygen species, causing increased ROS, following peroxide treatment, with subsequent increased apoptosis of lung cancer cells under oxidative stress conditions; conversely, FHIT-negative cells escape ROS overproduction and ROS-induced apoptosis, likely carrying oxidative damage. Thus, characterization of FHIT-interacting proteins has identified direct effectors of a FHIT-mediated apoptotic signal pathway that is lost in many cancers. This is of translational interest considering the very recent emphasis in a number of high-profile publications, concerning the role of oxidative phosphorylation in the treatment of human cancers, and especially cancer stem cells that rely upon oxidative phosphorylation for survival. Additionally, we have shown that cells from a FHIT-deficient lung cancer cell line, are sensitive to killing by exposure to atovaquone, thought to act as a selective oxidative phosphorylation inhibitor by targeting the CoQ10 dependence of the mitochondrial complex III, while the FHIT-expressing sister clone is resistant to this treatment.
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Nit1 and FHIT Tumor Suppressor Activities Are Additive
Journal of cellular biochemistry, 2009Co-Authors: Jin Sun, Teresa Druck, Louise Y.y. Fong, Hiroshi Okumura, Martha Yearsley, Wendy L. Frankel, Kay HuebnerAbstract:The fragile histidine triad gene (human FHIT, mouse FHIT) has been shown to act as a tumor suppressor gene. Nit1 and FHIT form a fusion protein, encoded by the NitFHIT gene in flies and worms, suggesting that mammalian Nit1 and FHIT proteins, which are encoded by genes on different chromosomes in mammals, may function in the same signal pathway(s). A previous study showed that Nit1 deficiency in knockout mice confers a cancer prone phenotype, as does FHIT deficiency. We have now assessed the tumor susceptibility of FHIT−/−Nit1−/− mice and observed that double knockout mice develop more spontaneous and carcinogen-induced tumors than FHIT−/− mice, suggesting that the extent of tumor susceptibility due to Nit1 and FHIT deficiency is additive, and that Nit1 and FHIT affect distinct signal pathways in mammals. Nit1, like FHIT, is present in cytoplasm and mitochondria but not nuclei. Because FHIT deficiency affects responses to replicative and oxidative stress, we sought evidence for Nit1 function in response to such stresses in tissues and cultured cells: when treated with hydroxyurea, the normal kidney-derived double-deficient cells appear not to activate the pChk2 pathway and when treated with H2O2, show little evidence of DNA damage, compared with wild type and FHIT−/− cells. The relevance of Nit1 deficiency to human cancers was examined in human esophageal cancer tissues, and loss of Nit1 expression was observed in 48% of esophageal adenocarcinomas. J. Cell. Biochem. 107: 1097–1106, 2009. © 2009 Wiley-Liss, Inc.
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fragile gene product FHIT in oxidative and replicative stress responses
Cancer Science, 2009Co-Authors: Hiroshi Okumura, Carlo M. Croce, Masaki Mori, Flavia Pichiorri, Hideshi Ishii, Kay HuebnerAbstract:Though the fragile histidine triad gene product, FHIT, was discovered and characterized as a tumor suppressor 13 years ago, its sequence, structure, and cellular location did not provide clues to aid discovery of its mechanisms of suppression. Recently, using chemical cross-linkers and immunoprecipitation, a FHIT protein complex was identified that includes Hsp60 and Hsp10 which may mediate FHIT stability and mitochondrial localization, where FHIT binds and stabilizes ferredoxin reductase (Fdxr); when Fdxr is overexpressed, it can lead to production of reactive oxygen species (ROS) that induce apoptosis. Cancer cells expressing endogenous or exogenous FHIT, when exposed to H2O2, an oxidative stress, produce higher levels of apoptosis-inducing ROS than matched, FHIT-negative cells; the FHIT-negative cancer cells survive, carrying DNA damage. In addition to this mitochondrial function, FHIT-overexpression in cancer cells exposed to replicative stress-inducing agents leads to enhanced caspase 3 activation and apoptosis, due to defective Chk1 activation. Thus, damage to the fragile FHIT locus leads to reduced expression of FHIT protein, and makes a two-pronged contribution to development of preneoplastic clonal expansion: (1) absence or reduction of FHIT leads to reduced expression of Fdxr and reduced ROS-induced apoptosis; (2) cells that escape ROS- or replicative stress-induced apoptosis can carry misrepaired DNA damage. The aberrant DNA damage response checkpoint in FHIT-deficient preneoplasias and cancers may make these lesions targets for inhibitors of proteins such as Parp1 and Chk1 with important roles in checkpoint responses, as observed for BRCA1-deficient cancer cells that also exhibit DNA damage repair deficiencies. (Cancer Sci 2009; 100: 1145–1150)
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correlation of fragile histidine triad FHIT protein structural features with effector interactions and biological functions
Journal of Biological Chemistry, 2009Co-Authors: Flavia Pichiorri, Teresa Druck, Sung-suk Suh, Hiroshi Okumura, Larry D. Barnes, Tatsuya Nakamura, Preston N Garrison, Pierluigi Gasparini, Kelly A Mccorkell, Carlo M. CroceAbstract:We have previously shown that FHIT tumor suppressor protein interacts with Hsp60 chaperone machinery and ferredoxin reductase (Fdxr) protein. FHIT-effector interactions are associated with a FHIT-dependent increase in Fdxr stability, followed by generation of reactive oxygen species and apoptosis induction under conditions of oxidative stress. To define FHIT structural features that affect interactions, downstream signaling, and biological outcomes, we used cancer cells expressing FHIT mutants with amino acid substitutions that alter enzymatic activity, enzyme substrate binding, or phosphorylation at tyrosine 114. Gastric cancer cell clones stably expressing mutants that do not bind substrate or cannot be phosphorylated showed decreased binding to Hsp60 and Fdxr and reduced mitochondrial localization. Expression of FHIT or mutants that bind interactor proteins results in oxidative damage and accumulation of cells in G2/M or sub-G1 fractions after peroxide treatment; noninteracting mutants are defective in these biological effects. Gastric cancer clones expressing noncomplexing FHIT mutants show reduction of FHIT tumor suppressor activity, confirming that substrate binding, interaction with heat shock proteins, mitochondrial localization, and interaction with Fdxr are important for FHIT tumor suppressor function.
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FHIT interaction with ferredoxin reductase triggers generation of reactive oxygen species and apoptosis of cancer cells
Journal of Biological Chemistry, 2008Co-Authors: Francesco Trapasso, Flavia Pichiorri, Tiziana Palumbo, Marco Gaspari, Rami I Aqeilan, Eugenio Gaudio, Hiroshi Okumura, Rodolfo Iuliano, Giampiero Di Leva, Muller FabbriAbstract:FHIT protein is lost in most cancers, its restoration suppresses tumorigenicity, and virus-mediated FHIT gene therapy induces apoptosis and suppresses tumors in preclinical models. We have used protein cross-linking and proteomics methods to characterize a FHIT protein complex involved in triggering FHIT-mediated apoptosis. The complex includes Hsp60 and Hsp10 that mediate FHIT stability and may affect import into mitochondria, where it interacts with ferredoxin reductase, responsible for transferring electrons from NADPH to cytochrome P450 via ferredoxin. Viral-mediated FHIT restoration increases production of intracellular reactive oxygen species, followed by increased apoptosis of lung cancer cells under oxidative stress conditions; conversely, FHIT-negative cells escape apoptosis, carrying serious oxidative DNA damage that may contribute to an increased mutation rate. Characterization of FHIT interacting proteins has identified direct effectors of the FHIT-mediated apoptotic pathway that is lost in most cancers through loss of FHIT.
Flavia Pichiorri - One of the best experts on this subject based on the ideXlab platform.
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FHIT fdxr interaction in the mitochondria modulation of reactive oxygen species generation and apoptosis in cancer cells
Cell Death and Disease, 2019Co-Authors: Teresa Druck, Flavia Pichiorri, Tiziana Palumbo, Francesco Trapasso, Marco Gaspari, Rami I Aqeilan, Dongju Park, Douglas G. Cheung, Eugenio GaudioAbstract:FHIT protein is lost in cancers of most, perhaps all, cancer types; when restored, it can induce apoptosis and suppress tumorigenicity, as shown in vitro and in mouse tumor models in vivo. Following protein cross-linking and proteomics analyses, we characterized a FHIT protein complex involved in triggering FHIT-mediated apoptosis. The complex includes the heat-shock chaperonin pair, HSP60/10, which is likely involved in importing FHIT into the mitochondria, where it interacts with ferredoxin reductase, responsible for transferring electrons from NADPH to cytochrome P450 via ferredoxin, in electron transport chain complex III. Overexpression of FHIT protein in FHIT-deficient cancer cells modulates the production of intracellular reactive oxygen species, causing increased ROS, following peroxide treatment, with subsequent increased apoptosis of lung cancer cells under oxidative stress conditions; conversely, FHIT-negative cells escape ROS overproduction and ROS-induced apoptosis, likely carrying oxidative damage. Thus, characterization of FHIT-interacting proteins has identified direct effectors of a FHIT-mediated apoptotic signal pathway that is lost in many cancers. This is of translational interest considering the very recent emphasis in a number of high-profile publications, concerning the role of oxidative phosphorylation in the treatment of human cancers, and especially cancer stem cells that rely upon oxidative phosphorylation for survival. Additionally, we have shown that cells from a FHIT-deficient lung cancer cell line, are sensitive to killing by exposure to atovaquone, thought to act as a selective oxidative phosphorylation inhibitor by targeting the CoQ10 dependence of the mitochondrial complex III, while the FHIT-expressing sister clone is resistant to this treatment.
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FHIT–Fdxr interaction in the mitochondria: modulation of reactive oxygen species generation and apoptosis in cancer cells
Cell Death & Disease, 2019Co-Authors: Teresa Druck, Flavia Pichiorri, Tiziana Palumbo, Francesco Trapasso, Marco Gaspari, Rami I Aqeilan, Eugenio Gaudio, Dongju Park, Douglas G. Cheung, Hiroshi OkumuraAbstract:FHIT protein is lost in cancers of most, perhaps all, cancer types; when restored, it can induce apoptosis and suppress tumorigenicity, as shown in vitro and in mouse tumor models in vivo. Following protein cross-linking and proteomics analyses, we characterized a FHIT protein complex involved in triggering FHIT-mediated apoptosis. The complex includes the heat-shock chaperonin pair, HSP60/10, which is likely involved in importing FHIT into the mitochondria, where it interacts with ferredoxin reductase, responsible for transferring electrons from NADPH to cytochrome P450 via ferredoxin, in electron transport chain complex III. Overexpression of FHIT protein in FHIT-deficient cancer cells modulates the production of intracellular reactive oxygen species, causing increased ROS, following peroxide treatment, with subsequent increased apoptosis of lung cancer cells under oxidative stress conditions; conversely, FHIT-negative cells escape ROS overproduction and ROS-induced apoptosis, likely carrying oxidative damage. Thus, characterization of FHIT-interacting proteins has identified direct effectors of a FHIT-mediated apoptotic signal pathway that is lost in many cancers. This is of translational interest considering the very recent emphasis in a number of high-profile publications, concerning the role of oxidative phosphorylation in the treatment of human cancers, and especially cancer stem cells that rely upon oxidative phosphorylation for survival. Additionally, we have shown that cells from a FHIT-deficient lung cancer cell line, are sensitive to killing by exposure to atovaquone, thought to act as a selective oxidative phosphorylation inhibitor by targeting the CoQ10 dependence of the mitochondrial complex III, while the FHIT-expressing sister clone is resistant to this treatment.
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fragile gene product FHIT in oxidative and replicative stress responses
Cancer Science, 2009Co-Authors: Hiroshi Okumura, Carlo M. Croce, Masaki Mori, Flavia Pichiorri, Hideshi Ishii, Kay HuebnerAbstract:Though the fragile histidine triad gene product, FHIT, was discovered and characterized as a tumor suppressor 13 years ago, its sequence, structure, and cellular location did not provide clues to aid discovery of its mechanisms of suppression. Recently, using chemical cross-linkers and immunoprecipitation, a FHIT protein complex was identified that includes Hsp60 and Hsp10 which may mediate FHIT stability and mitochondrial localization, where FHIT binds and stabilizes ferredoxin reductase (Fdxr); when Fdxr is overexpressed, it can lead to production of reactive oxygen species (ROS) that induce apoptosis. Cancer cells expressing endogenous or exogenous FHIT, when exposed to H2O2, an oxidative stress, produce higher levels of apoptosis-inducing ROS than matched, FHIT-negative cells; the FHIT-negative cancer cells survive, carrying DNA damage. In addition to this mitochondrial function, FHIT-overexpression in cancer cells exposed to replicative stress-inducing agents leads to enhanced caspase 3 activation and apoptosis, due to defective Chk1 activation. Thus, damage to the fragile FHIT locus leads to reduced expression of FHIT protein, and makes a two-pronged contribution to development of preneoplastic clonal expansion: (1) absence or reduction of FHIT leads to reduced expression of Fdxr and reduced ROS-induced apoptosis; (2) cells that escape ROS- or replicative stress-induced apoptosis can carry misrepaired DNA damage. The aberrant DNA damage response checkpoint in FHIT-deficient preneoplasias and cancers may make these lesions targets for inhibitors of proteins such as Parp1 and Chk1 with important roles in checkpoint responses, as observed for BRCA1-deficient cancer cells that also exhibit DNA damage repair deficiencies. (Cancer Sci 2009; 100: 1145–1150)
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correlation of fragile histidine triad FHIT protein structural features with effector interactions and biological functions
Journal of Biological Chemistry, 2009Co-Authors: Flavia Pichiorri, Teresa Druck, Sung-suk Suh, Hiroshi Okumura, Larry D. Barnes, Tatsuya Nakamura, Preston N Garrison, Pierluigi Gasparini, Kelly A Mccorkell, Carlo M. CroceAbstract:We have previously shown that FHIT tumor suppressor protein interacts with Hsp60 chaperone machinery and ferredoxin reductase (Fdxr) protein. FHIT-effector interactions are associated with a FHIT-dependent increase in Fdxr stability, followed by generation of reactive oxygen species and apoptosis induction under conditions of oxidative stress. To define FHIT structural features that affect interactions, downstream signaling, and biological outcomes, we used cancer cells expressing FHIT mutants with amino acid substitutions that alter enzymatic activity, enzyme substrate binding, or phosphorylation at tyrosine 114. Gastric cancer cell clones stably expressing mutants that do not bind substrate or cannot be phosphorylated showed decreased binding to Hsp60 and Fdxr and reduced mitochondrial localization. Expression of FHIT or mutants that bind interactor proteins results in oxidative damage and accumulation of cells in G2/M or sub-G1 fractions after peroxide treatment; noninteracting mutants are defective in these biological effects. Gastric cancer clones expressing noncomplexing FHIT mutants show reduction of FHIT tumor suppressor activity, confirming that substrate binding, interaction with heat shock proteins, mitochondrial localization, and interaction with Fdxr are important for FHIT tumor suppressor function.
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FHIT interaction with ferredoxin reductase triggers generation of reactive oxygen species and apoptosis of cancer cells
Journal of Biological Chemistry, 2008Co-Authors: Francesco Trapasso, Flavia Pichiorri, Tiziana Palumbo, Marco Gaspari, Rami I Aqeilan, Eugenio Gaudio, Hiroshi Okumura, Rodolfo Iuliano, Giampiero Di Leva, Muller FabbriAbstract:FHIT protein is lost in most cancers, its restoration suppresses tumorigenicity, and virus-mediated FHIT gene therapy induces apoptosis and suppresses tumors in preclinical models. We have used protein cross-linking and proteomics methods to characterize a FHIT protein complex involved in triggering FHIT-mediated apoptosis. The complex includes Hsp60 and Hsp10 that mediate FHIT stability and may affect import into mitochondria, where it interacts with ferredoxin reductase, responsible for transferring electrons from NADPH to cytochrome P450 via ferredoxin. Viral-mediated FHIT restoration increases production of intracellular reactive oxygen species, followed by increased apoptosis of lung cancer cells under oxidative stress conditions; conversely, FHIT-negative cells escape apoptosis, carrying serious oxidative DNA damage that may contribute to an increased mutation rate. Characterization of FHIT interacting proteins has identified direct effectors of the FHIT-mediated apoptotic pathway that is lost in most cancers through loss of FHIT.
Francesco Trapasso - One of the best experts on this subject based on the ideXlab platform.
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FHIT fdxr interaction in the mitochondria modulation of reactive oxygen species generation and apoptosis in cancer cells
Cell Death and Disease, 2019Co-Authors: Teresa Druck, Flavia Pichiorri, Tiziana Palumbo, Francesco Trapasso, Marco Gaspari, Rami I Aqeilan, Dongju Park, Douglas G. Cheung, Eugenio GaudioAbstract:FHIT protein is lost in cancers of most, perhaps all, cancer types; when restored, it can induce apoptosis and suppress tumorigenicity, as shown in vitro and in mouse tumor models in vivo. Following protein cross-linking and proteomics analyses, we characterized a FHIT protein complex involved in triggering FHIT-mediated apoptosis. The complex includes the heat-shock chaperonin pair, HSP60/10, which is likely involved in importing FHIT into the mitochondria, where it interacts with ferredoxin reductase, responsible for transferring electrons from NADPH to cytochrome P450 via ferredoxin, in electron transport chain complex III. Overexpression of FHIT protein in FHIT-deficient cancer cells modulates the production of intracellular reactive oxygen species, causing increased ROS, following peroxide treatment, with subsequent increased apoptosis of lung cancer cells under oxidative stress conditions; conversely, FHIT-negative cells escape ROS overproduction and ROS-induced apoptosis, likely carrying oxidative damage. Thus, characterization of FHIT-interacting proteins has identified direct effectors of a FHIT-mediated apoptotic signal pathway that is lost in many cancers. This is of translational interest considering the very recent emphasis in a number of high-profile publications, concerning the role of oxidative phosphorylation in the treatment of human cancers, and especially cancer stem cells that rely upon oxidative phosphorylation for survival. Additionally, we have shown that cells from a FHIT-deficient lung cancer cell line, are sensitive to killing by exposure to atovaquone, thought to act as a selective oxidative phosphorylation inhibitor by targeting the CoQ10 dependence of the mitochondrial complex III, while the FHIT-expressing sister clone is resistant to this treatment.
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FHIT–Fdxr interaction in the mitochondria: modulation of reactive oxygen species generation and apoptosis in cancer cells
Cell Death & Disease, 2019Co-Authors: Teresa Druck, Flavia Pichiorri, Tiziana Palumbo, Francesco Trapasso, Marco Gaspari, Rami I Aqeilan, Eugenio Gaudio, Dongju Park, Douglas G. Cheung, Hiroshi OkumuraAbstract:FHIT protein is lost in cancers of most, perhaps all, cancer types; when restored, it can induce apoptosis and suppress tumorigenicity, as shown in vitro and in mouse tumor models in vivo. Following protein cross-linking and proteomics analyses, we characterized a FHIT protein complex involved in triggering FHIT-mediated apoptosis. The complex includes the heat-shock chaperonin pair, HSP60/10, which is likely involved in importing FHIT into the mitochondria, where it interacts with ferredoxin reductase, responsible for transferring electrons from NADPH to cytochrome P450 via ferredoxin, in electron transport chain complex III. Overexpression of FHIT protein in FHIT-deficient cancer cells modulates the production of intracellular reactive oxygen species, causing increased ROS, following peroxide treatment, with subsequent increased apoptosis of lung cancer cells under oxidative stress conditions; conversely, FHIT-negative cells escape ROS overproduction and ROS-induced apoptosis, likely carrying oxidative damage. Thus, characterization of FHIT-interacting proteins has identified direct effectors of a FHIT-mediated apoptotic signal pathway that is lost in many cancers. This is of translational interest considering the very recent emphasis in a number of high-profile publications, concerning the role of oxidative phosphorylation in the treatment of human cancers, and especially cancer stem cells that rely upon oxidative phosphorylation for survival. Additionally, we have shown that cells from a FHIT-deficient lung cancer cell line, are sensitive to killing by exposure to atovaquone, thought to act as a selective oxidative phosphorylation inhibitor by targeting the CoQ10 dependence of the mitochondrial complex III, while the FHIT-expressing sister clone is resistant to this treatment.
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FHIT interaction with ferredoxin reductase triggers generation of reactive oxygen species and apoptosis of cancer cells
Journal of Biological Chemistry, 2008Co-Authors: Francesco Trapasso, Flavia Pichiorri, Tiziana Palumbo, Marco Gaspari, Rami I Aqeilan, Eugenio Gaudio, Hiroshi Okumura, Rodolfo Iuliano, Giampiero Di Leva, Muller FabbriAbstract:FHIT protein is lost in most cancers, its restoration suppresses tumorigenicity, and virus-mediated FHIT gene therapy induces apoptosis and suppresses tumors in preclinical models. We have used protein cross-linking and proteomics methods to characterize a FHIT protein complex involved in triggering FHIT-mediated apoptosis. The complex includes Hsp60 and Hsp10 that mediate FHIT stability and may affect import into mitochondria, where it interacts with ferredoxin reductase, responsible for transferring electrons from NADPH to cytochrome P450 via ferredoxin. Viral-mediated FHIT restoration increases production of intracellular reactive oxygen species, followed by increased apoptosis of lung cancer cells under oxidative stress conditions; conversely, FHIT-negative cells escape apoptosis, carrying serious oxidative DNA damage that may contribute to an increased mutation rate. Characterization of FHIT interacting proteins has identified direct effectors of the FHIT-mediated apoptotic pathway that is lost in most cancers through loss of FHIT.
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FHIT tumor suppressor: guardian of the preneoplastic genome
Future oncology (London England), 2008Co-Authors: Flavia Pichiorri, Kay Huebner, Tiziana Palumbo, Sung-suk Suh, Hiroshi Okamura, Francesco Trapasso, Hideshi Ishii, Carlo M. CroceAbstract:Environmental agents induce intragenic alterations in the FRA3B/FHIT chromosome fragile site, resulting in fragile FHIT allele loss early in cancer development. FHIT knockout mice are predisposed to tumor development and FHIT gene therapy reduces tumor burden. Repair-deficient cancers are likely to be FHIT-deficient and FHIT-deficient cells show enhanced resistance to ultraviolet C, mitomycin C, camptothecin and oxidative stress-induced cell killing. Loss of FHIT leads to alterations in the DNA damage response checkpoint and contributes to DNA instability. Hsp60/Hsp10 are FHIT interactors, suggesting a direct role for FHIT in stress responses. FHIT also interacts with and stabilizes ferrodoxin reductase (Fdxr), a mitochondrial flavoprotein that transfers electrons from NADPH to cytochrome P450, suggesting a role for FHIT in the modulation of reactive oxygen species production and of genomic damage.
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characterization of a FHIT protein complex suggests a novel pathway to apoptosis of cancer cells
Cancer Research, 2007Co-Authors: Flavia Pichiorri, Tiziana Palumbo, Francesco Trapasso, Marco Gaspari, Eugenio Gaudio, Hiroshi Okumura, Rodolfo Iuliano, Muller Fabbri, Di Leva Giampiero, Salvatore VenutaAbstract:451 The fragile histidine triad (FHIT) gene, spanning FRA3B at 3p14.2, the most active human common fragile site, shows abnormal transcripts, promoter hypermethylation and/or allelic loss in a large fraction of human cancers of various organs. Furthermore, FHIT restoration in a number of malignant cell lines triggers apoptosis in vitro and in preclinical models. However, though more than 600 publications document involvement of FHIT alterations in cancers, the mechanisms through which FHIT protein signals tumor suppression have not been defined, partially because interacting proteins have been difficult to identify. We have now used protein-protein cross-linking followed by identification of cross-llinked proteins by proteomics methods to identify a FHIT protein complex responsible for triggering apoptosis in FHIT virally-transduced lung cancer cells. Our experimental results demonstrate that FHIT localizes in both cytosol (where it interacts with Hsp60 and Hsp10) and mitochondria where it interacts with main mitochondria chain respiratory proteins. Interestingly, FHIT restoration significantly increases the production of reactive oxygen species (ROS) in lung and colon cancer-derived cells in response to stressful treatments. Delineation of direct downstream effectors of the FHIT suppressor pathway will lead to intensification of mechanistic studies of FHIT function that may influence future preventive and therapeutic strategies to activate the FHIT pathway. The finding that ROS generation is crucial for FHIT-mediated apoptosis emphasizes the importance of FHIT loss as a negative prognostic factor in various clinical settings; for example, assessment of FHIT status in preneoplastic or neoplastic conditions may be predictive of responses to antioxidant treatments.