Cancer Preventive Isothiocyanates Induce Selective Degradation of Cellular α- and β-Tubulins by Proteasomes

Abstract

Although it is conceivable that cancer preventive isothiocyanates (ITCs), a family of compounds in cruciferous vegetables, induce cell cycle arrest and apoptosis through a mechanism involving oxidative stress, our study shows that binding to cellular proteins correlates with their potencies of apoptosis induction. More recently, we showed that ITCs bind selectively to tubulins. The differential binding affinities toward tubulin among benzyl isothiocyanate, phenethyl isothiocyanate, and sulforaphane correlate well with their potencies of inducing tubulin conformation changes, microtubule depolymerization, and eventual cell cycle arrest and apoptosis in human lung cancer A549 cells. These results support that tubulin binding by ITCs is an early event for cell growth inhibition. Here we demonstrate that ITCs can selectively induce degradation of both α- and β-tubulins in a variety of human cancer cell lines in a dose- and time-dependent manner. The onset of degradation, a rapid and irreversible process, is initiated by tubulin aggregation, and the degradation is proteasome-dependent. Results indicate that the degradation is triggered by ITC binding to tubulin and is irrelevant to oxidative stress. This is the first report that tubulin, a stable and abundant cytoskeleton protein required for cell cycle progression, can be selectively degraded by a small molecule. Although it is conceivable that cancer preventive isothiocyanates (ITCs), a family of compounds in cruciferous vegetables, induce cell cycle arrest and apoptosis through a mechanism involving oxidative stress, our study shows that binding to cellular proteins correlates with their potencies of apoptosis induction. More recently, we showed that ITCs bind selectively to tubulins. The differential binding affinities toward tubulin among benzyl isothiocyanate, phenethyl isothiocyanate, and sulforaphane correlate well with their potencies of inducing tubulin conformation changes, microtubule depolymerization, and eventual cell cycle arrest and apoptosis in human lung cancer A549 cells. These results support that tubulin binding by ITCs is an early event for cell growth inhibition. Here we demonstrate that ITCs can selectively induce degradation of both α- and β-tubulins in a variety of human cancer cell lines in a dose- and time-dependent manner. The onset of degradation, a rapid and irreversible process, is initiated by tubulin aggregation, and the degradation is proteasome-dependent. Results indicate that the degradation is triggered by ITC binding to tubulin and is irrelevant to oxidative stress. This is the first report that tubulin, a stable and abundant cytoskeleton protein required for cell cycle progression, can be selectively degraded by a small molecule. Microtubules as a major cytoskeleton component in all eukaryotic cells play essential roles such as maintenance of cell polarity, intracellular traffic, organization, and cell motility (1Hartwell L.H. Kastan M.B. Science. 1994; 266: 1821-1828Crossref PubMed Scopus (2303) Google Scholar, 2Lengauer C. Kinzler K.W. Vogelstein B. Nature. 1998; 96: 643-649Crossref Scopus (3366) Google Scholar, 3White E. Genes Dev. 1996; 10: 1-15Crossref PubMed Scopus (1322) Google Scholar, 4Jordan M.A. Wilson L. Nat. Rev. Cancer. 2004; 4: 253-265Crossref PubMed Scopus (3468) Google Scholar). During cell division, the microtubule-formed mitotic spindle ensures the replicated chromosomes separate evenly at the end of the mitotic phase to the two daughter cells (1Hartwell L.H. Kastan M.B. Science. 1994; 266: 1821-1828Crossref PubMed Scopus (2303) Google Scholar). It is because of its essential roles in cell growth that microtubules become a valid target for the development of anti-microtubule drugs against the rapidly growing cancer cells (2Lengauer C. Kinzler K.W. Vogelstein B. Nature. 1998; 96: 643-649Crossref Scopus (3366) Google Scholar), as interference of microtubule dynamics arrests cell cycle progression and induces apoptosis (3White E. Genes Dev. 1996; 10: 1-15Crossref PubMed Scopus (1322) Google Scholar). Therefore, microtubules have been considered one of the best targets to date for cancer chemotherapy (4Jordan M.A. Wilson L. Nat. Rev. Cancer. 2004; 4: 253-265Crossref PubMed Scopus (3468) Google Scholar).Isothiocyanates (ITCs) 3The abbreviations used are: ITCisothiocyanateBITCbenzyl isothiocyanatePEITCphenethyl isothiocyanateSFNsulforaphane; N-methylphenethylamineCHAPS3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulfonic acidATZaminotriazoleCHXcycloheximidePBSphosphate-buffered salinePEGpolyethylene glycolRTreverse transcription2-DG2-deoxyglucoseBisTris2-[bis(2-hydroxyethyl)amino]-2-(hydroxymethyl)propane-1,3-diolZbenzyloxycarbonylfmkfluoromethyl ketonePSIproteasome inhibitor IROSreactive oxygen speciesMS/MStandem mass spectrometryIPimmunoprecipitation. 3The abbreviations used are: ITCisothiocyanateBITCbenzyl isothiocyanatePEITCphenethyl isothiocyanateSFNsulforaphane; N-methylphenethylamineCHAPS3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulfonic acidATZaminotriazoleCHXcycloheximidePBSphosphate-buffered salinePEGpolyethylene glycolRTreverse transcription2-DG2-deoxyglucoseBisTris2-[bis(2-hydroxyethyl)amino]-2-(hydroxymethyl)propane-1,3-diolZbenzyloxycarbonylfmkfluoromethyl ketonePSIproteasome inhibitor IROSreactive oxygen speciesMS/MStandem mass spectrometryIPimmunoprecipitation. are among the best studied chemopreventive small molecules (5WHOIARC Handbook on Cancer Prevention. 9. France, Lyon2004: 43-171Google Scholar). The three most studied ITCs, including benzyl-ITC (BITC; abundant in garden cress), phenethyl-ITC (PEITC; in watercress), and sulforaphane (SFN; in broccoli sprouts), have been shown to induce apoptosis and cell cycle arrest (5WHOIARC Handbook on Cancer Prevention. 9. France, Lyon2004: 43-171Google Scholar, 6Chen Y.R. Wang W. Kong A.N. Tan T.H. J. Biol. Chem. 1998; 273: 1769-1775Abstract Full Text Full Text PDF PubMed Scopus (237) Google Scholar, 7Chiao J.W. Chung F.L. Kancherla R. Ahmed T. Mittelman A. Conaway C.C. Int. J. Oncol. 2002; 20: 631-636PubMed Google Scholar, 8Xiao D. Choi S. Lee Y.J. Singh S.V. Mol. Carcinog. 2005; 43: 130-140Crossref PubMed Scopus (31) Google Scholar). Although it is believed that oxidative stress plays a role in cell cycle arrest and apoptosis induced by ITCs (6Chen Y.R. Wang W. Kong A.N. Tan T.H. J. Biol. Chem. 1998; 273: 1769-1775Abstract Full Text Full Text PDF PubMed Scopus (237) Google Scholar, 7Chiao J.W. Chung F.L. Kancherla R. Ahmed T. Mittelman A. Conaway C.C. Int. J. Oncol. 2002; 20: 631-636PubMed Google Scholar, 8Xiao D. Choi S. Lee Y.J. Singh S.V. Mol. Carcinog. 2005; 43: 130-140Crossref PubMed Scopus (31) Google Scholar, 9Zhang Y. Talalay P. Cancer Res. 1998; 58: 4632-4639PubMed Google Scholar, 10Zhang Y. Li J. Tang L. Free Radic. Biol. Med. 2005; 38: 70-77Crossref PubMed Scopus (93) Google Scholar, 11Singh S.V. Srivastava S.K. Choi S. Lew K.L. Antosiewicz J. Xiao D. Zeng Y. Watkins S.C. Johnson C.S. Trump D.L. Lee Y.J. Xiao H. Herman-Antosiewicz A. J. Biol. Chem. 2005; 280: 19911-19924Abstract Full Text Full Text PDF PubMed Scopus (315) Google Scholar, 12Trachootham D. Zhou Y. Zhang H. Demizu Y. Chen Z. Pelicano H. Chiao P.J. Achanta G. Arlinghaus R.B. Liu J. Huang P. Cancer Cell. 2006; 10: 241-252Abstract Full Text Full Text PDF PubMed Scopus (897) Google Scholar), we found that binding to proteins is a predominant intracellular chemical reaction of ITCs, and their protein binding affinities correlate well with inhibition of cell proliferation and induction of apoptosis (13Mi L. Wang X. Govind S. Hood B.L. Veenstra T.D. Conrads T.P. Saha D.T. Goldman R. Chung F.L. Cancer Res. 2007; 67: 6409-6416Crossref PubMed Scopus (128) Google Scholar). Recently, we identified tubulin, the microtubule constituent, as an in vivo target of ITCs by two-dimensional gel electrophoresis and mass spectrometry (14Mi L. Xiao Z. Hood B.L. Dakshanamurthy S. Wang X. Govind S. Conrads T.P. Veenstra T.D. Chung F.L. J. Biol. Chem. 2008; 283: 22136-22146Abstract Full Text Full Text PDF PubMed Scopus (181) Google Scholar). The growth inhibition of human non-small lung cancer A549 cells by ITCs followed the order of BITC > PEITC > SFN. The same order of potency was seen in their binding affinities toward tubulin, induction of its conformational changes, and inhibition of its polymerization. The study provides the first evidence of an in vivo ITC-tubulin binding adduct, indicating that direct modification of cysteines in tubulin by ITCs, rather than oxidative stress, may trigger cell cycle arrest and apoptosis.Here we report an unexpected novel finding that tubulin is selectively degraded in a variety of human cancer cells treated with ITCs. We provide evidence that the degradation is initiated by its binding with ITCs and mediated by the ubiquitin-proteasome pathway. Tubulin has long been viewed as a stable and abundant protein, and its levels in cells are tightly regulated (15Nogales E. Annu. Rev. Biochem. 2000; 69: 277-302Crossref PubMed Scopus (518) Google Scholar). In the literature, the only studies on cellular tubulin level change are related to “the auto-regulation theory,” i.e. when microtubules collapse, the presence of a massive amount of tubulin monomers would selectively destabilize tubulin mRNA and subsequently decrease tubulin protein synthesis (16Ben-Ze'ev A. Farmer S.R. Penman S. Cell. 1979; 17: 319-325Abstract Full Text PDF PubMed Scopus (221) Google Scholar, 17Cleveland D.W. Lopata M.A. Sherline P. Kirschner M.W. Cell. 1981; 25: 537-546Abstract Full Text PDF PubMed Scopus (259) Google Scholar, 18Cleveland D.W. Pittenger M.F. Feramisco J.R. Nature. 1983; 305: 738-740Crossref PubMed Scopus (80) Google Scholar). To our knowledge, there is no report on tubulin degradation as a result of treatment with any agents. Our studies provide strong evidence that tubulin as a target of ITCs for cell growth to a mechanism for the anti-microtubule of ITCs through binding to tubulin and a to study protein by modification with small we have shown that binding to intracellular proteins by ITCs is a predominant event that correlates with their apoptosis (13Mi L. Wang X. Govind S. Hood B.L. Veenstra T.D. Conrads T.P. Saha D.T. Goldman R. Chung F.L. Cancer Res. 2007; 67: 6409-6416Crossref PubMed Scopus (128) Google Scholar). We have in human lung cancer A549 cells that tubulin is a protein target of ITCs by two-dimensional gel and BITC a with tubulin in vivo (14Mi L. Xiao Z. Hood B.L. Dakshanamurthy S. Wang X. Govind S. Conrads T.P. Veenstra T.D. Chung F.L. J. Biol. Chem. 2008; 283: 22136-22146Abstract Full Text Full Text PDF PubMed Scopus (181) Google Scholar). we found that BITC and PEITC bind tubulin with are in inducing tubulin conformational changes, microtubule and growth inhibition and In and no binding to tubulin, are both of of the The study shows that BITC and PEITC can selectively induce and degradation of both α- and β-tubulins in a variety of cancer cells. of tubulin to ITCs is a The potency of inducing tubulin degradation the order BITC PEITC > correlates with their on the in and in vivo we a of tubulin degradation ITCs rapidly α- and β-tubulins in through binding to The modification by ITCs tubulin conformational changes, and The tubulin are degraded by the The including and the BITC binding to tubulin, conformational changes, and tubulin and In and can tubulin degradation This that and degradation are two related tubulin degradation is of oxidative stress. It be that results both and A549 cells indicating that the mechanism may be cell The tubulin degradation by small molecules is novel and as a of is that when a cell it two of P. J. Y. J. C. Nat. Biol. PubMed Scopus Google Scholar). it a variety of proteins to the protein to its the protein is the cell its degradation Nature. PubMed Scopus Google Scholar). event of proteins is that to Full Text Full Text PDF PubMed Scopus Google Scholar). The that the of proteins the of degradation of the cell the of the Y. M.A. R. D.L. D. C.C. T.P. Nature. 2007; PubMed Scopus Google Scholar, A. J. Biol. Chem. 2005; 280: Full Text Full Text PDF PubMed Scopus Google Scholar). novel finding in study is of proteins R. E. 2002; PubMed Scopus Google Scholar, Science. PubMed Scopus Google Scholar), ITCs may induce the of protein Our indicate that are α- and and and L. showed that aggregation, of a of the of the degradation may the for and tubulin degradation is a result of the of tubulin degradation is and it is to that is It is that the most for both BITC and PEITC to induce tubulin degradation in cells was and of tubulin degradation was when cells treated with than BITC tubulin was with that The inhibition may be by the that both BITC and PEITC at may the ubiquitin-proteasome in finding in study is that both and to tubulin degradation, indicating that tubulin degradation is on of two protein degradation tubulin and degradation A. Johnson Singh S.V. Cancer Res. 2006; PubMed Scopus Google our study a to study protein and degradation by small molecules and their tubulin is essential for cell division, it is conceivable that cells would be to cell growth arrest and it has been that human lung cells and human cells are to cell than the cells D. Zhou Y. Zhang H. Demizu Y. Chen Z. Pelicano H. Chiao P.J. Achanta G. Arlinghaus R.B. Liu J. Huang P. Cancer Cell. 2006; 10: 241-252Abstract Full Text Full Text PDF PubMed Scopus (897) Google Scholar, J. Conaway C.C. Chung F.L. Cancer Res. 2005; PubMed Scopus Google Scholar). we that the anti-microtubule by ITC is at for cell cycle arrest at the phase apoptosis (14Mi L. Xiao Z. Hood B.L. Dakshanamurthy S. Wang X. Govind S. Conrads T.P. Veenstra T.D. Chung F.L. J. Biol. Chem. 2008; 283: 22136-22146Abstract Full Text Full Text PDF PubMed Scopus (181) Google Scholar). The study that tubulin degradation as a of ITC binding may to Although is study provides for the and development of compounds for cancer and Microtubules as a major cytoskeleton component in all eukaryotic cells play essential roles such as maintenance of cell polarity, intracellular traffic, organization, and cell motility (1Hartwell L.H. Kastan M.B. Science. 1994; 266: 1821-1828Crossref PubMed Scopus (2303) Google Scholar, 2Lengauer C. Kinzler K.W. Vogelstein B. Nature. 1998; 96: 643-649Crossref Scopus (3366) Google Scholar, 3White E. Genes Dev. 1996; 10: 1-15Crossref PubMed Scopus (1322) Google Scholar, 4Jordan M.A. Wilson L. Nat. Rev. Cancer. 2004; 4: 253-265Crossref PubMed Scopus (3468) Google Scholar). During cell division, the microtubule-formed mitotic spindle ensures the replicated chromosomes separate evenly at the end of the mitotic phase to the two daughter cells (1Hartwell L.H. Kastan M.B. Science. 1994; 266: 1821-1828Crossref PubMed Scopus (2303) Google Scholar). It is because of its essential roles in cell growth that microtubules become a valid target for the development of anti-microtubule drugs against the rapidly growing cancer cells (2Lengauer C. Kinzler K.W. Vogelstein B. Nature. 1998; 96: 643-649Crossref Scopus (3366) Google Scholar), as interference of microtubule dynamics arrests cell cycle progression and induces apoptosis (3White E. Genes Dev. 1996; 10: 1-15Crossref PubMed Scopus (1322) Google Scholar). Therefore, microtubules have been considered one of the best targets to date for cancer chemotherapy (4Jordan M.A. Wilson L. Nat. Rev. Cancer. 2004; 4: 253-265Crossref PubMed Scopus (3468) Google Scholar). (ITCs) 3The abbreviations used are: ITCisothiocyanateBITCbenzyl isothiocyanatePEITCphenethyl isothiocyanateSFNsulforaphane; N-methylphenethylamineCHAPS3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulfonic acidATZaminotriazoleCHXcycloheximidePBSphosphate-buffered salinePEGpolyethylene glycolRTreverse transcription2-DG2-deoxyglucoseBisTris2-[bis(2-hydroxyethyl)amino]-2-(hydroxymethyl)propane-1,3-diolZbenzyloxycarbonylfmkfluoromethyl ketonePSIproteasome inhibitor IROSreactive oxygen speciesMS/MStandem mass spectrometryIPimmunoprecipitation. 3The abbreviations used are: ITCisothiocyanateBITCbenzyl isothiocyanatePEITCphenethyl isothiocyanateSFNsulforaphane; N-methylphenethylamineCHAPS3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulfonic acidATZaminotriazoleCHXcycloheximidePBSphosphate-buffered salinePEGpolyethylene glycolRTreverse transcription2-DG2-deoxyglucoseBisTris2-[bis(2-hydroxyethyl)amino]-2-(hydroxymethyl)propane-1,3-diolZbenzyloxycarbonylfmkfluoromethyl ketonePSIproteasome inhibitor IROSreactive oxygen speciesMS/MStandem mass spectrometryIPimmunoprecipitation. are among the best studied chemopreventive small molecules (5WHOIARC Handbook on Cancer Prevention. 9. France, Lyon2004: 43-171Google Scholar). The three most studied ITCs, including benzyl-ITC (BITC; abundant in garden cress), phenethyl-ITC (PEITC; in watercress), and sulforaphane (SFN; in broccoli sprouts), have been shown to induce apoptosis and cell cycle arrest (5WHOIARC Handbook on Cancer Prevention. 9. France, Lyon2004: 43-171Google Scholar, 6Chen Y.R. Wang W. Kong A.N. Tan T.H. J. Biol. Chem. 1998; 273: 1769-1775Abstract Full Text Full Text PDF PubMed Scopus (237) Google Scholar, 7Chiao J.W. Chung F.L. Kancherla R. Ahmed T. Mittelman A. Conaway C.C. Int. J. Oncol. 2002; 20: 631-636PubMed Google Scholar, 8Xiao D. Choi S. Lee Y.J. Singh S.V. Mol. Carcinog. 2005; 43: 130-140Crossref PubMed Scopus (31) Google Scholar). Although it is believed that oxidative stress plays a role in cell cycle arrest and apoptosis induced by ITCs (6Chen Y.R. Wang W. Kong A.N. Tan T.H. J. Biol. Chem. 1998; 273: 1769-1775Abstract Full Text Full Text PDF PubMed Scopus (237) Google Scholar, 7Chiao J.W. Chung F.L. Kancherla R. Ahmed T. Mittelman A. Conaway C.C. Int. J. Oncol. 2002; 20: 631-636PubMed Google Scholar, 8Xiao D. Choi S. Lee Y.J. Singh S.V. Mol. Carcinog. 2005; 43: 130-140Crossref PubMed Scopus (31) Google Scholar, 9Zhang Y. Talalay P. Cancer Res. 1998; 58: 4632-4639PubMed Google Scholar, 10Zhang Y. Li J. Tang L. Free Radic. Biol. Med. 2005; 38: 70-77Crossref PubMed Scopus (93) Google Scholar, 11Singh S.V. Srivastava S.K. Choi S. Lew K.L. Antosiewicz J. Xiao D. Zeng Y. Watkins S.C. Johnson C.S. Trump D.L. Lee Y.J. Xiao H. Herman-Antosiewicz A. J. Biol. Chem. 2005; 280: 19911-19924Abstract Full Text Full Text PDF PubMed Scopus (315) Google Scholar, 12Trachootham D. Zhou Y. Zhang H. Demizu Y. Chen Z. Pelicano H. Chiao P.J. Achanta G. Arlinghaus R.B. Liu J. Huang P. Cancer Cell. 2006; 10: 241-252Abstract Full Text Full Text PDF PubMed Scopus (897) Google Scholar), we found that binding to proteins is a predominant intracellular chemical reaction of ITCs, and their protein binding affinities correlate well with inhibition of cell proliferation and induction of apoptosis (13Mi L. Wang X. Govind S. Hood B.L. Veenstra T.D. Conrads T.P. Saha D.T. Goldman R. Chung F.L. Cancer Res. 2007; 67: 6409-6416Crossref PubMed Scopus (128) Google Scholar). Recently, we identified tubulin, the microtubule constituent, as an in vivo target of ITCs by two-dimensional gel electrophoresis and mass spectrometry (14Mi L. Xiao Z. Hood B.L. Dakshanamurthy S. Wang X. Govind S. Conrads T.P. Veenstra T.D. Chung F.L. J. Biol. Chem. 2008; 283: 22136-22146Abstract Full Text Full Text PDF PubMed Scopus (181) Google Scholar). The growth inhibition of human non-small lung cancer A549 cells by ITCs followed the order of BITC > PEITC > SFN. The same order of potency was seen in their binding affinities toward tubulin, induction of its conformational changes, and inhibition of its polymerization. The study provides the first evidence of an in vivo ITC-tubulin binding adduct, indicating that direct modification of cysteines in tubulin by ITCs, rather than oxidative stress, may trigger cell cycle arrest and benzyl phenethyl inhibitor oxygen mass spectrometry benzyl phenethyl inhibitor oxygen mass spectrometry Here we report an unexpected novel finding that tubulin is selectively degraded in a variety of human cancer cells treated with ITCs. We provide evidence that the degradation is initiated by its binding with ITCs and mediated by the ubiquitin-proteasome pathway. Tubulin has long been viewed as a stable and abundant protein, and its levels in cells are tightly regulated (15Nogales E. Annu. Rev. Biochem. 2000; 69: 277-302Crossref PubMed Scopus (518) Google Scholar). In the literature, the only studies on cellular tubulin level change are related to “the auto-regulation theory,” i.e. when microtubules collapse, the presence of a massive amount of tubulin monomers would selectively destabilize tubulin mRNA and subsequently decrease tubulin protein synthesis (16Ben-Ze'ev A. Farmer S.R. Penman S. Cell. 1979; 17: 319-325Abstract Full Text PDF PubMed Scopus (221) Google Scholar, 17Cleveland D.W. Lopata M.A. Sherline P. Kirschner M.W. Cell. 1981; 25: 537-546Abstract Full Text PDF PubMed Scopus (259) Google Scholar, 18Cleveland D.W. Pittenger M.F. Feramisco J.R. Nature. 1983; 305: 738-740Crossref PubMed Scopus (80) Google Scholar). To our knowledge, there is no report on tubulin degradation as a result of treatment with any agents. Our studies provide strong evidence that tubulin as a target of ITCs for cell growth to a mechanism for the anti-microtubule of ITCs through binding to tubulin and a to study protein by modification with small we have shown that binding to intracellular proteins by ITCs is a predominant event that correlates with their apoptosis (13Mi L. Wang X. Govind S. Hood B.L. Veenstra T.D. Conrads T.P. Saha D.T. Goldman R. Chung F.L. Cancer Res. 2007; 67: 6409-6416Crossref PubMed Scopus (128) Google Scholar). We have in human lung cancer A549 cells that tubulin is a protein target of ITCs by two-dimensional gel and BITC a with tubulin in vivo (14Mi L. Xiao Z. Hood B.L. Dakshanamurthy S. Wang X. Govind S. Conrads T.P. Veenstra T.D. Chung F.L. J. Biol. Chem. 2008; 283: 22136-22146Abstract Full Text Full Text PDF PubMed Scopus (181) Google Scholar). we found that BITC and PEITC bind tubulin with are in inducing tubulin conformational changes, microtubule and growth inhibition and In and no binding to tubulin, are both of of the The study shows that BITC and PEITC can selectively induce and degradation of both α- and β-tubulins in a variety of cancer cells. of tubulin to ITCs is a The potency of inducing tubulin degradation the order BITC PEITC > correlates with their on the in and in vivo we a of tubulin degradation ITCs rapidly α- and β-tubulins in through binding to The modification by ITCs tubulin conformational changes, and The tubulin are degraded by the The including and the BITC binding to tubulin, conformational changes, and tubulin and In and can tubulin degradation This that and degradation are two related tubulin degradation is of oxidative stress. It be that results both and A549 cells indicating that the mechanism may be cell The tubulin degradation by small molecules is novel and as a of is that when a cell it two of P. J. Y. J. C. Nat. Biol. PubMed Scopus Google Scholar). it a variety of proteins to the protein to its the protein is the cell its degradation Nature. PubMed Scopus Google Scholar). event of proteins is that to Full Text Full Text PDF PubMed Scopus Google Scholar). The that the of proteins the of degradation of the cell the of the Y. M.A. R. D.L. D. C.C. T.P. Nature. 2007; PubMed Scopus Google Scholar, A. J. Biol. Chem. 2005; 280: Full Text Full Text PDF PubMed Scopus Google Scholar). novel finding in study is of proteins R. E. 2002; PubMed Scopus Google Scholar, Science. PubMed Scopus Google Scholar), ITCs may induce the of protein Our indicate that are α- and and and L. showed that aggregation, of a of the of the degradation may the for and tubulin degradation is a result of the of tubulin degradation is and it is to that is It is that the most for both BITC and PEITC to induce tubulin degradation in cells was and of tubulin degradation was when cells treated with than BITC tubulin was with that The inhibition may be by the that both BITC and PEITC at may the ubiquitin-proteasome in finding in study is that both and to tubulin degradation, indicating that tubulin degradation is on of two protein degradation tubulin and degradation A. Johnson Singh S.V. Cancer Res. 2006; PubMed Scopus Google our study a to study protein and degradation by small molecules and their tubulin is essential for cell division, it is conceivable that cells would be to cell growth arrest and it has been that human lung cells and human cells are to cell than the cells D. Zhou Y. Zhang H. Demizu Y. Chen Z. Pelicano H. Chiao P.J. Achanta G. Arlinghaus R.B. Liu J. Huang P. Cancer Cell. 2006; 10: 241-252Abstract Full Text Full Text PDF PubMed Scopus (897) Google Scholar, J. Conaway C.C. Chung F.L. Cancer Res. 2005; PubMed Scopus Google Scholar). we that the anti-microtubule by ITC is at for cell cycle arrest at the phase apoptosis (14Mi L. Xiao Z. Hood B.L. Dakshanamurthy S. Wang X. Govind S. Conrads T.P. Veenstra T.D. Chung F.L. J. Biol. Chem. 2008; 283: 22136-22146Abstract Full Text Full Text PDF PubMed Scopus (181) Google Scholar). The study that tubulin degradation as a of ITC binding may to Although is study provides for the and development of compounds for cancer and we have shown that binding to intracellular proteins by ITCs is a predominant event that correlates with their apoptosis (13Mi L. Wang X. Govind S. Hood B.L. Veenstra T.D. Conrads T.P. Saha D.T. Goldman R. Chung F.L. Cancer Res. 2007; 67: 6409-6416Crossref PubMed Scopus (128) Google Scholar). We have in human lung cancer A549 cells that tubulin is a protein target of ITCs by two-dimensional gel and BITC a with tubulin in vivo (14Mi L. Xiao Z. Hood B.L. Dakshanamurthy S. Wang X. Govind S. Conrads T.P. Veenstra T.D. Chung F.L. J. Biol. Chem. 2008; 283: 22136-22146Abstract Full Text Full Text PDF PubMed Scopus (181) Google Scholar). we found that BITC and PEITC bind tubulin with are in inducing tubulin conformational changes, microtubule and growth inhibition and In and no binding to tubulin, are both of of the The study shows that BITC and PEITC can selectively induce and degradation of both α- and β-tubulins in a variety of cancer cells. of tubulin to ITCs is a The potency of inducing tubulin degradation the order BITC PEITC > correlates with their on the in and in vivo we a of tubulin degradation ITCs rapidly α- and β-tubulins in through binding to The modification by ITCs tubulin conformational changes, and The tubulin are degraded by the The including and the BITC binding to tubulin, conformational changes, and tubulin and In and can tubulin degradation This that and degradation are two related tubulin degradation is of oxidative stress. It be that results both and A549 cells indicating that the mechanism may be cell The tubulin degradation by small molecules is novel and as a of It is that when a cell it two of P. J. Y. J. C. Nat. Biol. PubMed Scopus Google Scholar). it a variety of proteins to the protein to its the protein is the cell its degradation Nature. PubMed Scopus Google Scholar). event of proteins is that to Full Text Full Text PDF PubMed Scopus Google Scholar). The that the of proteins the of degradation of the cell the of the Y. M.A. R. D.L. D. C.C. T.P. Nature. 2007; PubMed Scopus Google Scholar, A. J. Biol. Chem. 2005; 280: Full Text Full Text PDF PubMed Scopus Google Scholar). novel finding in study is of proteins R. E. 2002; PubMed Scopus Google Scholar, Science. PubMed Scopus Google Scholar), ITCs may induce the of protein Our indicate that are α- and and and L. showed that aggregation, of a of the of the degradation may the for and tubulin degradation is a result of the of tubulin degradation is and it is to that is It is that the most for both BITC and PEITC to induce tubulin degradation in cells was and of tubulin degradation was when cells treated with than BITC tubulin was with that The inhibition may be by the that both BITC and PEITC at may the ubiquitin-proteasome in finding in study is that both and to tubulin degradation, indicating that tubulin degradation is on of two protein degradation tubulin and degradation A. Johnson Singh S.V. Cancer Res. 2006; PubMed Scopus Google our study a to study protein and degradation by small molecules and their tubulin is essential for cell division, it is conceivable that cells would be to cell growth arrest and it has been that human lung cells and human cells are to cell than the cells D. Zhou Y. Zhang H. Demizu Y. Chen Z. Pelicano H. Chiao P.J. Achanta G. Arlinghaus R.B. Liu J. Huang P. Cancer Cell. 2006; 10: 241-252Abstract Full Text Full Text PDF PubMed Scopus (897) Google Scholar, J. Conaway C.C. Chung F.L. Cancer Res. 2005; PubMed Scopus Google Scholar). we that the anti-microtubule by ITC is at for cell cycle arrest at the phase apoptosis (14Mi L. Xiao Z. Hood B.L. Dakshanamurthy S. Wang X. Govind S. Conrads T.P. Veenstra T.D. Chung F.L. J. Biol. Chem. 2008; 283: 22136-22146Abstract Full Text Full Text PDF PubMed Scopus (181) Google Scholar). The study that tubulin degradation as a of ITC binding may to Although is study provides for the and development of compounds for cancer and We S. of of and of Cancer at for and We and for and of for and for with with