Stress, Chaperones and Cell Death

Group composition Leader: André-Patrick ARRIGO arrigo@univ-lyon1.fr http://www.heatshock.net/showlab.php?sort=score&surname=Arrigo&initials=AP Staff members: André-Patrick ARRIGO, Professor Chantal DIAZ-LATOUD, Assistant-Professor Carole KRETZ-REMY, Assistant-Professor Nicole MOUNIER, Assistant-Professor Post-doct, Ph.D students: Stéphanie SIMON, Post-doct, AFM. Maryline MOULIN, Post-doct Benjamin GIBERT, Ph.D student, Région Rhône-Alpes fellow Mathieu NIVON, Ph.D student, MRT fellow Emma RICHET, Undergraduate M2 Lydie FASQUELLE, Undergraduate M2 Technical staff: Dominique GUILLET, T2 UCB Grants: Région Rhône-Alpes, Ligue, AFM

Protective activity of Hsp27 against H2O2 induced necrotic cell death

Scientific themes

Our research aims are focused towards a better understanding of the molecular mechanisms and significance of the cellular response to stress.

Most physical and chemical stress interfere with the folding of intracellular proteins. Consequently, heat shock or stress proteins (Hsps) are synthetized. Hsps, through their chaperone activity, refold altered polypeptides and are indispensable to counteract the deleterious damages induced by stress. Among the Hsps, the so-called Small Hsps (sHsps) are characterized by their common α-crystallin domain.

Hsp27 is a phospho-oligomeric polypeptide. In cells exposed to stress, Hsp27 large oligomeric structures store abberantly folded polypeptides before they are refolded by ATP-dependent chaperones (Hsp70, Hsp90..). In contrast, Hsp27 small oligomers act towards F-actin and stabilize cytoskeleton. We were first to demonstrate that Hsp27 and alphaB-crystallin have anti-oxidant and anti-apoptotic properties. In pathologies characterized by enhanced cell death (i.e asthma, neurodegeneration) Hsp27 expression can be beneficial. Indeed, we have shown that in cells derived from these pathologies, Hsp27 expression decreases the deleterious effects induced by aggregated or oxidized proteins. In pathologies characterized by deficient cell death, Hsp27 expression has often deleterious effects. For example, Hsp27 is overexpressed in numerous cancer cells (breast, head-neck, colon...) and appears essential for the tumorigenic and invasive potential of these cells. The molecular mechanism of the protective activity of Hsp27 in cancer cells is studied. We are also involved in a project aimed at finding Hsp27 inhibitors using a peptide aptamer approach.

alphaB-crystallin (αB) is constitutively expressed in the eye crystallin as well as in skeletal and cardiac muscles. In muscle cells, this Hsp regulates desmin cytoskeleton assembly and acts, in response to stress, as a molecular chaperone that prevents intermediate filaments aggregation. αB has a major role in control of muscle integrity. Indeed, mutations in αB gene induce alphaB-crystallinopathies (myofibrillar myopathies often associated with cardiomyopathies and cataract). Inhibition of aggresomes formation (resulting of pathological mutations in αB) appears as a valid strategy to attenuate the developement alphaB-crystallinopathies. Our aim is to obtain peptide aptamers that specifically recognize the mutated pathological forms (but not wild-type) of αB. The ability of these peptides to restore mutated αB chaperone activity will be tested. This may lead to the discovery of chemical chaperones that counteract the pathological effect induced by mutated αB.

Apoptosis induced by TRAIL is efficient only in cancer cells. However, since in vivo this cell death process is not very efficient new stimulators of TRAIL apoptosis are needed. In this regard we recently reported that a mild heat shock treatment (1h at 42°C) strongly stimulates TRAIL apoptosis in human leukemic (lymphocytic and promyelocytic) cells. The same phenomenon is observed in primary cells from patients suffering from chronical lymphocytic leukemia (CLL). Of interest, the co-treatment (heat shock plus TRAIL) has no effect in normal non leukemic cells. The molecular mechanism reveals that, in leukemic cells, heat shock increases plasma membrane fluidity as well as ceramide levels and consequently enhances the recognition of DR4 and DR5 receptors by TRAIL. Drugs that enhance membrane fluidity are tested in combination with TRAIL. In this regard, we have reported that alcohols are potent stimulators of TRAIL apoptosis. The project will next be oriented towards a medical application.

We have studied several mechanisms of activation of NF-κB. This factor regulates the transcription of the major genes involved in the inflammatory process. The activation depends on either a phosphorylation and degradation of the inhibitory sub-unit IκB-alpha or on its dissociation from the dimeric p65-p50 NF-κB complex. Thereafter, p65-p50 migrates inside the nucleus and activates target genes that bear κB sites in their promoter region. We have shown that NF-κB is activated during the recovery phase that follows a heat shock treatment. This activation is characterized by the dissociation of IκB-alpha from p65-p50 complex. The nature of the responsive mechanism is studied. We are also analyzing the heat shock response in cells devoid of p65 expression (RNAi approach). We have observed a strong decrease in cell survival in cells unable to mount an NF-κB driven response after heat shock. This enhanced cell death appears to be a consequence of a reduced autophagic response after heat shock. We are now testing where and how in the autophagy pathway NF-κB acts: autophagosomes formation or upstream transduction signals ? In vivo, the observation may have considerable impact towards fever and inflammation processes.

Publications since 2000

Aloy, M.-T., Hadchity, E., Bonda C., Diaz-Latoud, C., Line C., Rousson, R., Arrigo, A.-P. and Rodriguez-Lafrasse, C. Protective role of Hsp27 protein against gamma-radiation induced apoptosis and radiosensitization effects of Hsp27 gene silencing in different human tumor cells.
The International Journal of Radiation Oncology, Biology, Physics. Epub ahead of print. Oct 31 (2007).

Arrigo A.-P. Anti-apoptotic, tumorigenic and metastatic potential of Hsp27 (HspB1) and alphaB-crystallin (HspB5): emerging targets for the development of new anti-cancer therapeutic strategies. In "Heat Shock Proteins in Cancer". Eds. Stuart K Calderwood, Michael Sherman and Daniel Ciocca. Springer-Verlag. pp 73-92 (2007).

Moulin, M, Carpentier,, S., Levade,, T., and A.-P. Arrigo. Potential roles of membrane fluidity and ceramide elevation in the mechanism of hyperthermia- and alcohol-stimulated TRAIL apoptosis. Apoptosis 12, 1703-1720 (2007).

Arrigo, A.-P., Simon, S., Gibert, B., Kretz-Remy, C., Nivon M., Czekalla, A., Guillet, D., Moulin, M., Diaz-Latoud, C., and P. Vicart, P. Hsp27 (HspB1) and aB-crystallin (HspB5) as therapeutic targets. FEBS Lett. 581, 3665-3674 (2007).

Arrigo, A.-P. The cellular "Networking" of Hsp27 and its functions in the control of protein folding, redox state and apoptosis. In : Molecular Aspects of the Stress Response: Chaperones, Membranes and Networks. Lazlo Vight and Peter Csermely Eds. Landes Bioscience and Springer Science+Business Media.
Advances in Experimental Medicine and Biology. 594, 16-26 (2007).

Lemaire F., Mandon CA, Reboud J., Papine, A., Angulo J., Pointu, H., Diaz-Latoud C., Lajaunie C., Chatelain F., Arrigo, A.-P., and B. Schaack. Toxicity assays in nanodrops combining bioassay and morphometric Endpoints. PloS One. 2(1) e163 (2007).

Wyttenbach, A., and A.-P. Arrigo. The role of heat shock proteins during neurodegeneration in Alzheimer's, Parkinson's and Huntington's diseases. In : Heat Shock Proteins in the Nervous System. Eds. Christiane Richter-Landsberg. Landes Biosciences. pp 81-99 (2007).

Moulin M., Dumontet, C., and A.-P. Arrigo. Sensitization of chronic lymphocytic leukemia cells to TRAIL-induced apoptosis by hyperthermia.
Cancer Lett. 250, 117-127.(2007).

Firdaus, W.J.J, Wyttenbach, A., Kretz-Remy, C., Currie, W., and A.-P. Arrigo. Huntingtin granules are iron-dependent centers of oxidative events.
FEBS Journal. 273, 5428-5441 (2006).

Mandon, C., Diaz-Latoud, C., Arrigo, A.-P. and L. Blum. Chemical stress sensitive luminescent human cells: molecular biology approach using inducible Drosophila melanogaster hsp22 promoter. Biochemical and Biophysical Research Communications. 335, 536-544 (2005).

Firdaus, W.J.J, Wyttenbach, A., Currie, W., Diaz-Latoud, C., and A.-P. Arrigo. Analysis of oxidative events induced by expanded polyglutamine huntingtin exon1 which are differentially restored by expression of Hsps or treatment with an anti-oxidant. FEBS Journal. 273, 3076-3093 (2006).

Mandon, C., Diaz-Latoud, C., Arrigo, A.-P. and L. Blum. Dithiocarbamate fungicide thiram detection: comparison of bioluminescent and fluorescent whole-cell bioassays based on hsp22 promoter induction. J. of Biotechnology, 124, 392-402 (2006).

Moulin, M. and A.-P. Arrigo. Long lasting heat shock stimulation of TRAIL-induced apoptosis in leukemic T lymphocytes.Exp. Cell Res. 312, 1765-1784 (2006).

Arrigo, A.-P., Virot, S., Chaufour, S., Firdaus, W.J.J., Kretz-Remy, C., and C. Diaz-Latoud. Hsp27 consolidates intracellular redox homeostasis by upholding glutathione in its reduced form and by decreasing iron intracellular levels.
Antioxidants and Redox Signaling7, 414-422 (2005).

Arrigo, A.-P., Firdaus, W.J.J , Mellier G., Moulin, M., Paul, C. Diaz-Latoud, C., and C. Kretz-Remy. Cytotoxic effects induced by oxidative stress in cultured mammalian cells and protection provided by Hsp27 expression. Methods 35, 126-138 (2005).

Favre-Bonvin A, Reynaud C, Kretz-Remy C, Jalinot P. Human papillomavirus type 18 E6 protein binds the cellular PDZ protein TIP-2/GIPC, which is involved in transforming growth factor beta signaling and triggers its degradation by the proteasome. J Virol. 2005 Apr;79(7):4229-37.

Arrigo, A.-P. In search of the molecular mechanism by which small stress proteins counteract apoptosis during cellular differentiation.
J. Cellular Biochemistry. 94, 241-246 (2005).

Arrigo, A.-P. Chaperons moléculaires : l'exemple des protéines de choc thermique. Médecine/Sciences. 21 (6-7), 619-625 (2005).

Franklin, T.B., Krueger-Naug, A.M., Clarke, D.B., Arrigo, A.-P., and R.W. Currie. Heat shock proteins and cellular protection in the central nervous system.
Int. J. Hyperhermia. 21, 379-392 (2005).

Diaz-Latoud, C., Buache E., Javoueh, E., and A.-P. Arrigo. Substitution of the unique cysteine residue of murine Hsp25 interferes with the protective activity of this stress protein through dimer formation inhibition. Antioxidants and Redox Signaling. 7, 436-445 (2005).

Giraud S, Diaz-Latoud C, Hacot S, Textoris J, Bourette RP, Diaz JJ. US11 of herpes simplex virus type 1 interacts with HIPK2 and antagonizes HIPK2-induced cell growth arrest. J Virol. 78(6):2984-2993 (2004).

Manero, F., Lujbic-Thibal, V., Moulin, M., Yvin, J.-Cl., and A.-P. Arrigo. Heparin-like agents stimulate low levels of apoptosis induced by Fas agonistic antibody : different inhibitory effects of Hsp27 and Bcl-2. Cell Stress and Chaperones. 9,150-166 (2004).

Chen Yu, Arrigo, A.-P., and R.W. Currie. Heat shock treatment suppresses angiotensin II-activation of NF-kB pathway and heart inflammation : a role for IKK depletion by heat shock ? American Journal of Physiology, Heart and Circulation Physiology. 287, 1104-1114 (2004).

Aït-Aïssa, S., Pandard, P., Magaud, H., Arrigo, A.-P., Thybaud, E, and J.-M. Porcher.. Evaluation of an in vitro hsp70 induction test for toxicity assessment of complex mixtures : comparison with chemical analyses and ecotoxicity tests. Ecotoxicol Environ Saf. 54(1), 92-104 (2003).

Kretz-Remy, C. and A.-P. Arrigo. Modulation of the chymotrypsin-like activity of the 20S proteasome by intracellular redox status : effects of glutathione peroxidase overexpression and anti-oxidant drugs. Chem. Biol. 384, 589-595 (2003).

Kretz-Remy, C. and Arrigo, A.-P. Gene expression ansd the thiol redox state. In: Protein sensors of reactive oxygen species: Thiol enzymes and proteins. Methods in Enzymology. (Helmut Sies and Lester Packer, Eds). Academic Press. Inc. Orlando, Florida 32887 USA.vol. 348, pp 200-215 (2002).

Arrigo, A.-P. and Mehlen, P. ES cells as a model for stress proteins and apoptosis during differentiation. Methods in Molecular Biology,vol.184:Embryonic Stem Cells:Methods and Protocols. Edited by:K.Turksen © Humana Press Inc.,Totowa, NJ, pp 35-44 (2002).

Arrigo, A.-P., Paul, C., Ducasse, C., Sauvageot, O., and Kretz-Remy , C. Small stress proteins : modulation of intracellular redox state and protection against oxidative stress induced cytotoxicity. In : Small stress proteins. Progress in Molecular and Subcellular Biology, Vol. 28. (A.-P. Arrigo and W. E.G. Muller, Eds). Springer Verlag, Berlin, Heidelberg, New-York, Tokyo. pp 171-184 (2002).

Arrigo, A.-P., Paul, C., Ducasse, C., Manero, F., Kretz-Remy, C., Javouhey, E., Virot, S., Mounier, N., and Diaz-Latoud, C. Small stress proteins : novel regulators of apoptosis induced independently of reactive oxygen species. In : Small stress proteins. Progress in Molecular and Subcellular Biology, Vol. 28. (A.-P. Arrigo and W. E.G. Muller, Eds). Springer Verlag, Berlin, Heidelberg, New-York, Tokyo. pp 185- 204 (2002).

Paul, C., Manero, F., Gonin, S., Kretz-Remy, C., Virot, S., and Arrigo, A.-P. Hsp27 as a negative regulator of cytochrome c release.
Mol. Cell Biol. 22, 816-834 (2002).

Mounier, N. and Arrigo, A.-P. sHSP and actin: How do they interact ? Cell Stress and Chaperones. 7, 167-176 (2002).

Wyttenbach, A., Sauvageot, O.,Carmichael, J., Diaz-Latoud, C., Arrigo, A.-P., Rubinsztein, D.C. Heat shock protein 27 prevents cellular polyglutamine toxicity and suppresses the increase of reactive oxygen species caused by huntingtin.
Human Molecular Genetics. 11, 1137-1151 (2002).

Merendino, A., Paul, C., Costa, M A., Melis, M., Chiappara, G., Izzo, V., Vignola, A.M., Arrigo, A.-P. Heat shock protein-27 protects human bronchial epithelial cells against oxidative stress-mediated apoptosis : implication in asthma.
Cell Stress and Chaperones. 7, 269-280 (2002).

Manero, F., and Arrigo, A.-P. The Fas/FasL system as a critical pathway of cancer cell apoptosis. Transworld Research Network.
Recent Research Developments In Cancer. 4, 353-377 (2002).

Arrigo, A.-P., and Ducasse, C. Expression of the anti-apoptotic protein Hsp27 during both the keratinocyte differentiation and dedifferentiation of HaCat keratinocytes : expression linked to changes in intracellular protein organization ?
Experimental Gerontology. 37, 1247-1255 (2002).

Samali, A., Robertson, J.D., Peterson, E., Manero, F., van Zeijl ,L., Paul, C., Cotgreave, I. A., Arrigo, A.-P. and S. Ohrrenius. Hsp27 protects mitochondria of thermotolerant cells against apoptotic stimuli. Cell Stress and Chaperones. 6, 49-58 (2001).

Pieri, I. , Cifuentez-Diaz, C., Oudinet, JP., Blondet , B., Rieger, F., Gonin, S., Arrigo, A.-P., and Y Thomas. Regulation of Hsp25 expression during anterior horn motor neuron degeneration in the paralyze mouse mutant.
Journal of Neuroscience Research. 65, 247-253 (2001).

Kretz-Remy, C. and Arrigo, A.-P. Selenium : a key element that controls NF-kB activation and IkBa half life.
BioFactors. 14, 117-125 (2001).

Kretz-Remy, C., Munsch, B., and A.-P. Arrigo. kB-dependent transcriptional activation during heat shock recovery: a new mechanism of NF-kB activation.
J. Biol. Chem. 47, 43723-43733 (2001).

Siena, L., Merendino, A.M., Chanez, P., Polla, B., Chiappara, G., Arrigo. A.-P., Reina, C., Bousquet, J., Bonsignore, G., Vignola, A.M. Increased expression of antiapoptotic factors by bronchial epithelial cells is associated with decreased apoptosis in patients with asthma and chronic bronchitis.
J of Allergy and Clinical Immunology.108, 738-746 (2001).

Arrigo, A.-P. Small stress proteins: novel regulators of intracellular redox state. IUBMB Life.52, 303-307 (2001).

Carrier, H., Flocard, F., Tagliati, V., Arrigo, A.-P. and Godinot, C. Immunolabelling of mitochondrial superoxide dismutase and of Hsp60 in muscles harboring a respiratory chain deficiency. Neuromuscular Disorders. 10, 144-149 (2000).

Garrido, C., Arrigo, A.-P., and Solari, E. Hsp27 : A small heat shock protein with protective and tumorigenic effects. Transworld Research Network.
Recent Res. Devel. Cancer. 2, 105-114 (2000).

Aït-Aïssa, S., Porchera, J-M. , Arrigo, A.-P., and Lambré, C. Activation of the hsp70 promoter by environmental inorganic and organic chemicals: relationships with cytotoxicity and lipophilicity. Toxicology. 145, 147-157 (2000).

Arrigo, A.-P. sHsp as novel regulators of programmed cell death and tumorigenicity. Pathologie Biologie. 48, 280-288 (2000).

Bruey, J.-M., Ducasse, C., Bonniaud, P., Ravagnan, L., Susin, S.A., Diaz-Latoud, C., Arrigo, A.-P., Kroemer, G., Solary, E., Garrido, C. Hsp27 negatively regulates cell death by interacting with cytochrome c. Nature Cell Biol. 2, 645-652 (2000).

Paul, C., Bruey, J.M., Hilpert, S., Arrigo, A.-P., Solari, E., and Garrido, C. Hsp27 large unphosphorylated aggregates mediate its antiapoptotic and tumorigenic effects. Oncogene. 19, 4855-4863 (2000).

Paul C.and Arrigo, A.-P. Comparison of the protective activities generated by two survival proteins Bcl-2 and Hsp27 in L929 murine fibroblasts exposed to menadione or staurosporine. Experimental Gerontology. 35, 757-766 (2000).