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Use of micro-rotation imaging to study JNK-mediated cell survival in Theileria parva-infected B-lymphocytes

Published online by Cambridge University Press:  16 February 2005

R. LIZUNDIA ,
L. SENGMANIVONG ,
J. GUERGNON ,
T. MÜLLER ,
T. SCHNELLE ,
G. LANGSLEY  and
S. L. SHORTE
R. LIZUNDIA
Affiliation:
Laboratoire du Signalisation Immunoparasitaire, URA CNRS 2851, Department Parasitologie, Institut Pasteur, Paris, France
L. SENGMANIVONG
Affiliation:
Platefome d'Imagerie Dynamique, Department Biologie Cellulaire et Infection, Institut Pasteur, 25–28 rue du Dr. Roux, 75015, Paris, France
J. GUERGNON
Affiliation:
Laboratoire du Signalisation Immunoparasitaire, URA CNRS 2851, Department Parasitologie, Institut Pasteur, Paris, France
T. MÜLLER
Affiliation:
Cell Handling and Analysis, Evotec Technologies GmbH, c/o Institut für Biologie der Humboldt-Universität zu Berlin, Invalidenstrasse 42, D-10115 Berlin, Germany
T. SCHNELLE
Affiliation:
Cell Handling and Analysis, Evotec Technologies GmbH, c/o Institut für Biologie der Humboldt-Universität zu Berlin, Invalidenstrasse 42, D-10115 Berlin, Germany
G. LANGSLEY
Affiliation:
Laboratoire du Signalisation Immunoparasitaire, URA CNRS 2851, Department Parasitologie, Institut Pasteur, Paris, France
S. L. SHORTE
Affiliation:
Platefome d'Imagerie Dynamique, Department Biologie Cellulaire et Infection, Institut Pasteur, 25–28 rue du Dr. Roux, 75015, Paris, France
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Abstract

Lymphocytes infected with the protozoan parasite Theileria parva are transformed to permanently proliferating cells, an event underlying the pathology of the disease. However, the molecular signalling mediating this process is complex and poorly understood. Here, we show that down-regulation of JNK signalling by transient over expression of a dominant-negative mutant of JNK (JNK-APF) significantly increases Annexin-V-phycoerythrin (V-PE) labelling on infected B cell populations observed using flow cytometry. To establish whether this increase was specifically due to apoptosis, we used a novel single-cell imaging method: micro-rotation (MR)-imaging, designed to allow high-resolution 3-dimensional imaging of single cells in suspension. With this method we visualized subcellular patterns of V-PE uptake and chromatin organization in lymphocytes co-transfected with JNK-APF and GFP-tagged histone-H2B. This single-cell approach allowed us to clearly reveal characteristic apoptotic phenotypes, whose patterns reflected progressive states of programmed cell death due to JNK down-regulation. Our results strongly suggest a role for JNK in the survival of Theileria-infected B cells, and demonstrate the powerful utility of a new and unique 3-dimensional imaging method for living cells in suspension.

Keywords

TheileriaapoptosisB-lymphocytechromatinJNKmicro-rotation imaging
Type
Research Article
Information
Parasitology , Volume 130 , Issue 6 , June 2005 , pp. 629 - 635
Copyright
© 2005 Cambridge University Press

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References

REFERENCES

AMEYAR, M., WISNIEWSKA, M. & WEITZMAN, J. B. ( 2003). A role for AP-1 in apoptosis: the case for and against. Biochimie 85, 747–752.CrossRefGoogle Scholar
BAUMGARTNER, M., CHAUSSEPIED, M., MOREAU, M. F., WERLING, D., DAVIS, W. C., GARCIA, A. & LANGSLEY, G. ( 2000). Constitutive PI3-K activity is essential for proliferation, but not survival, of Theileria parva-transformed B cells. Cellular Microbiology 2, 329–339.CrossRefGoogle Scholar
BAUMGARTNER, M., ANGELISOVA, P., SETTERBLAD, N., MOONEY, N., WERLING, D., HOREJSI, V. & LANGSLEY, G. ( 2003). Constitutive exclusion of Csk from Hck-positive membrane microdomains permits Src kinase-dependent proliferation of Theileria-transformed B lymphocytes. Blood 101, 1874–1881.CrossRefGoogle Scholar
BOTTERON, C. & DOBBELAERE, D. ( 1998). AP-1 and ATF-2 are constitutively activated via the JNK pathway in Theileria parva-transformed T-cells. BBRC 246, 418–421.Google Scholar
CHAUSSEPIED, M., LALLEMAND, D., MOREAU, M. F., ADAMSON, R., HALL, R. & LANGSLEY, G. ( 1998). Upregulation of Jun and Fos family members and permanent JNK activity lead to constitutive AP-1 activation in Theileria-transformed leukocytes. Mol Biochem Parasitol 94, 215–226.CrossRefGoogle Scholar
CHAUSSEPIED, M. & LANGSLEY, G. ( 1996). Theileria transformation of bovine leukocytes: a parasite model for the study of lymphoproliferation. Res Immunol 147, 127–138.CrossRefGoogle Scholar
DOBBELAERE, D. A. & ROTTENBERG, S. ( 2003) Theileria-induced leukocyte transformation. Curr Opin Microbiol 6, 377–382.CrossRefGoogle Scholar
GUERGNON, J., DESSAUGE, F., LANGSLEY, G. & GARCIA, A. ( 2003 a). Apoptosis of Theileria-infected lymphocytes induced upon parasite death involves activation of caspases 9 and 3. Biochimie 85, 771–776.Google Scholar
GUERGNON, J., CHAUSSEPIED, M., SOPP, P., LIZUNDIA, R., MOREAU, M. F., BLUMEN, B., WERLING, D., HAWARD, C. J. & LANGSLEY, G. ( 2003 b). A tumour necrosis factor alpha autocrine loop contributes to proliferation and nuclear factor-kappaB activation of Theileria parva-transformed B cells. Cell Microbiol 5, 709–716.Google Scholar
HESS, P., PIHAN, G., SAWYERS, C. L., FLAVELL, R. A. & DAVIS, R. J. ( 2002). Survival signaling mediated by c-Jun NH(2)-terminal kinase in transformed B lymphoblasts. Nat Genet. 32, 201–205.CrossRefGoogle Scholar
HEUSSLER, V. T., KUENZI, P., FRAGA, F., SCHWAB, R. A., HEMMINGS, B. A. & DOBBELAERE, D. A. ( 2001). The Akt/PKB pathway is constitutively activated in Theileria- transformed leucocytes, but does not directly control constitutive NF- kappaB activation. Cell Microbiol 3, 537–550.CrossRefGoogle Scholar
HEUSSLER, V. T., ROTTENBERG, S., SCHWAB, R., KUENZI, P., FERNANDEZ, P. C., McKELLAR, S., SHIELS, B., CHEN, Z. J., ORTH, K., WALLACH, D. & DOBBELAERE, D. A. ( 2002). Hijacking of host cell IKK signalosomes by the transforming parasite Theileria. Science 298, 1033–1036.CrossRefGoogle Scholar
HUANG, C., RAJFUR, Z., BORCHERS, C., SCHALLER, M. D. & JACOBSON, K. ( 2003). JNK phosphorylates paxillin and regulates cell migration. Nature, London 424, 219–223.CrossRefGoogle Scholar
INOSHITA, S., TAKEDA, K., HATAI, T., TERADA, Y., SANO, M., HATA, J., UMEZAWA, A. & ICHIJO, H. ( 2002). Phosphorylation and inactivation of myeloid cell leukemia 1 by JNK in response to oxidative stress. J Biol Chem 277, 43730–43734.CrossRefGoogle Scholar
IRVIN, A. D., DOBBELAERE, D. A., MWAMACHI, D. M., MINAMI, T., SPOONER, P. R. & OCAMA, J. G. ( 1983). Immunisation against East Coast fever: correlation between monoclonal antibody profiles of Theileria parva stocks and cross immunity in vivo. Res. Vet. Sci. 35, 341–346.Google Scholar
KANDA, T., SULLIVAN, K. F. & WAHL, G. M. ( 1998) Histone-GFP fusion protein enables sensitive analysis of chromosome dynamics in living mammalian cells. Curr Biol 8, 377–385.CrossRefGoogle Scholar
KUENZI, P., SCHNEIDER, P. & DOBBELAERE, D. A. ( 2003) Theileria parva-transformed T cells show enhanced resistance to Fas/Fas ligand-induced apoptosis. J Immunol 171, 1224–1231.CrossRefGoogle Scholar
MOREAU, M.-F., THIBAUT, J.-L., BEN MILED, L., CHAUSSEPIED, M., BAUMGARTNER, M., DAVIS, W. C., MINOPRIO, P., LANGSLEY, G. ( 1999). Theileria annulata and CD5-positive macrophages and B1 cells. Inf Immunity 67, 6678–6682.Google Scholar
TOURNIER, C., DONG, C., TURNER, T. K., JONES, S. N., FLAVELL, R. A. & DAVIS, R. J. ( 2001). MKK7 is an essential component of the JNK signal transduction pathway activated by proinflammatory cytokines. Genes Dev 15, 1419–1426.CrossRefGoogle Scholar
WESTON, C. R. & DAVIS, R. J. ( 2002). The JNK signal transduction pathway. Curr Opin Genet Dev 12, 14–21.CrossRefGoogle Scholar
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M1B Supplementary movie

Movie 1B. MR-imaging of H2B-GFP fluorescence signal inside individual JNKN-APF co-transfected B cell.

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M2 Supplementary movie

Movie 2. MR-imaging of H2B-GFP and anexin V-PE labelling inside apoptotic Theileria-infected B cell.

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