Advertisement

Neurochemical Research

, Volume 21, Issue 4, pp 417–422 | Cite as

Antisense oligonucleotide to the 70-kDa heat shock cognate protein inhibits synthesis of myelin basic protein

  • Dennis A. Aquino
  • Carmen Lopez
  • Muhammad Farooq
Original Articles

Abstract

Transfection of rat oligodendrocytes with an oligonucleotide sequence complementary to the mRNA encoding the initial ten amino acids of the rat 70-kDa heat shock cognate protein (HSC70) resulted in a rapid (within 24 h) and significant reduction in HSC70 synthesis (69% of control cells transfected with sense oligonucleotide). A further decrease to approximately 44% of controls was detected after 2 days. At that time, HSC70 protein content fell to approximately 49% of controls, and a significant reduction in the synthesis of myelin basic protein (MBP) was first detected (66% of controls). After 5 days, HSC70 synthesis returned to control levels. As HSC70 protein content recovered, so did the synthesis of MBP. Throughout the 5-day experimental period, only minor changes were detected in cell morphology, overall pattern of protein synthesis and the synthesis and content of proteolipid protein (PLP) and the pi isoenzyme of glutathione-S-transferase (pi). These data show that when HSC70 protein content is sufficiently reduced by antisense oligonucleotide, synthesis of MBP (but not PLP or pi) is correspondingly down-regulated, and provide evidence consistent with the role of HSC70 as a chaperone for MBP.

Key Words

Chaperone 70-kDa heat shock cognate protein myelin basic protein oligodendrocyte protein synthesis antisense oligonucleotide 

Preview

Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.

References

  1. 1.
    Morimoto, R. I., Tissières, A., and Georgopoulos, C. 1994. Progress and perspectives on the biology of heat shock proteins and molecular chaperones. Pages 1–30,in Morimoto, R. I., Tissières, A., and Georgopoulos, C. (eds.), The Biology of Heat Shock Proteins and Molecular Chaperones, Cold Spring Harbor Laboratory Press, Plainview.Google Scholar
  2. 2.
    Roobol, A., and Carden, M. J. 1993. Identification of chaperonin particles in mammalian brain cytosol and of T-complex polypeptide 1 as one of their components. J. Neurochem. 60:2327–2330.PubMedCrossRefGoogle Scholar
  3. 3.
    Burns, R. G., and Surridge, C. D. 1994. Functional role of a consensus peptide which is common to alpha-, beta-, and gammatubulin, to actin and centractin, to phytochrome A, and to the TCP1 alpha chaperonin protein. FEBS Lett. 347:105–111.PubMedCrossRefGoogle Scholar
  4. 4.
    Melki, R., Vainberg, I. E., Chow, R. L., and Cowan, N. J. 1993. Chaperonin-mediated folding of vertebrate actin-related protein and gamma-tubulin. J. Cell Biol. 122:1301–1310.PubMedCrossRefGoogle Scholar
  5. 5.
    Sternlicht, H., Farr, G. W., Sternlicht, M. L., Driscoll, J. K., Willison, K., and Yaffe, M. B. 1993. The t-complex polypeptide 1 complex is a chaperonin for tubulin and actin in vivo. Proc. Natl. Acad. Sci. 90:9422–9426.PubMedCrossRefGoogle Scholar
  6. 6.
    Yaffe, M. B., Farr, G. W., Miklos, D., Horwich, A. L., Sternlicht, M. L., and Sternlicht, H. 1992. TCP1 complex is a molecular chaperone in tubulin biogenesis. Nature. 358:245–248.PubMedCrossRefGoogle Scholar
  7. 7.
    Lewis, V. A., Hynes, G. M., Zheng, D., Saibil, H., and Willison, K. 1992. T-complex polypeptide-1 is a subunit of a heteromeric particle in the eukaryotic cytosol. Nature. 358:249–252.PubMedCrossRefGoogle Scholar
  8. 8.
    Beckmann, R. P., Mizzen, L. E., and Welch, W. J. 1990. Interaction of Hsp 70 with newly synthesized proteins: implications for protein folding and assembly. Science. 248:850–854.PubMedCrossRefGoogle Scholar
  9. 9.
    Beckmann, R. P., Lovett, M., and Welch, W. J. 1992. Examining the function and regulation of hsp 70 in cells subjected to metabolic stress. J. Cell Biol. 117:1137–1150.PubMedCrossRefGoogle Scholar
  10. 10.
    Brown, C. R., Martin, R. L., Hansen, W. J., Beckmann, R. P., and Welch, W. J. 1993. The constitutive and stress inducible forms of hsp 70 exhibit functional similarities and interact with one another in an ATP-dependent fashion. J. Cell Biol. 120:1101–1112.PubMedCrossRefGoogle Scholar
  11. 11.
    Langer, T., and Neupert, W. 1994. Chaperoning mitochondrial biogenesis. Pages 53–83,in Morimoto, R. I., Tissières, A., and Georgopoulos, C. (eds.), The Biology of Heat Shock Proteins and Molecular Chaperones, Cold Spring Harbor Laboratory Press, Plainview.Google Scholar
  12. 12.
    Craig, E. A., Gambill, B. D., and Nelson, R. J. 1993. Heat shock proteins: molecular chaperones of protein biogenesis. Microbiol. Rev. 57:402–414.PubMedGoogle Scholar
  13. 13.
    Aquino, D. A., Klipfel, A. A., Brosman, C. F., and Norton, W. T. 1993. The 70-kDa heat shock cognate protein (HSC70) is a major constituent of the central nervous system and is up-regulated only at the mRNA level in acute experimental autoimmune encephalomyelitis. J. Neurochem. 61:1340–1348.PubMedGoogle Scholar
  14. 14.
    Aquino, D. A., Tourtellotte, W. W., and Norton, W. T. 1993. Heat shock proteins in multiple sclerosis. J. Neurochem. 61 (Suppl): S228B.Google Scholar
  15. 15.
    Cammer, W., Tansey, F., Abramovitz, M., Ishigaki, S., and Listowsky, I. 1989. Differential localization of glutathione-S-transferase Yp and Yb subunits in oligodendrocytes and astrocytes of rat brain. J. Neurochem. 52:876–883.PubMedCrossRefGoogle Scholar
  16. 16.
    Norton, W. T., and Farooq, M. 1990. Bulk isolation and culture of oligodendroglia from mature brain. Pages 171–178,in Conn, P. M. (ed.), Methods in Neurosciences, Vol. 2, Academic Press, Inc., San Diego.Google Scholar
  17. 17.
    Wagner, R. W., Matteucci, M. D., Lewis, J. G., Gutierrez, A. J., Moulds, C., and Froehler, B. C. 1993. Antisense gene inhibition by oligonucleotides containing C-5 propyne pyrimidines. Science. 260:1510–1513.PubMedCrossRefGoogle Scholar
  18. 18.
    Sorger, P. K., and Pelham, H. R. 1987. Cloning and expression of a gene encoding hsc73, the major hsp70-like protein in unstressed rat cells. EMBO J. 6:993–998.PubMedGoogle Scholar
  19. 19.
    Norton, W. T., Farooq, M., Chiu, F.-C., and Bottenstein, J. E. 1988. Pure astrocyte cultures derived from cells isolated from mature brain. Glia. 1:403–414.PubMedCrossRefGoogle Scholar
  20. 20.
    Aquino, D. A., Chiu, F.-C., Brosnan, C. F., and Norton, W. T. 1988. Glial fibrillary acidic protein increases in the spinal cord of Lewis rats with acute experimental autoimmune encephalomyelitis. J. Neurochem. 51:1085–1096.PubMedCrossRefGoogle Scholar
  21. 21.
    Caceres, A., and Kosik, K. S. 1990. Inhibition of neurite polarity by tau antisense oligonucleotides in primary cerebellar neurons. Nature. 343:461–463.PubMedCrossRefGoogle Scholar
  22. 22.
    Selinfreund, R. H., Barger, S. W., Welsh, M. J., and Van Eldik, L. J. 1990. Antisense inhibition of glial S100 beta production results in alterations in cell morphology, cytoskeletal organization, and cell proliferation. J. Cell Biol. 111:2021–2028.PubMedCrossRefGoogle Scholar
  23. 23.
    Ulloa, L., Diaz-Nido, J., and Avila, J. 1993. Depletion of casein kinase II by antisense oligonucleotide prevents neuritogenesis in neuroblastoma cells. EMBO J. 12:1633–1640.PubMedGoogle Scholar
  24. 24.
    Yu, A. C. H., Lee, Y. L., and Eng, L. F. 1993. Astrogliosis in culture: I. The model and the effect of antisense oligonucleotides on glial fibrillary acidic protein synthesis. J. Neurosci. Res. 34: 295–303.PubMedCrossRefGoogle Scholar
  25. 25.
    Felgner, P. L., and Ringold, G. M. 1989. Cationic liposome-mediated transfection. Nature. 337:387–388.PubMedCrossRefGoogle Scholar
  26. 26.
    Langer, T., Lu, C., Echols, H., Flanagan, J., Hayer, M. K., and Hartl, F. U. 1992 Successive action of DnaK, DnaJ and GroEL along the pathway of chaperone-mediated protein folding. Nature. 356:683–689.PubMedCrossRefGoogle Scholar
  27. 27.
    Gething, M.-J., and Sambrook, J. 1992. Protein folding in the cell. Nature. 355:33–45.PubMedCrossRefGoogle Scholar
  28. 28.
    Chirico, W. J., Waters, M. G., and Blobel, G. 1988. 70K heat shock related proteins stimulate protein translocation into microsomes. Nature. 332:805–810.PubMedCrossRefGoogle Scholar
  29. 29.
    Deshaies, R. J., Koch, B. D., Werner-Washburne, M., Craig, E. A., and Schekman, R. 1988. A subfamily of stress proteins facilitates translocation of secretory and mitochondrial precursor polypeptides. Nature. 332:800–805.PubMedCrossRefGoogle Scholar
  30. 30.
    Normington, K., Kohno, K., Kozutsumi, Y., Gething, M.-J., and Sambrook, J. 1989. S. cerevisiae encodes an essential protein homologous in sequence and function to mammalian BiP. Cell. 57:1223–1236.PubMedCrossRefGoogle Scholar

Copyright information

© Plenum Publishing Corporation 1996

Authors and Affiliations

  • Dennis A. Aquino
    • 1
  • Carmen Lopez
    • 1
  • Muhammad Farooq
    • 1
  1. 1.Department of NeurologyAlbert Einstein College of MedicineBronx

Personalised recommendations