Skip to main content
NCBI home page
Cellular and Molecular Neurobiology logoLink to Cellular and Molecular Neurobiology
. 1992 Oct;12(5):397–409. doi: 10.1007/BF00711541

Measles virus-substance P receptor interactions: Jurkat lymphocytes transfected with substance P receptor cDNA enhance measles virus fusion and replication

Greg Harrowe 1,2, Julie Sudduth-Klinger 1,2, Donald G Payan 1,2,3,✉
PMCID: PMC11567401  PMID: 1281755

Abstract

  1. We have demonstrated previously (Harroweet al., 1990), using a lymphoblastoid cell line that constitutively expresses the substance P receptor (SPR) (Payanet al., 1984, 1986), that this receptor may facilitate measles virus (MV) fusion with these cells. In order to test this hypothesis further, a stable cell line transfected with SPR cDNA has been established, and various stages of MV infection in SPR positive and negative cells compared.

  2. Jurkat cells, a human T-lymphoblastoid cell line, were transfected with a cDNA clone encoding the SPR. Cells transfected with only the plasmid were used as controls. Jurkat cells and Jurkat vector control cells (J-vo) failed to demonstrate any detectable125I-SP binding, whereas a clonally selected population of cells transfected with SPR cDNA (J-SPR) expressed about 50,000 receptors/cell (Sudduth-Klingeret al., 1992).

  3. Using the J-vo- and J-SPR-transfected cell lines, the following experiments were conducted to investigate the effect of SPR expression on MV infection. To determine if MV would preferentially attach to J-SPR as compared to J-vo, we absorbed virus to cells at 37°C for various times and measured bound MV using a fluorescence activated cell sorter (FACS). Using this approach, we found that MV bound to a greater degree to J-SPR compared with J-vo. In addition to equilibrium being reached faster for J-SPR, the total amount of bound MV was higher on J-SPR. The effect was greater at lower MOIs, suggesting that there existed multiple binding sites for MV on these cells and that the affinity is higher for those cells expressing the SPR.

  4. Since binding does not necessitate a successful viral infection, we needed to know if this difference in binding reflected a difference in infection. This was demonstrated by showing an approximate twofold increase in infected cells after a 2-hr binding period with J-SPR as compared to J-vo at an MOI of 1 in an infectious cell-center assay. Moreover, when both cells types were subjected to continuous infection in culture, J-SPR-infected cells produced a seven- to ninefold increase in measles viral titer in 24 hr as compared with J-vo. The observed increase in viral titer may have resulted in more of the J-SPR cells binding virus, as indicated by our binding and infectious cell-center data, or alternatively, the virus might have entered the J-SPR cells faster and begun replication before the J-vo-infected cells.

  5. The possibility of a difference in the rate of fusion was investigated using the octadecyl rhodamine B (R18) dequenching assay. This assay utilizes the ability of R18 to quench its own fluorescence at high concentrations in the viral membrane (Blumenthalet al., 1987). Upon fusion of R18-loaded virus with cell membrane, a release of quenching occurs as the dye dilutes itself in the cell's plasma membrane, causing an increase in fluorescence. This allows for a kinetic measurement of viral fusion. Employing this technique, we looked at the fusion kinetics of R18 MV absorbed to J-SPR and J-vo. After a 2-h preincubation with cells and R18-loaded virus at 4°C, the virally bound cells were warmed to 37°C in a fluorimeter and fluorescence was monitored. The results demonstrated a rapid increase in fusion with J-SPR relative to J-vo. In addition, the level of fluorescence was higher for J-SPR compared with J-vo. These data suggest that fusion was enhanced by the SPR, and per unit time, more virus was bound and fusing.

  6. SPR expression in Jurkat cells facilitates and enhances MV infection. The molecular mechanisms by which this facilitation takes place are an area of ongoing investigation in this laboratory.

Key words: substance P, substance P receptor, measles virus, neuroimmunology, viral fusion, viral replications

References

  1. Blumenthal, R., Bali-Puri, A., Walter, A., Covell, D., and Eidelman, O. (1987). pH-dependent fusion of vesicular stomatitis virus with Vero cells.J. Biol. Chem.26213614–13619. [PubMed] [Google Scholar]
  2. Desai, D. M., Goldsmith, M., and Weiss, A. (1990a). A transfected human muscarinic receptor fails to substitute for the T cell antigen receptor complex in CD2-initiated signal transduction.Int. Immunol.2615–620. [DOI] [PubMed] [Google Scholar]
  3. Desai, D. M., Newton, M. E., Kadlecek, T., and Weiss, A. (1990b). Stimulation of the phosphatidyl-inositol pathway can induce T-cell activation.Nature34866–69. [DOI] [PubMed] [Google Scholar]
  4. Goldsmith, M. A., Desai, D. M., Schultz, T., and Weiss, A. (1989). Function of a heterologous muscarinic receptor in T cell antigen receptor signal transduction mutants.J. Biol. Chem.26417190–17197. [PubMed] [Google Scholar]
  5. Harrowe, G., Mitsuhashi, M., and Payan, D. G. (1990). Measles virus-substance P receptor interactions. Possible novel mechanism of viral fusion.J. Clin. Invest.851324–1327. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Horth, M., Lambrecht, B., Khim, M. E. L., Bex, F., Thiriat, C., Rysschaert, J. M., Buray, A., and Brasseur, R. (1991). Theoretical and functional analysis of the SIV fusion peptide.EMBO102747–2755. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Krah, D. L., and Choppin, P. W. (1988). Mice immunized with measles virus develop antibodies to a cell surface receptor for binding virus.J. Virol.621565–1572. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Lentz, T. L., Burrage, T. G., Smith, A. L., Crick, J., and Tignor, G. H. (1982). Is the acetylcholine receptor a rabies receptor?Science215182–184. [DOI] [PubMed] [Google Scholar]
  9. McGillis, J. P., Organist, M. L., and Payan, D. G. (1987). Immunoaffinity purification of membrane protein constituents of the IM-9 lymphoblast receptor for substance P.Anal. Biochem.164502–513. [DOI] [PubMed] [Google Scholar]
  10. Mitsuhashi, M., Mitsuhashi, T., and Payan, D. G. (1989). Multiple signaling pathways of histamine H2 receptors.J. Biol. Chem.26418356–18362. [DOI] [PubMed] [Google Scholar]
  11. Mitsuhashi, M., Shichijo, S., Christian, C., Sudduth-Klinger, J., Harrowe, G., and Payan, D. G. (1992). Multiple intracellular signaling pathways of the neuropeptide substance P receptor.J. Neurosci. Res. (in press). [DOI] [PubMed]
  12. Payan, D. G., Brewster, D. R., and Goetzl, E. J. (1984). Stereospecific receptors for substance P on cultured human IM-9 lymphoblasts.J. Immunol.1333260–3265. [PubMed] [Google Scholar]
  13. Payan, D. G., McGillis, J. P., and Organist, M. L. (1986). Binding characteristics and affinity labeling of protein constituents of the human IM-9 lymphoblast receptor for substance P.J. Biol. Chem.26114321–14329. [PubMed] [Google Scholar]
  14. Richardson, C. D., and Choppin, P. W. (1983). Oligopeptides that specifically inhibit membrane fusion by paramyxoviruses.Virology131518–532. [DOI] [PubMed] [Google Scholar]
  15. Richardson, C. D., Scheid, A., and Choppin, P. W. (1980). Specific inhibition of paramyxovirus replication by oligopeptides with amino acid sequences similar to those at the N-termini of the F1 or HA2 viral polypeptides.Virology105205–222. [DOI] [PubMed] [Google Scholar]
  16. Sambrook, J., Fritsch, E. F., and Maniatis, T. (1989).Molecular Cloning: A Laboratory Manual, 2nd ed., Cold Spring Harbor Laboratory Press, Cold Spring Harbor, New York. [Google Scholar]
  17. Scheid, A., and Choppin, P. W. (1977). Two disulfide-linked polypeptide chains constitute the active F protein of paramyxoviruses.Virology8054–66. [DOI] [PubMed] [Google Scholar]
  18. Schroeder, C. (1986). Substance P, a neuropeptide, inhibits measles virus replication in cell culture.Acta Virol.30432–435. [PubMed] [Google Scholar]
  19. Slepushkin, V. A., Melikyan, G. B., Sidorova, M. S., Chumakov, V. M., Andrew, S. M. Manulyan, R. A., and Karamov, E. V. (1990). Interaction of human immunodeficiency virus (HIV-1) fusion peptides with artifical lipid membranes.Biochem. Biophys. Res. Commun.172952–957. [DOI] [PubMed] [Google Scholar]
  20. Sudduth-Klinger, J., Schumann, M., Gardner, P., and Payan, D. G. (1992). Functional and immunological responses of Jurkat lymphocytes transfected with the substance P receptor.Cell. Mol. Neurobiol.12379–395. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Vile, R. G., and Weiss, R. A. (1991). Virus receptors as permeases.Nature352666. [DOI] [PubMed] [Google Scholar]
  22. White, J., and Littman, D. R. (1989). Viral receptors of the immunoglobulin superfamily.Cell56725. [DOI] [PubMed] [Google Scholar]
  23. Yakota, Y., Sasai, Y., Tanaka, K., Fujiwara, T., Tsuchida, K., Shigemoto, R., Kakizuka, A., Ohkubo, H., Nakanishi, S. (1989). Molecular characterization of a functional cDNA for rat substance P receptor.J. Biol. Chem.26417649–17652. [PubMed] [Google Scholar]

Articles from Cellular and Molecular Neurobiology are provided here courtesy of Springer

RESOURCES