Skip to main content
Genetics logoLink to Genetics
. 1992 Mar;130(3):461–470. doi: 10.1093/genetics/130.3.461

Interactions between Genes Involved in Exocytotic Membrane Fusion in Paramecium

H Bonnemain 1, T Gulik-Krzywicki 1, C Grandchamp 1, J Cohen 1
PMCID: PMC1204865  PMID: 1551571

Abstract

Crosses between members of two independent collections of Paramecium tetraurelia mutants blocked in the final membrane fusion step of trichocyst release (nd mutants) allowed us to define 13 complementation groups comprising 23 alleles. The mutant nd9(a) was then used as a target in a mutagenesis experiment designed to screen both revertants and new mutants in order to identify interacting genes. This mutant was chosen because it is the best known of its class to date and seems to be altered in assembly of the material connecting the trichocyst membrane to the plasma membrane and in assembly of the ``rosette,'' a complex array of intramembranous particles in the plasma membrane at the trichocyst insertion sites. No revertants were obtained but two new mutants deficient for rosette assembly were identified, nd16(b) and nd18, whose gene products appear to interact with that of nd9. Indeed, the double mutants grown at 18°, a permissive temperature for each of the single mutants, are characterized by a deficiency in exocytosis and in rosette assembly, as are also double mutants combining other allelic forms of the same genes. Moreover, aberrant dominance relationships among alleles of nd9 and of nd16 indicate the existence of interactions between identical subunits, which most likely assemble into multimeric structures. The nd16 gene product was shown by microinjection experiments to be a cytosolic factor, as in the nd9 gene product. It is therefore tempting to propose that the nd16 gene product also belongs to the connecting material and is involved in rosette assembly, in cooperation with nd9 and nd18.

Full Text

The Full Text of this article is available as a PDF (3.1 MB).

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Aggerbeck L. P., Gulik-Krzywicki T. Studies of lipoproteins by freeze-fracture and etching electron microscopy. Methods Enzymol. 1986;128:457–472. doi: 10.1016/0076-6879(86)28087-8. [DOI] [PubMed] [Google Scholar]
  2. Aunis D., Hesketh J. E., Devilliers G. Freeze-fracture study of the chromaffin cell during exocytosis: evidence for connections between the plasma membrane and secretory granules and for movements of plasma membrane-associated particles. Cell Tissue Res. 1979 Apr 12;197(3):433–441. doi: 10.1007/BF00233568. [DOI] [PubMed] [Google Scholar]
  3. Beisson J., Cohen J., Lefort-Tran M., Pouphile M., Rossignol M. Control of membrane fusion in exocytosis. Physiological studies on a Paramecium mutant blocked in the final step of the trichocyst extrusion process. J Cell Biol. 1980 May;85(2):213–227. doi: 10.1083/jcb.85.2.213. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Beisson J., Lefort-Tran M., Pouphile M., Rossignol M., Satir B. Genetic analysis of membrane differentiation in Paramecium. Freeze-fracture study of the trichocyst cycle in wild-type and mutant strains. J Cell Biol. 1976 Apr;69(1):126–143. doi: 10.1083/jcb.69.1.126. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Burgoyne R. D. Calpactin in exocytosis. Nature. 1988 Jan 7;331(6151):20–20. doi: 10.1038/331020a0. [DOI] [PubMed] [Google Scholar]
  6. Cohen J., Beisson J. Genetic analysis of the relationships between the cell surface and the nuclei in Paramecium tetraurella. Genetics. 1980 Aug;95(4):797–818. doi: 10.1093/genetics/95.4.797. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Cohen J. Cytotoxic versus mutagenic effect of ethyl methanesulfonate on Paramecium tetraurelia. Mutat Res. 1980 Apr;70(2):251–254. doi: 10.1016/0027-5107(80)90165-7. [DOI] [PubMed] [Google Scholar]
  8. Gilligan D. M., Satir B. H. Protein phosphorylation/dephosphorylation and stimulus-secretion coupling in wild type and mutant Paramecium. J Biol Chem. 1982 Dec 10;257(23):13903–13906. [PubMed] [Google Scholar]
  9. Gilligan D. M., Satir B. H. Stimulation and inhibition of secretion in Paramecium: role of divalent cations. J Cell Biol. 1983 Jul;97(1):224–234. doi: 10.1083/jcb.97.1.224. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Gomperts B. D. GE: a GTP-binding protein mediating exocytosis. Annu Rev Physiol. 1990;52:591–606. doi: 10.1146/annurev.ph.52.030190.003111. [DOI] [PubMed] [Google Scholar]
  11. Goud B., Salminen A., Walworth N. C., Novick P. J. A GTP-binding protein required for secretion rapidly associates with secretory vesicles and the plasma membrane in yeast. Cell. 1988 Jun 3;53(5):753–768. doi: 10.1016/0092-8674(88)90093-1. [DOI] [PubMed] [Google Scholar]
  12. Kerboeuf D., Cohen J. A Ca2+ influx associated with exocytosis is specifically abolished in a Paramecium exocytotic mutant. J Cell Biol. 1990 Dec;111(6 Pt 1):2527–2535. doi: 10.1083/jcb.111.6.2527. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Koffer A., Gomperts B. D. Soluble proteins as modulators of the exocytotic reaction of permeabilised rat mast cells. J Cell Sci. 1989 Nov;94(Pt 3):585–591. doi: 10.1242/jcs.94.3.585. [DOI] [PubMed] [Google Scholar]
  14. Lawson D., Raff M. C., Gomperts B., Fewtrell C., Gilula N. B. Molecular events during membrane fusion. A study of exocytosis in rat peritoneal mast cells. J Cell Biol. 1977 Feb;72(2):242–259. doi: 10.1083/jcb.72.2.242. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Lumpert C. J., Kersken H., Plattner H. Cell surface complexes ('cortices') isolated from Paramecium tetraurelia cells as a model system for analysing exocytosis in vitro in conjunction with microinjection studies. Biochem J. 1990 Aug 1;269(3):639–645. doi: 10.1042/bj2690639. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Matt H., Bilinski M., Plattner H. Adenosinetriphosphate, calcium and temperature requirements for the final steps of exocytosis in Paramecium cells. J Cell Sci. 1978 Aug;32:67–86. doi: 10.1242/jcs.32.1.67. [DOI] [PubMed] [Google Scholar]
  17. Nadin C. Y., Rogers J., Tomlinson S., Edwardson J. M. A specific interaction in vitro between pancreatic zymogen granules and plasma membranes: stimulation by G-protein activators but not by Ca2+. J Cell Biol. 1989 Dec;109(6 Pt 1):2801–2808. doi: 10.1083/jcb.109.6.2801. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Nakata T., Sobue K., Hirokawa N. Conformational change and localization of calpactin I complex involved in exocytosis as revealed by quick-freeze, deep-etch electron microscopy and immunocytochemistry. J Cell Biol. 1990 Jan;110(1):13–25. doi: 10.1083/jcb.110.1.13. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Novick P., Field C., Schekman R. Identification of 23 complementation groups required for post-translational events in the yeast secretory pathway. Cell. 1980 Aug;21(1):205–215. doi: 10.1016/0092-8674(80)90128-2. [DOI] [PubMed] [Google Scholar]
  20. Orci L., Perrelet A., Friend D. S. Freeze-fracture of membrane fusions during exocytosis in pancreatic B-cells. J Cell Biol. 1977 Oct;75(1):23–30. doi: 10.1083/jcb.75.1.23. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Orias E., Flacks M., Satir B. H. Isolation and ultrastructural characterization of secretory mutants of Tetrahymena thermophila. J Cell Sci. 1983 Nov;64:49–67. doi: 10.1242/jcs.64.1.49. [DOI] [PubMed] [Google Scholar]
  22. Palade G. Intracellular aspects of the process of protein synthesis. Science. 1975 Aug 1;189(4200):347–358. doi: 10.1126/science.1096303. [DOI] [PubMed] [Google Scholar]
  23. Pinto da Silva P., Nogueira M. L. Membrane fusion during secretion. A hypothesis based on electron microscope observation of Phytophthora Palmivora zoospores during encystment. J Cell Biol. 1977 Apr;73(1):161–181. doi: 10.1083/jcb.73.1.161. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Plattner H., Miller F., Bachmann L. Membrane specializations in the form of regular membrane-to-membrane attachment sites in Paramecium. A correlated freeze-etching and ultrathin-sectioning analysis. J Cell Sci. 1973 Nov;13(3):687–719. doi: 10.1242/jcs.13.3.687. [DOI] [PubMed] [Google Scholar]
  25. Plattner H. Regulation of membrane fusion during exocytosis. Int Rev Cytol. 1989;119:197–286. doi: 10.1016/s0074-7696(08)60652-x. [DOI] [PubMed] [Google Scholar]
  26. Plattner H., Reichel K., Matt H., Beisson J., Lefort-Tran M., Pouphile M. Genetic dissection of the final exocytosis steps in Paramecium tetraurelia cells: cytochemical determination of Ca2+-ATPase activity over performed exocytosis sites. J Cell Sci. 1980 Dec;46:17–40. doi: 10.1242/jcs.46.1.17. [DOI] [PubMed] [Google Scholar]
  27. Pollack S. Mutations affecting the trichocysts in Paramecium aurelia. I. Morphology and description of the mutants. J Protozool. 1974 May;21(2):352–362. doi: 10.1111/j.1550-7408.1974.tb03669.x. [DOI] [PubMed] [Google Scholar]
  28. Ruiz F., Adoutte A., Rossignol M., Beisson J. Genetic analysis of morphogenetic processes in Paramecium. I. A mutation affecting trichocyst formation and nuclear division. Genet Res. 1976 Apr;27(2):109–122. doi: 10.1017/s0016672300016323. [DOI] [PubMed] [Google Scholar]
  29. Schekman R. Protein localization and membrane traffic in yeast. Annu Rev Cell Biol. 1985;1:115–143. doi: 10.1146/annurev.cb.01.110185.000555. [DOI] [PubMed] [Google Scholar]
  30. Tanaka Y., De Camilli P., Meldolesi J. Membrane interactions between secretion granules and plasmalemma in three exocrine glands. J Cell Biol. 1980 Feb;84(2):438–453. doi: 10.1083/jcb.84.2.438. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Turkewitz A. P., Madeddu L., Kelly R. B. Maturation of dense core granules in wild type and mutant Tetrahymena thermophila. EMBO J. 1991 Aug;10(8):1979–1987. doi: 10.1002/j.1460-2075.1991.tb07727.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Vilmart J., Plattner H. Membrane-integrated proteins at preformed exocytosis sites. J Histochem Cytochem. 1983 May;31(5):626–632. doi: 10.1177/31.5.6841968. [DOI] [PubMed] [Google Scholar]
  33. Zieseniss E., Plattner H. Synchronous exocytosis in Paramecium cells involves very rapid (less than or equal to 1 s), reversible dephosphorylation of a 65-kD phosphoprotein in exocytosis-competent strains. J Cell Biol. 1985 Dec;101(6):2028–2035. doi: 10.1083/jcb.101.6.2028. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. de Curtis I., Simons K. Isolation of exocytic carrier vesicles from BHK cells. Cell. 1989 Aug 25;58(4):719–727. doi: 10.1016/0092-8674(89)90106-2. [DOI] [PubMed] [Google Scholar]

Articles from Genetics are provided here courtesy of Oxford University Press

RESOURCES