Protoplasma

, Volume 168, Issue 3–4, pp 113–124

Biochemical and immunological characterization of intermicrotubular cement in the feeding apparatus of phagotrophic eugienoids:Entosiphon, Peranema, andPloeotia

  • A. Belhadri
  • Danielle Bayle
  • G. Brugerolle
Article

Summary

A monoclonal antibody (IIID12) obtained from mice immunized against the entireEntosiphon cytoskeleton highlights the feeding apparatus ofEntosiphon, Peranema, andPloeotia by IF. IGS at the ultrastructural level shows that it labels the cementing material surrounding the microtubular bundles in the three species studied. InEntosiphon additional structures, such as the supplementary plaque, the scaffold structure and the lenticular structure or canal thickening, are also detected by the antibody. Immunoblotting analysis after SDS-PAGE reveals a positive reaction with this antibody to the 58 and 66kDa protein bands inEntosiphon, 82 and 84kDa inPeranema, and 56 and 60kDa inPloeotia. These results demonstrate biochemical homologies in the proteins of the cement material in the three heteronematal eugienoids studied. The possible role of these proteins in microtubule assembly and stabilization is discussed, as well as the role of the cementing material in the mode of the feeding apparatus motion during the ingestion of food.

Keywords

Cytoskeleton Phagotrophic Eugienoids Immunochemistry Ultrastructure 

Abbreviations

BSA

bovine serum albumin

DMSO

dimethyl sulfoxide

EDTA

ethylenediaminetetraacetic acid

IF

immunofluorescence

IGS

immunogold staining

HAT

hypoxantin-aminopterinthymidine

MAb

monoclonal antibody

MEM

minimum essential medium

PBS

phosphate buffered saline

PMSF

phenylmethanesulfonyl fluoride

TAME

Nα-p-tosyl-L-arginine methyl-ester

TLCK

Nα-p-tosyl-L-lysine chloromethyl ketone

SDS-PAGE

sodium dodecyl sulfate-polyacrylamide gel electrophoresis

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References

  1. Bricheux G, Brugerolle G (1986) The membrane cytoskeleton complex of euglenoids. I. Biochemical and immunological characterization of the epiplasmic proteins ofEuglena acus. Eur J Cell Biol 40: 150–159Google Scholar
  2. — — (1987) The pellicular complex of euglenoids. II. Biochemical and immunological comparative study of major epiplasmic proteins. Protoplasma 140: 43–54Google Scholar
  3. Dubreuil RR, Bouck GB (1985) The membrane skeleton of a unicellular organism consists of bridged, articulating strips. J Cell Biol 101: 1884–1896Google Scholar
  4. — — (1988) Interrelationships among the plasma membrane, the membrane skeleton and surface form in a unicellular flagellate. Protoplasma 143: 150–164Google Scholar
  5. —, Rosiere TK, Rosner MC, Bouck GB (1988) Properties and topography of the major integral plasma membrane protein of a unicellular organism. J Cell Biol 107: 191–200Google Scholar
  6. Farmer MA, Triemer RE (1988) A redescription of the genusPloeotia Duj. (Euglenophyceae). Taxon 37: 319–325Google Scholar
  7. Galfré G, Milstein C (1981) Preparation of monoclonal antibodies. Strategies and procedure. Methods Enzymol 73: 3–47Google Scholar
  8. Gallo JM, Schrével J (1985) Homologies between paraflagellar rod proteins from trypanosomes and euglenoids revealed by a monoclonal antibody. Eur J Cell Biol 36: 163–168Google Scholar
  9. —, Karsenti E, Bornens M, Delacourte A, Schrével J (1982) Euglenoid movement inDistigma proteus. II. Presence and localization of an actin-like protein. Biol Cell 44: 149–156Google Scholar
  10. Hilenski LL, Walne PL (1985) Ultrastructure of the flagella of the colorless phagotrophPeranema trichophorum (Euglenophyceae). II. Flagellar roots. J Phycol 21: 125–134Google Scholar
  11. Hoffmann C, Bouck GB (1976) Immunological and structural evidence for patterned intussusceptive surface growth in a unicellular organism. A postulated role for submembranous proteins and microtubules. J Cell Biol 69: 693–715Google Scholar
  12. Hyams JS (1982) TheEuglena paraflagellar rod: structure, relationship to other flagellar components and preliminary biochemical characterization. J Cell Sci 55: 199–210Google Scholar
  13. Kivic PA, Walne PL (1984) An evaluation of a possible phylogenetic relationship between the Euglenophyta and the Kinetoplastida. Origins Life 13: 269–288Google Scholar
  14. Laemmli UK (1970) Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature 227: 680–685Google Scholar
  15. Larsen J, Patterson DJ (1990) Some flagellates (Protista) from tropical marine sediments. J Nat Hist 24: 801–937Google Scholar
  16. Lefort-Tran M, Bré MH, Ranck JL, Pouphile M (1980)Euglena plasma membrane during normal and vitamin B12 starvation growth. J Cell Sci 41: 245–261Google Scholar
  17. Leedale GF (1967) Euglenoid flagellates. Prentice Hall, Englewood Cliffs, NJGoogle Scholar
  18. — (1978) Phylogenetic criteria in euglenoid flagellates. BioSystems 10: 183–187Google Scholar
  19. Lonergan TA (1985) Regulation of cell shape inEuglena gracilis. IV. Localization of actin, myosin and calmodulin. J Cell Sci 77: 197–208Google Scholar
  20. Levine ND, Corliss JO, Cox FEG, Deroux G, Grain J, Honigberg BM, Leedale GF, Loeblich AR, Lom J, Lynn D, Merinfeld EG, Page FC, Poljansky G, Sprague V, Vavra J, Wallace FG (1980) A newly revised classification of the protozoa. J Protozool 27: 37–58Google Scholar
  21. Mignot JP (1963) Quelques particularités de l'ultrastructure d'Entosiphon sulcatum (Duj.) Stein, flagellé euglénien. CR Acad Sci Paris 257: 2530–2533Google Scholar
  22. — (1966) Structure et ultrastructure de quelques euglénomonadines. Protistologica 2: 51–117Google Scholar
  23. Miller KR, Miller GJ (1978) Organization of the cell membrane inEuglena. Protoplasma 95: 11–24Google Scholar
  24. Nisbet B (1974) An ultrastructural study of the feeding apparatus ofPeranema trichophorum. J Protozool 21: 39–48Google Scholar
  25. Nahrebne DK, Triemer RE (1990) Proteins involved in movement of the microtubular components of the feeding apparatus inEntosiphon sulcatum. J Protozool 37: abstract 3A-18Google Scholar
  26. O'Farrel PH (1975) High resolution two dimensional electrophoresis of proteins. J Biol Chem 250: 4007–4021Google Scholar
  27. Rosiere TK, Marrs JA, Bouck GB (1990) A 39-kD plasma membrane protein (IP 39) is an anchor for the unusual membrane skeleton ofEuglena gracilis. J Cell Biol 110: 1077–1088Google Scholar
  28. Sogin ML (1989) Evolution of eukaryotic microorganisms and their small subunit ribosomal RNAs. Amer Zool 29: 487–499Google Scholar
  29. —, Gunderson JH (1987) Structural diversity of eukaryotic small subunit ribosomal RNAs. Ann N Y Acad Sci 503: 125–139Google Scholar
  30. Surek B, Melkonian M (1986) A cryptic cytostome is present inEuglena. Protoplasma 133: 39–49Google Scholar
  31. Suzaki T, Williamson RE (1986) Pellicular ultrastructure and euglenoid movement inEuglena ehrenbergii Klebs andEuglena oxyuris Schmarda. J Protozool 33: 165–184Google Scholar
  32. Towbin H, Staehelin T, Gordon J (1979) Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets: procedure and some applications. Proc Natl Acad Sci USA 76: 4350–4354Google Scholar
  33. Triemer RE (1986) Light and electron microscopic description of a colorless euglenoid,Serpenomonas costata n.g., n.sp. J Protozool 33: 412–415Google Scholar
  34. —, Fanner MA (1991 a) An ultrastructural comparison of the mitotic apparatus, feeding apparatus, flagellar apparatus and cytoskeleton in euglenoids and kinetoplastids. Protoplasma 164: 91–104Google Scholar
  35. — — (1991 b) The ultrastructural organization of the heterotrophic euglenoids and its evolutionary implications. In: Patterson DJ, Larsen J (eds) The biology of free-living heterotrophic flagellates. Clarendon Press, Oxford, pp 185–203 (The Systematic Association, vol 45)Google Scholar
  36. —, Fritz L (1987) Structure and operation of the feeding apparatus on a colorless euglenoid.Entosiphon sulcatum. J Protozool 34: 39–47Google Scholar
  37. Viguès B, Bricheux G, Metivier C, Brugerolle G, Peck RK (1987) Evidence for common epitopes among proteins of the membrane skeleton of a ciliate, an euglenoid and a dinoflagellate. Eur J Protistol 23: 101–110Google Scholar
  38. Willey RL, Walne PL, Kivic K (1988) Phagotrophy and the origins of the euglenoid flagellates. CRC Crit Rev Plant Sci 7: 303–340Google Scholar

Copyright information

© Springer-Verlag 1992

Authors and Affiliations

  • A. Belhadri
    • 1
  • Danielle Bayle
    • 1
  • G. Brugerolle
    • 1
  1. 1.Laboratoire de Zoologie-ProtistologieUniversité Blaise PascalAubièreFrance

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