Abstract
High resolution two-dimensional polyacrylamide gel electrophoresis (2D-PAGE) and autoradiography were used to analyze the protein phenotypes ofTrypanosoma brucei (T.b. brucei andT.b. gambiense) clones suspected of being hybrids. Procyclic culture forms of parental and suspected hybrid trypanosomes were biosynthetically labeled with [35S]methionine and labeled proteins were resolved by multiple 2D-PAGE (the ISO-DALTTM system) to allow accurate inter-gel comparisons. Autoradiography of the gels showed that the parental clones had qualitative differences in at least seven sets of spots. Five of these sets represented charge differences and one represented proteins of altered relative molecular mass (Mr) and charge. Autoradiographs of the gels of the putative hybrid trypanosomes showed both forms of the proteins found separately in the parental clones indicating that new, nonparental phenotypes had been generated by transmission of mixed trypanosome clones through tsetse flies. The 2D-PAGE patterns from parasites cultivated for extended periods were identical, showing that the individual cloned parasites were phenotypically stable. The results indicate that analytical 2-D gels can be used to study the phenotypes of “parental” or “hybrid” African trypanosomes without having any previous knowledge of the molecular characteristics of the parasites. In addition, the technique allows an extension of phenotypic analysis to hundreds of different proteins in populations of cloned parasites.
Similar content being viewed by others
References
Anderson NG, Anderson NL (1978a) Analytical techniques for cell fractions. XXI. Two-dimensional analysis of serum and tissue proteins: multiple isoelectric focusing. Anal Biochem 85:331–340
Anderson NL, Anderson NG (1978b) Analytical techniques for cell fractions. XXII. Two-dimensional analysis of serum and tissue proteins: multiple gradient slab gel electrophoresis. Anal Biochem 85:341–354
Anderson NL, Anderson NG (1979) Microheterogeneity of serum transferrin, haptoglobin and α2HS glycoprotein examined by high resolution two-dimensional electrophoresis. Biochem Biophys Res Comm 88:258–265
Anderson NL, Parish NM, Richardson JP, Pearson TW (1985) Comparison of African trypanosomes of different antigenic phenotypes, subspecies and life cycle stages by two-dimensional gel electrophoresis. Mol Biochem Parasitol 16:299–314
Borst P, Fase-Fowler F, Frasch ACC, Hoeijmakers JHJ, Weijers PJ (1980) Characterization of DNA fromTrypanosoma brucei and related trypanosomes by restriction endonuclease digestion. Mol Biochem Parasitol 1:221–246
Borst P, Van der Ploeg M, Van Hoek JFM, Tas J, James J (1982) On the DNA content and ploidy of trypanosomes. Mol Biochem Parasitol 6:13–23
Brun R, Schönenberger M (1979) Cultivation andin vitro cloning of procyclic culture forms ofTrypanosoma brucei in a semi-defined medium. Acta Trop 36:289–292
Brun R, Schönenberger M (1981) Stimulating effect of citrate and cis-aconitate on the transformation ofTrypanosoma brucei bloodstream forms to procyclic formsin vitro. Z Parasitenkd 66:17–24
Cunningham I (1973) New culture medium for maintenance of tsetse tissues and growth of trypanosomatids. J Protozool 24:325–329
Felgner P, Brinkmann U, Zillmann U, Mehlitz D, Abu-Ishira S (1981) Epidemiological studies on the animal reservoir of gambiense sleeping sickness, Part II. Parasitological and immunodiagnostic examination of the human population. Tropenmed Parasitol 32:134–140
Geigy R, Kaufmann M (1973) Sleeping sickness survey in the Serengeti area (Tanzania). Examination of large animals for trypanosomes. Acta Trop 30:12–23
Gibson WC, Marshall TF de C, Godfrey DG (1980) Numerical analysis of enzyme polymorphism: a new approach to the epidemiology and taxonomy of the subgenusTrypanozoon. Adv Parasitol 18:175–246
Gibson WC, Osinga KA, Michels PAM, Borst P (1985) Trypanosomes of subgenus Trypanozoon are diploid for houskeeping genes. Mol Biochem Parasitol 16:231–242
Jenni L, Marti S, Schweizer J, Betschart B, LePage RWF, Wells JM, Tait A, Paindavoine P, Pays E, Steinert M (1986) Hybrid formation between African trypanosomes during cyclical transmission. Nature 322:173–175
Paindavoine P, Zampetti-Bosseler R, Pays E, Schweizer J, Guyaux M, Jenni L, Steinert M (1986) Trypanosome hybrids generated in tsetse flies by nuclear fusion. EMBO J 5:3631–3636
Pearson TW, Anderson NL (1983) Use of high-resolution two-dimensional gel electrophoresis for analysis of monoclonal antibodies and their specific antigens. Meth Enzymol 92:196–220
Sternberg J, Tait A, Haley S, Wells JM, Prospero TD, LePage RWF, Schweizer J, Jenni L (1988) Gene exchange in African trypanosomes: characterization of a new hybrid genotype. Mol Biochem Parasitol 27:191–200
Tait A (1980) Evidence for diploidy and mating in trypanosomes. Nature 287:536–538
Tait A (1983) Sexual processes in the Kinetoplastida. Parasitology 86:29–57
Van Meirvenne N, Janssens PG, Magnus E (1975) Antigenic variation in syringe passaged populations ofTrypanosoma (Trypanozoon) brucei. Ann Soc Belge Med Trop 55:1–23
Vickerman K (1985) Developmental cycles and biology of pathogenic trypanosomes. Br Med Bull 41:105–114
Wells JM, Prospero TD, Jenni L, LePage RWF (1987) DNA contents and molecular karyotypes of hybridTrypanosoma brucei. Mol Biochem Parasitol 249:103–116
Author information
Authors and Affiliations
Rights and permissions
About this article
Cite this article
Pearson, T.W., Jenni, L. Detection of hybrid phenotypes in African trypanosomes by high resolution two-dimensional gel electrophoresis. Parasitol Res 76, 63–67 (1989). https://doi.org/10.1007/BF00931074
Accepted:
Issue Date:
DOI: https://doi.org/10.1007/BF00931074