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. 1997 Feb;17(2):833–843. doi: 10.1128/mcb.17.2.833

Gene conversions mediating antigenic variation in Trypanosoma brucei can occur in variant surface glycoprotein expression sites lacking 70-base-pair repeat sequences.

R McCulloch 1, G Rudenko 1, P Borst 1
PMCID: PMC231809  PMID: 9001237

Abstract

African trypanosomes undergo antigenic variation of their variant surface glycoprotein (VSG) coat to avoid immune system-mediated killing by their mammalian host. An important mechanism for switching the expressed VSG gene is the duplicative transposition of a silent VSG gene into one of the telomeric VSG expression sites of the trypanosome, resulting in the replacement of the previously expressed VSG gene. This process appears to be a gene conversion reaction, and it has been postulated that sequences within the expression site may act to initiate and direct the reaction. All bloodstream form expression sites contain huge arrays (many kilobase pairs) of 70-bp repeat sequences that act as the 5' boundary of gene conversion reactions involving most silent VSG genes. For this reason, the 70-bp repeats seemed a likely candidate to be involved in the initiation of switching. Here, we show that deletion of the 70-bp repeats from the active expression site does not affect duplicative transposition of VSG genes from silent expression sites. We conclude that the 70-bp repeats do not appear to function as indispensable initiation sites for duplicative transposition and are unlikely to be the recognition sequence for a sequence-specific enzyme which initiates recombination-based VSG switching.

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Selected References

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

  1. Aline R., Jr, MacDonald G., Brown E., Allison J., Myler P., Rothwell V., Stuart K. (TAA)n within sequences flanking several intrachromosomal variant surface glycoprotein genes in Trypanosoma brucei. Nucleic Acids Res. 1985 May 10;13(9):3161–3177. doi: 10.1093/nar/13.9.3161. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Bernards A., De Lange T., Michels P. A., Liu A. Y., Huisman M. J., Borst P. Two modes of activation of a single surface antigen gene of Trypanosoma brucei. Cell. 1984 Jan;36(1):163–170. doi: 10.1016/0092-8674(84)90085-0. [DOI] [PubMed] [Google Scholar]
  3. Bernards A., Kooter J. M., Borst P. Structure and transcription of a telomeric surface antigen gene of Trypanosoma brucei. Mol Cell Biol. 1985 Mar;5(3):545–553. doi: 10.1128/mcb.5.3.545. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Bernards A., Van der Ploeg L. H., Frasch A. C., Borst P., Boothroyd J. C., Coleman S., Cross G. A. Activation of trypanosome surface glycoprotein genes involves a duplication-transposition leading to an altered 3' end. Cell. 1981 Dec;27(3 Pt 2):497–505. doi: 10.1016/0092-8674(81)90391-3. [DOI] [PubMed] [Google Scholar]
  5. Blundell P. A., Rudenko G., Borst P. Targeting of exogenous DNA into Trypanosoma brucei requires a high degree of homology between donor and target DNA. Mol Biochem Parasitol. 1996 Feb-Mar;76(1-2):215–229. doi: 10.1016/0166-6851(95)02560-x. [DOI] [PubMed] [Google Scholar]
  6. Borst P., Bitter W., McCulloch R., Van Leeuwen F., Rudenko G. Antigenic variation in malaria. Cell. 1995 Jul 14;82(1):1–4. doi: 10.1016/0092-8674(95)90044-6. [DOI] [PubMed] [Google Scholar]
  7. Borst P., Cross G. A. Molecular basis for trypanosome antigenic variation. Cell. 1982 Jun;29(2):291–303. doi: 10.1016/0092-8674(82)90146-5. [DOI] [PubMed] [Google Scholar]
  8. Borst P., Greaves D. R. Programmed gene rearrangements altering gene expression. Science. 1987 Feb 6;235(4789):658–667. doi: 10.1126/science.3544215. [DOI] [PubMed] [Google Scholar]
  9. Borst P., Rudenko G. Antigenic variation in African trypanosomes. Science. 1994 Jun 24;264(5167):1872–1873. doi: 10.1126/science.7516579. [DOI] [PubMed] [Google Scholar]
  10. Borst P., Rudenko G., Taylor M. C., Blundell P. A., Van Leeuwen F., Bitter W., Cross M., McCulloch R. Antigenic variation in trypanosomes. Arch Med Res. 1996 Autumn;27(3):379–388. [PubMed] [Google Scholar]
  11. Campbell D. A., van Bree M. P., Boothroyd J. C. The 5'-limit of transposition and upstream barren region of a trypanosome VSG gene: tandem 76 base-pair repeats flanking (TAA)90. Nucleic Acids Res. 1984 Mar 26;12(6):2759–2774. doi: 10.1093/nar/12.6.2759. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Carruthers V. B., Cross G. A. High-efficiency clonal growth of bloodstream- and insect-form Trypanosoma brucei on agarose plates. Proc Natl Acad Sci U S A. 1992 Sep 15;89(18):8818–8821. doi: 10.1073/pnas.89.18.8818. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Cross G. A. Antigenic variation in trypanosomes: secrets surface slowly. Bioessays. 1996 Apr;18(4):283–291. doi: 10.1002/bies.950180406. [DOI] [PubMed] [Google Scholar]
  14. Cross G. A. Identification, purification and properties of clone-specific glycoprotein antigens constituting the surface coat of Trypanosoma brucei. Parasitology. 1975 Dec;71(3):393–417. doi: 10.1017/s003118200004717x. [DOI] [PubMed] [Google Scholar]
  15. Cross G. A., Manning J. C. Cultivation of Trypanosoma brucei sspp. in semi-defined and defined media. Parasitology. 1973 Dec;67(3):315–331. doi: 10.1017/s0031182000046540. [DOI] [PubMed] [Google Scholar]
  16. Cully D. F., Ip H. S., Cross G. A. Coordinate transcription of variant surface glycoprotein genes and an expression site associated gene family in Trypanosoma brucei. Cell. 1985 Aug;42(1):173–182. doi: 10.1016/s0092-8674(85)80113-6. [DOI] [PubMed] [Google Scholar]
  17. De Lange T., Kooter J. M., Luirink J., Borst P. Transcription of a transposed trypanosome surface antigen gene starts upstream of the transposed segment. EMBO J. 1985 Dec 1;4(12):3299–3306. doi: 10.1002/j.1460-2075.1985.tb04080.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. De Lange T., Kooter J. M., Michels P. A., Borst P. Telomere conversion in trypanosomes. Nucleic Acids Res. 1983 Dec 10;11(23):8149–8165. doi: 10.1093/nar/11.23.8149. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Eid J., Sollner-Webb B. Stable integrative transformation of Trypanosoma brucei that occurs exclusively by homologous recombination. Proc Natl Acad Sci U S A. 1991 Mar 15;88(6):2118–2121. doi: 10.1073/pnas.88.6.2118. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Florent I., Baltz T., Raibaud A., Eisen H. On the role of repeated sequences 5' to variant surface glycoprotein genes in African trypanosomes. Gene. 1987;53(1):55–62. doi: 10.1016/0378-1119(87)90092-8. [DOI] [PubMed] [Google Scholar]
  21. Greaves D. R., Borst P. Trypanosoma brucei variant-specific glycoprotein gene chromatin is sensitive to single-strand-specific endonuclease digestion. J Mol Biol. 1987 Oct 5;197(3):471–483. doi: 10.1016/0022-2836(87)90559-6. [DOI] [PubMed] [Google Scholar]
  22. Hirumi H., Hirumi K. Continuous cultivation of Trypanosoma brucei blood stream forms in a medium containing a low concentration of serum protein without feeder cell layers. J Parasitol. 1989 Dec;75(6):985–989. [PubMed] [Google Scholar]
  23. Hoeijmakers J. H., Frasch A. C., Bernards A., Borst P., Cross G. A. Novel expression-linked copies of the genes for variant surface antigens in trypanosomes. Nature. 1980 Mar 6;284(5751):78–80. doi: 10.1038/284078a0. [DOI] [PubMed] [Google Scholar]
  24. Horn D., Cross G. A. A developmentally regulated position effect at a telomeric locus in Trypanosoma brucei. Cell. 1995 Nov 17;83(4):555–561. doi: 10.1016/0092-8674(95)90095-0. [DOI] [PubMed] [Google Scholar]
  25. Johnson P. J., Kooter J. M., Borst P. Inactivation of transcription by UV irradiation of T. brucei provides evidence for a multicistronic transcription unit including a VSG gene. Cell. 1987 Oct 23;51(2):273–281. doi: 10.1016/0092-8674(87)90154-1. [DOI] [PubMed] [Google Scholar]
  26. Kooter J. M., van der Spek H. J., Wagter R., d'Oliveira C. E., van der Hoeven F., Johnson P. J., Borst P. The anatomy and transcription of a telomeric expression site for variant-specific surface antigens in T. brucei. Cell. 1987 Oct 23;51(2):261–272. doi: 10.1016/0092-8674(87)90153-x. [DOI] [PubMed] [Google Scholar]
  27. Lambowitz A. M., Belfort M. Introns as mobile genetic elements. Annu Rev Biochem. 1993;62:587–622. doi: 10.1146/annurev.bi.62.070193.003103. [DOI] [PubMed] [Google Scholar]
  28. Lamont G. S., Tucker R. S., Cross G. A. Analysis of antigen switching rates in Trypanosoma brucei. Parasitology. 1986 Apr;92(Pt 2):355–367. doi: 10.1017/s003118200006412x. [DOI] [PubMed] [Google Scholar]
  29. Lee M. G., Van der Ploeg L. H. Homologous recombination and stable transfection in the parasitic protozoan Trypanosoma brucei. Science. 1990 Dec 14;250(4987):1583–1587. doi: 10.1126/science.2177225. [DOI] [PubMed] [Google Scholar]
  30. Ligtenberg M. J., Bitter W., Kieft R., Steverding D., Janssen H., Calafat J., Borst P. Reconstitution of a surface transferrin binding complex in insect form Trypanosoma brucei. EMBO J. 1994 Jun 1;13(11):2565–2573. doi: 10.1002/j.1460-2075.1994.tb06546.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Liu A. Y., Michels P. A., Bernards A., Borst P. Trypanosome variant surface glycoprotein genes expressed early in infection. J Mol Biol. 1985 Apr 5;182(3):383–396. doi: 10.1016/0022-2836(85)90198-6. [DOI] [PubMed] [Google Scholar]
  32. Liu A. Y., Van der Ploeg L. H., Rijsewijk F. A., Borst P. The transposition unit of variant surface glycoprotein gene 118 of Trypanosoma brucei. Presence of repeated elements at its border and absence of promoter-associated sequences. J Mol Biol. 1983 Jun 15;167(1):57–75. doi: 10.1016/s0022-2836(83)80034-5. [DOI] [PubMed] [Google Scholar]
  33. Matthews K. R., Shiels P. G., Graham S. V., Cowan C., Barry J. D. Duplicative activation mechanisms of two trypanosome telomeric VSG genes with structurally simple 5' flanks. Nucleic Acids Res. 1990 Dec 25;18(24):7219–7227. doi: 10.1093/nar/18.24.7219. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Matthews K. R., Tschudi C., Ullu E. A common pyrimidine-rich motif governs trans-splicing and polyadenylation of tubulin polycistronic pre-mRNA in trypanosomes. Genes Dev. 1994 Feb 15;8(4):491–501. doi: 10.1101/gad.8.4.491. [DOI] [PubMed] [Google Scholar]
  35. Michels P. A., Liu A. Y., Bernards A., Sloof P., Van der Bijl M. M., Schinkel A. H., Menke H. H., Borst P., Veeneman G. H., Tromp M. C. Activation of the genes for variant surface glycoproteins 117 and 118 in Trypanosoma brucei. J Mol Biol. 1983 Jun 5;166(4):537–556. doi: 10.1016/s0022-2836(83)80283-6. [DOI] [PubMed] [Google Scholar]
  36. Michels P. A., Van der Ploeg L. H., Liu A. Y., Borst P. The inactivation and reactivation of an expression-linked gene copy for a variant surface glycoprotein in Trypanosoma brucei. EMBO J. 1984 Jun;3(6):1345–1351. doi: 10.1002/j.1460-2075.1984.tb01975.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Navarro M., Cross G. A. DNA rearrangements associated with multiple consecutive directed antigenic switches in Trypanosoma brucei. Mol Cell Biol. 1996 Jul;16(7):3615–3625. doi: 10.1128/mcb.16.7.3615. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Pays E. Gene conversion in trypanosome antigenic variation. Prog Nucleic Acid Res Mol Biol. 1985;32:1–26. doi: 10.1016/s0079-6603(08)60344-x. [DOI] [PubMed] [Google Scholar]
  39. Pays E., Guyaux M., Aerts D., Van Meirvenne N., Steinert M. Telomeric reciprocal recombination as a possible mechanism for antigenic variation in trypanosomes. Nature. 1985 Aug 8;316(6028):562–564. doi: 10.1038/316562a0. [DOI] [PubMed] [Google Scholar]
  40. Pays E. Pseudogenes, chimaeric genes and the timing of antigen variation in African trypanosomes. Trends Genet. 1989 Dec;5(12):389–391. doi: 10.1016/0168-9525(89)90181-9. [DOI] [PubMed] [Google Scholar]
  41. Pays E., Tebabi P., Pays A., Coquelet H., Revelard P., Salmon D., Steinert M. The genes and transcripts of an antigen gene expression site from T. brucei. Cell. 1989 Jun 2;57(5):835–845. doi: 10.1016/0092-8674(89)90798-8. [DOI] [PubMed] [Google Scholar]
  42. Pays E., Van Assel S., Laurent M., Darville M., Vervoort T., Van Meirvenne N., Steinert M. Gene conversion as a mechanism for antigenic variation in trypanosomes. Cell. 1983 Sep;34(2):371–381. doi: 10.1016/0092-8674(83)90371-9. [DOI] [PubMed] [Google Scholar]
  43. Pays E., Van Meirvenne N., Le Ray D., Steinert M. Gene duplication and transposition linked to antigenic variation in Trypanosoma brucei. Proc Natl Acad Sci U S A. 1981 May;78(5):2673–2677. doi: 10.1073/pnas.78.5.2673. [DOI] [PMC free article] [PubMed] [Google Scholar]
  44. Pays E., Vanhamme L., Berberof M. Genetic controls for the expression of surface antigens in African trypanosomes. Annu Rev Microbiol. 1994;48:25–52. doi: 10.1146/annurev.mi.48.100194.000325. [DOI] [PubMed] [Google Scholar]
  45. Robertson B. D., Meyer T. F. Genetic variation in pathogenic bacteria. Trends Genet. 1992 Dec;8(12):422–427. doi: 10.1016/0168-9525(92)90325-x. [DOI] [PubMed] [Google Scholar]
  46. Roth C., Bringaud F., Layden R. E., Baltz T., Eisen H. Active late-appearing variable surface antigen genes in Trypanosoma equiperdum are constructed entirely from pseudogenes. Proc Natl Acad Sci U S A. 1989 Dec;86(23):9375–9379. doi: 10.1073/pnas.86.23.9375. [DOI] [PMC free article] [PubMed] [Google Scholar]
  47. Rothwell V., Aline R., Jr, Parsons M., Agabian N., Stuart K. Expression of a minichromosomal variant surface glycoprotein gene in Trypanosoma brucei. Nature. 1985 Feb 14;313(6003):595–597. doi: 10.1038/313595a0. [DOI] [PubMed] [Google Scholar]
  48. Rudenko G., Blundell P. A., Dirks-Mulder A., Kieft R., Borst P. A ribosomal DNA promoter replacing the promoter of a telomeric VSG gene expression site can be efficiently switched on and off in T. brucei. Cell. 1995 Nov 17;83(4):547–553. doi: 10.1016/0092-8674(95)90094-2. [DOI] [PubMed] [Google Scholar]
  49. Rudenko G., McCulloch R., Dirks-Mulder A., Borst P. Telomere exchange can be an important mechanism of variant surface glycoprotein gene switching in Trypanosoma brucei. Mol Biochem Parasitol. 1996 Sep;80(1):65–75. doi: 10.1016/0166-6851(96)02669-2. [DOI] [PubMed] [Google Scholar]
  50. Scholler J. K., Myler P. J., Stuart K. D. A novel telomeric gene conversion in Trypanosoma brucei. Mol Biochem Parasitol. 1989 Jun 1;35(1):11–19. doi: 10.1016/0166-6851(89)90137-0. [DOI] [PubMed] [Google Scholar]
  51. Schürch N., Hehl A., Vassella E., Braun R., Roditi I. Accurate polyadenylation of procyclin mRNAs in Trypanosoma brucei is determined by pyrimidine-rich elements in the intergenic regions. Mol Cell Biol. 1994 Jun;14(6):3668–3675. doi: 10.1128/mcb.14.6.3668. [DOI] [PMC free article] [PubMed] [Google Scholar]
  52. Shah J. S., Young J. R., Kimmel B. E., Iams K. P., Williams R. O. The 5' flanking sequence of a Trypanosoma brucei variable surface glycoprotein gene. Mol Biochem Parasitol. 1987 Jun;24(2):163–174. doi: 10.1016/0166-6851(87)90103-4. [DOI] [PubMed] [Google Scholar]
  53. Shea C., Glass D. J., Parangi S., Van der Ploeg L. H. Variant surface glycoprotein gene expression site switches in Trypanosoma brucei. J Biol Chem. 1986 May 5;261(13):6056–6063. [PubMed] [Google Scholar]
  54. Stanhope-Baker P., Hudson K. M., Shaffer A. L., Constantinescu A., Schlissel M. S. Cell type-specific chromatin structure determines the targeting of V(D)J recombinase activity in vitro. Cell. 1996 Jun 14;85(6):887–897. doi: 10.1016/s0092-8674(00)81272-6. [DOI] [PubMed] [Google Scholar]
  55. Thon G., Baltz T., Eisen H. Antigenic diversity by the recombination of pseudogenes. Genes Dev. 1989 Aug;3(8):1247–1254. doi: 10.1101/gad.3.8.1247. [DOI] [PubMed] [Google Scholar]
  56. Thon G., Baltz T., Giroud C., Eisen H. Trypanosome variable surface glycoproteins: composite genes and order of expression. Genes Dev. 1990 Aug;4(8):1374–1383. doi: 10.1101/gad.4.8.1374. [DOI] [PubMed] [Google Scholar]
  57. Turner C. M., Barry J. D. High frequency of antigenic variation in Trypanosoma brucei rhodesiense infections. Parasitology. 1989 Aug;99(Pt 1):67–75. doi: 10.1017/s0031182000061035. [DOI] [PubMed] [Google Scholar]
  58. Valdés J., Taylor M. C., Cross M. A., Ligtenberg M. J., Rudenko G., Borst P. The viral thymidine kinase gene as a tool for the study of mutagenesis in Trypanosoma brucei. Nucleic Acids Res. 1996 May 15;24(10):1809–1815. doi: 10.1093/nar/24.10.1809. [DOI] [PMC free article] [PubMed] [Google Scholar]
  59. Van der Ploeg L. H., Gottesdiener K., Lee M. G. Antigenic variation in African trypanosomes. Trends Genet. 1992 Dec;8(12):452–457. doi: 10.1016/0168-9525(92)90330-7. [DOI] [PubMed] [Google Scholar]
  60. Van der Ploeg L. H., Schwartz D. C., Cantor C. R., Borst P. Antigenic variation in Trypanosoma brucei analyzed by electrophoretic separation of chromosome-sized DNA molecules. Cell. 1984 May;37(1):77–84. doi: 10.1016/0092-8674(84)90302-7. [DOI] [PubMed] [Google Scholar]
  61. Van der Ploeg L. H., Valerio D., De Lange T., Bernards A., Borst P., Grosveld F. G. An analysis of cosmid clones of nuclear DNA from Trypanosoma brucei shows that the genes for variant surface glycoproteins are clustered in the genome. Nucleic Acids Res. 1982 Oct 11;10(19):5905–5923. doi: 10.1093/nar/10.19.5905. [DOI] [PMC free article] [PubMed] [Google Scholar]
  62. Van der Werf A., Van Assel S., Aerts D., Steinert M., Pays E. Telomere interactions may condition the programming of antigen expression in Trypanosoma brucei. EMBO J. 1990 Apr;9(4):1035–1040. doi: 10.1002/j.1460-2075.1990.tb08207.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  63. Wu T. C., Lichten M. Meiosis-induced double-strand break sites determined by yeast chromatin structure. Science. 1994 Jan 28;263(5146):515–518. doi: 10.1126/science.8290959. [DOI] [PubMed] [Google Scholar]
  64. Zomerdijk J. C., Ouellette M., ten Asbroek A. L., Kieft R., Bommer A. M., Clayton C. E., Borst P. The promoter for a variant surface glycoprotein gene expression site in Trypanosoma brucei. EMBO J. 1990 Sep;9(9):2791–2801. doi: 10.1002/j.1460-2075.1990.tb07467.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  65. ten Asbroek A. L., Ouellette M., Borst P. Targeted insertion of the neomycin phosphotransferase gene into the tubulin gene cluster of Trypanosoma brucei. Nature. 1990 Nov 8;348(6297):174–175. doi: 10.1038/348174a0. [DOI] [PubMed] [Google Scholar]

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