Abstract
Trypanosoma brucei contains more than a hundred genes coding for the different variant surface glycoproteins (VSGs). Activation of some of these genes involves the duplication of the gene (the basic copy or BC) and transposition of the duplicate to an expression site (yielding the expression-linked copy or ELC). We have cloned large fragments of genomic DNA in cosmid vectors in Escherichia coli. Cosmids containing the BCs of genes 117, 118 and 121 were readily obtained, but DNA containing the ELCs was strongly selected against in the cosmid and plasmid cloning systems used. We have analysed the distribution of VSG genes in the genome using probes for the sequences at the edges of the transposed segment which are partially homologous among these genes. In genomic cosmid clone banks, about 9% of all colonies hybridize with probes from the 5'- and 3'-edges of the transposed segment, showing that these sequences are linked in the genome. Moreover, the 117 and 118 BC cosmids contain several additional putative VSG genes in tandem, as deduced from hybridization and sequence analyses. We conclude that the VSG genes are highly clustered and share common sequences at the borders of the transposed segment.
Full text
PDF


















Images in this article
Selected References
These references are in PubMed. This may not be the complete list of references from this article.
- 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]
- Birnboim H. C., Doly J. A rapid alkaline extraction procedure for screening recombinant plasmid DNA. Nucleic Acids Res. 1979 Nov 24;7(6):1513–1523. doi: 10.1093/nar/7.6.1513. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Boothroyd J. C., Cross G. A., Hoeijmakers J. H., Borst P. A variant surface glycoprotein of Trypanosoma brucei synthesized with a C-terminal hydrophobic 'tail' absent from purified glycoprotein. Nature. 1980 Dec 11;288(5791):624–626. doi: 10.1038/288624a0. [DOI] [PubMed] [Google Scholar]
- 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]
- Borst P., Fase-Fowler F., Gibson W. C. Quantitation of genetic differences between Trypanosoma brucei gambiense, rhodesiense and brucei by restriction enzyme analysis of kinetoplast DNA. Mol Biochem Parasitol. 1981 Jun;3(2):117–131. doi: 10.1016/0166-6851(81)90011-6. [DOI] [PubMed] [Google Scholar]
- Borst P., Fase-Fowler F. The maxi-circle of Trypanosoma brucei kinetoplast DNA. Biochim Biophys Acta. 1979 Nov 22;565(1):1–12. doi: 10.1016/0005-2787(79)90078-9. [DOI] [PubMed] [Google Scholar]
- Borst P., Frasch A. C., Bernards A., Van der Ploeg L. H., Hoeijmakers J. H., Arnberg A. C., Cross G. A. DNA rearrangements involving the genes for variant antigens in Trypanosoma brucei. Cold Spring Harb Symp Quant Biol. 1981;45(Pt 2):935–943. doi: 10.1101/sqb.1981.045.01.110. [DOI] [PubMed] [Google Scholar]
- Boyer H. W., Roulland-Dussoix D. A complementation analysis of the restriction and modification of DNA in Escherichia coli. J Mol Biol. 1969 May 14;41(3):459–472. doi: 10.1016/0022-2836(69)90288-5. [DOI] [PubMed] [Google Scholar]
- Cami B., Kourilsky P. Screening of cloned recombinant DNA in bacteria by in situ colony hybridization. Nucleic Acids Res. 1978 Jul;5(7):2381–2390. doi: 10.1093/nar/5.7.2381. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Capbern A., Giroud C., Baltz T., Mattern P. Trypanosoma equiperdum: etude des variations antigéniques au cours de la trypanosomose experimentale du lapin. Exp Parasitol. 1977 Jun;42(1):6–13. doi: 10.1016/0014-4894(77)90055-8. [DOI] [PubMed] [Google Scholar]
- Denhardt D. T. A membrane-filter technique for the detection of complementary DNA. Biochem Biophys Res Commun. 1966 Jun 13;23(5):641–646. doi: 10.1016/0006-291x(66)90447-5. [DOI] [PubMed] [Google Scholar]
- Fairlamb A. H., Weislogel P. O., Hoeijmakers J. H., Borst P. Isolation and characterization of kinetoplast DNA from bloodstream form of Trypanosoma brucei. J Cell Biol. 1978 Feb;76(2):293–309. doi: 10.1083/jcb.76.2.293. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Grosveld F. G., Dahl H. H., de Boer E., Flavell R. A. Isolation of beta-globin-related genes from a human cosmid library. Gene. 1981 Apr;13(3):227–237. doi: 10.1016/0378-1119(81)90028-7. [DOI] [PubMed] [Google Scholar]
- Hanahan D., Meselson M. Plasmid screening at high colony density. Gene. 1980 Jun;10(1):63–67. doi: 10.1016/0378-1119(80)90144-4. [DOI] [PubMed] [Google Scholar]
- 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]
- Ish-Horowicz D., Burke J. F. Rapid and efficient cosmid cloning. Nucleic Acids Res. 1981 Jul 10;9(13):2989–2998. doi: 10.1093/nar/9.13.2989. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Jeffreys A. J., Flavell R. A. A physical map of the DNA regions flanking the rabbit beta-globin gene. Cell. 1977 Oct;12(2):429–439. doi: 10.1016/0092-8674(77)90119-2. [DOI] [PubMed] [Google Scholar]
- Majumder H. K., Boothroyd J. C., Weber H. Homologous 3'-terminal regions of mRNAs for surface antigens of different antigenic variants of Trypanosoma brucei. Nucleic Acids Res. 1981 Sep 25;9(18):4745–4753. doi: 10.1093/nar/9.18.4745. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Matthyssens G., Michiels F., Hamers R., Pays E., Steinert M. Two variant surface glycoproteins of Trypanosoma brucei have a conserved C-terminus. Nature. 1981 Sep 17;293(5829):230–233. doi: 10.1038/293230a0. [DOI] [PubMed] [Google Scholar]
- Michels P. A., Bernards A., Van der Ploeg L. H., Borst P. Characterization of the expression-linked gene copies of variant surface glycoprotein 118 in two independently isolated clones of Trypanosoma brucei. Nucleic Acids Res. 1982 Apr 10;10(7):2353–2366. doi: 10.1093/nar/10.7.2353. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Morrison D. A. Transformation and preservation of competent bacterial cells by freezing. Methods Enzymol. 1979;68:326–331. doi: 10.1016/0076-6879(79)68023-0. [DOI] [PubMed] [Google Scholar]
- 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]
- Rigby P. W., Dieckmann M., Rhodes C., Berg P. Labeling deoxyribonucleic acid to high specific activity in vitro by nick translation with DNA polymerase I. J Mol Biol. 1977 Jun 15;113(1):237–251. doi: 10.1016/0022-2836(77)90052-3. [DOI] [PubMed] [Google Scholar]
- Sato S., Hutchinson C. A., 3rd, Harris J. I. A thermostable sequence-specific endonuclease from Thermus aquaticus. Proc Natl Acad Sci U S A. 1977 Feb;74(2):542–546. doi: 10.1073/pnas.74.2.542. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Southern E. M. Detection of specific sequences among DNA fragments separated by gel electrophoresis. J Mol Biol. 1975 Nov 5;98(3):503–517. doi: 10.1016/s0022-2836(75)80083-0. [DOI] [PubMed] [Google Scholar]
- Van der Ploeg L. H., Bernards A., Rijsewijk F. A., Borst P. Characterization of the DNA duplication-transposition that controls the expression of two genes for variant surface glycoproteins in Trypanosoma brucei. Nucleic Acids Res. 1982 Jan 22;10(2):593–609. doi: 10.1093/nar/10.2.593. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Van der Ploeg L. H., Liu A. Y., Michels P. A., De Lange T., Borst P., Majumder H. K., Weber H., Veeneman G. H., Van Boom J. RNA splicing is required to make the messenger RNA for a variant surface antigen in trypanosomes. Nucleic Acids Res. 1982 Jun 25;10(12):3591–3604. doi: 10.1093/nar/10.12.3591. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Vickerman K. Antigenic variation in trypanosomes. Nature. 1978 Jun 22;273(5664):613–617. doi: 10.1038/273613a0. [DOI] [PubMed] [Google Scholar]
- Vogelstein B., Gillespie D. Preparative and analytical purification of DNA from agarose. Proc Natl Acad Sci U S A. 1979 Feb;76(2):615–619. doi: 10.1073/pnas.76.2.615. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Wahl G. M., Stern M., Stark G. R. Efficient transfer of large DNA fragments from agarose gels to diazobenzyloxymethyl-paper and rapid hybridization by using dextran sulfate. Proc Natl Acad Sci U S A. 1979 Aug;76(8):3683–3687. doi: 10.1073/pnas.76.8.3683. [DOI] [PMC free article] [PubMed] [Google Scholar]