MECHANISM OF ANTIGENIC VARIATION IN TRYPANOSOMA BRUCEI

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Antigenic variation of trypanosomes is brought about by the periodic expression of genes that code for the Variant Cell Surface Glycoprotein (VSG) coat. The expression of immunologically distinct surface coats enables trypanosomes that switched to the expression of new coat to escape immune destruction. The genetic program underlying antigenic variation involves the more or less ordered expression of different VSG genes. Some VSG genes can be activated by a duplicative transposition of the gene to an expression site located at a chromosome end or telomere; other VSG genes that are always located at telomeres can be activated without detectable genomic recombinations in the vicinity of the gene. I have shown that several of these represent different telomeric expression sites situated on different chromosomes (4). It is the aim of this project to investigate the mutually exclusive activation of these telomeric expression sites and the involvement of chromosomal recombinations in their regulation. Recently I have been able to size separate trypanosomal chromosomes by a new electrophoretic technique. This allowed the detection of previously unobserved chromosome rearrangements, displacing hundreds of kilobasepairs. These could explain transcriptional regulation of the expression sites by a position effect on the VSG gene promoter due to the chromosomal recombinations. In order to examine the mutually exclusive regulation of the expression sites I will isolate recombination mutants, clone an area of 150 kb comprising the expression site of VSG gene 1.8 and investigate the regulatory effect of the recombinations on VSG gene expression: by localisation and comparison of the VSG gene promoter in different recombination mutants that activated the same site. I will also examine the frequency and mechanism underlying the chromosome recombinations: by cloning of the mutant chromosome recombination regions and analysis of the nature of the recombinations by determination of the nucleotide sequences at donor and acceptor sites. I will compare the chromosome repertoire and chromosomal stability in T.brucei, with that of other Kinetoplastida (species of the genera Trypanosoma, Leishmania, Leptomonas, and Herpetomonas). This will give insight in the chromosome stability and mutation frequency in these protozoa which is of importance for our understanding of the forces molding the parasites genome in its continuous adaptation to a highly variable environment.