M Laurent

Gene conversion as a mechanism for antigenic variation in Trypanosomes

Expression of the gene coding for the trypanosome AnTat 1.1 surface antigen is linked to the duplicative transposition of a basic copy (BC) of this gene to an expression site. In two trypanosome clones successively derived from AnTat 1.1 (AnTat 1.10 and AnTat 1.1B) we found evidence that gene conversions are involved in the transformation of the AnTat 1.1 transposed element into the two new surface antigen coding sequences. Although the three resultant mRNAs--AnTat 1.1, 1.10, and 1.1B--are different, they still share large homologies. Two of them, AnTat 1.1 and 1.1B, code for surface coats that are indistinguishable by conventional serological techniques, whereas AnTat 1.10 has been found different by the same methods. The three genomic rearrangements involve two of the five members of the AnTat 1.1 gene family. These two members are both located in unstable telomeric regions similar to the expression site, each in a different orientation with respect to the DNA terminus. We have concluded that the duplicative transposition is achieved by a gene conversion that may affect variable lengths of the same silent genes, and that different members of the same surface antigen gene family can contribute to the diversification of the antigen repertoire.

Modifications of a trypanosoma b. brucei antigen gene repertoire by different DNA recombinational mechanisms

In the Trypanosoma b. brucei AnTat 1.1C clone, the gene coding for the variant-specific surface antigen is telomeric and appears as a hybrid sequence, partially modified by gene conversion. This conversion is very similar to that observed in another AnTat 1.1-expressor clone (AnTat 1.1B). This sequence is not activated by duplicative transposition, although it could be activated by duplication in another clone (AnTat 1.10). Instead activation of the AnTat 1.1C gene seems operated by reciprocal recombination between its own telomere and the telomere carrying the previous (AnTat 1.16) ELC. Indeed, from the switch to AnTat 1.1C onward, the AnTat 1.16 ELC becomes a new silent member of its gene family, whereas in the variant directly derived from AnTat 1.1C (AnTat 1.3B), the AnTat 1.1C-containing telomere is lost, probably replaced by a large duplicate, at least 40 kb long, of the AnTat 1.3 gene-containing telomere. Different DNA rearrangement mechanisms used by the trypanosome to change its antigenic type thus contribute, by gain and loss of genes, to the evolution of the repertoire for surface antigens.

Possible DNA modification in GC dinucleotides of Trypanosoma brucei telomeric sequences; relationship with antigen gene transcription.

Polymorphism in restriction site cleavage (PstI, SphI, PvuII, HindIII) has been noticed in several occasions in the telomeric sequences harbouring trypanosome variant-specific antigen genes (1, 2, 3). This polymorphism has been further investigated and seems best interpreted as due to partial DNA modification in GC dinucleotides. The actively transcribed telomeric genes do not exhibit such a polymorphism; furthermore, in at least three independent cases, gene inactivation is linked to the appearance of polymorphism. It could thus be hypothesized that DNA modification prevents antigen gene transcription, or vice-versa. We report however that at least some telomeric antigen-specific sequences of the procyclic trypanosomes (in vitro culture form) are not polymorphic, although they do not synthesize any variant-specific antigen mRNA. There is thus no absolute relationship between the absence of polymorphism and antigen gene transcription.

Kinetoplast DNA. A unique macromolecular structure of considerable size and mechanical resistance

Abstract Highly purified k-DNA has been prepared from Crithidia luciliae by a two step procedure, involving differential sedimentation in sucrose followed by banding in a CsCl density gradient. This k-DNA is isolated as compact particles of homogeneous size and DNA content, each one corresponding to the whole DNA complement of individual kinetoplasts (3.72 × 10−14g).