Characterization of the Variable Antigen Genes Expressed by Trypanosoma brucei rhodesiense during Metacyclic Stage and in the Bloodstream

摘要

Two different cDNA libraries from the mRNAs of day 5 trypanosomes which have been shown to be expressing predominately metacyclic variable surface glycoproteins (M-VSGs) have been constructed. In these libraries have been identified cDNA clones that contain the coding sequences for 4 of the 12 - 15 different M-VSGs expressed by metacyclic Trypanosoma brucei rhodesiense. These cDNAs have been sequenced and used as probes to characterize the corresponding chromosomal genes. They have also been subcloned into E. coli expression vectors so that the bacteria synthesize large quantities of fusion proteins expressing vectors so that the bacteria synthesize large quantities of fusion proteins expressing the M-VSG epitopes. It is anticipated that these fusion proteins can be used for immunological studies. As a corollary to the M-VSG project, we are also investigating several aspects of the DNA rearrangements that are associated with the expression of VSGs during the bloodstream stages of the parasite infection. We have determined the complete sequence of a 1.35 kb repeat sequence that contains the coding sequence for the 35 nucleotides that are found at the 5'-terminal of all VSG mRNAs. Fragments containing this sequence are now being used as probes to investigate the transcription of these 35 nucleotides in isolated trypanosome nuclei under a variety of conditions. During the coming year we plan to continue to expand both the metacyclic VSG project and the transcription studies.