Bloodstream and metacyclic variant surface glycoprotein gene expression sites of Trypanosoma brucei gambiense

Abstract

Trypanosoma brucei gambiense is the causative agent of chronic human sleeping sickness. Previous studies have indicated that T. b. gambiense isolates expressed the antigens U1 or L2 in both the metacyclic and early bloodstream form of the parasite life cycle. These studies suggested that L2 and U1 were likely to be metacyclic variant surface glycoproteins (mVSG). The basic copies of the genes encoding the VSGs L2 and U1 are present in single copy in non-expressing isolates of T. b. gambiense. Furthermore, they have been found to be maintained stably in a large number of stocks isolated from a wide geographic area over a 30-year period. The genomic DNA comprising the upstream 5′ flanking regions of the U1 and L2 putative mVSG gene expression sites have been cloned from bloodstream forms of T. b. gambiense. The L2 expression site clone, containing 12.5 kb of sequences 5′ to the VSG gene, was found to lack the 72/76-bp repeat unit generally found in the ‘barren’ region upstream of bloodstream form expression sites. The U1 expression site clone, containing 13.5 kb of the 5′ flanking region, appeared to have these repeats, which were localized to 2 kb of DNA immediately 5′ to the U1 mVSG gene. Neither the U1 nor the L2 clone was found to have ESAG2 or ESAG3 gene sequences, but both were found to have ESAG1 genes. The ESAG1 genes from the putative metacyclic expression sites and from the U1 and L2 bloodstream form expression sites (in the form of cDNA clones) were sequenced and compared to all other published ESAG1 sequences.