The dual role of the 16mer motif within the 3' untranslated region of the variant surface glycoprotein of Trypanosoma brucei

Abstract

The variant surface glycoprotein (VSG) of African trypanosomes is essential for survival of bloodstream form parasites. These parasites undergo antigenic variation, an immune evasion strategy in which they periodically switch VSG expression from one isoform to another. The molecular processes central to the expression and regulation of the VSG are however not fully understood. In general, the regulation of gene expression in trypanosomes is largely post-transcriptional. Regulatory sequences, mostly present in the 3 UTRs, often serve as key elements in the modulation of the levels of individual mRNAs. In T. brucei VSG genes, a 16mer motif within the 3 UTR has been shown to be essential for the stability of VSG transcripts and abundant VSG expression. This motif is 100 % conserved in the 3 UTRs of all transcribed and non-transcribed VSG genes. As a stability-associated sequence element, the absence of nucleotide substitutions in the 16mer is however exceptional. We therefore hypothesised that the motif is involved in other essential roles/processes besides stability of the VSG transcripts. In this study, we demonstrate that the 100 % conservation of the 16mer motif is not essential for cell viability or for the maintenance of functional VSG protein levels. We further show that the intact motif in the active VSG 3 UTR is neither required to promote VSG silencing during switching nor is it needed during differentiation from bloodstream forms to procyclic forms. Ectopic overexpression of a second VSG, however, requires the intact 16mer motif within the ectopic VSG 3 UTR to trigger silencing and exchange of the active VSG, suggesting a role for the motif in transcriptional VSG switching. The enigmatic 16mer motif therefore appears to play a dual role in transcriptional VSG switching and VSG transcript stability.