Authors

Discussion

Antigenic variation allows Trypanosoma brucei to evade the immune system, survive, and proliferate in its mammalian host. This process is dependent on monoallelic expression of variant surface glycoproteins (VSGs). While the parasites possess an extensive repertoire of VSG genes, only one expression-site (VSG-ES) is active at one time. Stochastic antigenic switching allows the cells to evade the immune response, whereas cells expressing multiple VSGs are easily cleared by the immune system.

The expression-site body (ESB) is a subnuclear structure where the single active VSG-ES is transcribed by RNA polymerase I (Pol-I). VSG-Exclusion-2 (VEX2) is a large RNA:DNA helicase found at the ESB which has been shown to be essential for VSG monoallelic expression. Previous analysis using Hi-C and scRNA-Seq showed that VEX2 depletion leads to dramatic changes in genome organisation that result in multiple antigen expression, however the mechanism is not fully understood.

We sought to dissect VEX2 function and recruitment to the ESB by using a tet-inducible system to overexpress wild-type, mutated, and truncated versions of VEX2.These results were analysed using fluorescence microscopy and transcriptomics.

Super resolution microscopy showed that VEX2 overexpression leads to itsaccumulation at the active-VSG-ES and at the periphery of the nucleolus, revealing intrinsic affinity for sites of Pol-I transcription. We also found that such specificity is mediated by sequences within its N-terminus and that the helicase activity is essential for VSG expression control.

In addition, by specifically blocking transcription at the active-VSG-ES, we found that active transcription is crucial for VEX2 localization to the ESB, indicating the role of RNA in its compartmentalisation within this structure. Future research will aim to identify the RNA sequences that VEX2 interacts with.