The role of genomic location and flanking 3'UTR in the generation of functional levels of variant surface glycoprotein in Trypanosoma brucei

CHER-PHENG OOI, Sophie Ridewood, Belinda Hall, Anna Trenaman, Nadina Wand, Georgios Sioutas, Iris Scherwitzl, Rudenko Gloria

Research output: Contribution to journalArticle

7 Citations (Scopus)
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Abstract

Trypanosoma brucei faces relentless immune attack in the mammalian bloodstream, where it is protected by an essential coat of Variant Surface Glycoprotein (VSG) comprising ∼10% total protein. The active VSG gene is in a Pol I-transcribed telomeric expression site (ES). We investigated factors mediating these extremely high levels of VSG expression by inserting ectopic VSG117 into VSG221 expressing T. brucei. Mutational analysis of the ectopic VSG 3'UTR demonstrated the essentiality of a conserved 16-mer for mRNA stability. Expressing ectopic VSG117 from different genomic locations showed that functional VSG levels could be produced from a gene 60 kb upstream of its normal telomeric location. High, but very heterogeneous levels of VSG117 were obtained from the Pol I-transcribed rDNA. Blocking VSG synthesis normally triggers a precise precytokinesis cell-cycle checkpoint. VSG117 expression from the rDNA was not adequate for functional complementation, and the stalled cells arrested prior to cytokinesis. However, VSG levels were not consistently low enough to trigger a characteristic 'VSG synthesis block' cell-cycle checkpoint, as some cells reinitiated S phase. This demonstrates the essentiality of a Pol I-transcribed ES, as well as conserved VSG 3'UTR 16-mer sequences for the generation of functional levels of VSG expression in bloodstream form T. brucei.
Original languageEnglish
Pages (from-to)614 - 634
Number of pages21
JournalMolecular Microbiology
Volume106
Issue number4
Early online date14 Sep 2017
DOIs
Publication statusPublished - 14 Nov 2017

Keywords

  • genomic location
  • Trypanosoma brucei

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    CHER-PHENG OOI, PhD

    • Biology - Senior Lecturer in Microbiology

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