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A non-coding role for trypanosome VSG transcripts in allelic exclusion.

Douglas O Escrivani1, Sebastian Hutchinson1, Michele Tinti1

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African trypanosomes use Variant Surface Glycoprotein (VSG) transcripts to control gene expression. The VSG transcript actively participates in competition, influencing its own exclusion and nuclear organization.

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Area of Science:

  • Molecular Biology
  • Parasitology
  • Genetics

Background:

  • Bloodstream-form African trypanosomes exhibit antigenic variation through Variant Surface Glycoprotein (VSG) gene switching.
  • VSG expression is tightly regulated, requiring mono-telomeric expression and exclusion of other VSG genes, mediated by VSG exclusion factors (VEX1-2).
  • The precise mechanisms maintaining VSG exclusion and the role of the VSG transcript itself are not fully understood.

Purpose of the Study:

  • To investigate the role of the VSG transcript in allelic competition and VSG gene exclusion.
  • To determine if the VSG transcript influences the maintenance of VSG exclusion.
  • To explore the functional consequences of perturbing VSG transcript levels or translation.

Main Methods:

  • Recruitment of MS2 coat protein to the VSG 5'-untranslated region to block translation.
  • RNA interference (RNAi) to deplete VSG transcripts.
  • Monitoring VSG exclusion, DNA replication, mitosis, and VEX2 focus formation.

Main Results:

  • Neither translation blockade nor transcript depletion alone significantly affected native VSG exclusion.
  • A VSG transgene escaped exclusion specifically when the native VSG transcript was transiently depleted.
  • Perturbations blocked cytokinesis, but DNA replication and mitosis continued during translational blockade.
  • Increased VEX2 foci were observed in cells with a second active VSG.

Conclusions:

  • The VSG transcript acts as a bifunctional molecule, serving as both coding and non-coding RNA.
  • The VSG transcript actively participates in allelic competition to establish VSG exclusion.
  • VSG transcript-mediated exclusion represents a form of RNA-mediated symmetry breaking that influences nuclear architecture.