Depletion of Trypanosome CTR9 Leads to Gene Expression Defects

Benard A Ouna,Christine Clayton,Claudia Helbig,Benson Nyambega,Theresa Manful,Abeer Fadda,Matilda Males,Ziyin Li

doi:10.1371/journal.pone.0034256

Abstract

The Paf complex of Opisthokonts and plants contains at least five subunits: Paf1, Cdc73, Rtf1, Ctr9, and Leo1. Mutations in, or loss of Paf complex subunits have been shown to cause defects in histone modification, mRNA polyadenylation, and transcription by RNA polymerase I and RNA polymerase II. We here investigated trypanosome CTR9, which is essential for trypanosome survival. The results of tandem affinity purification suggested that trypanosome CTR9 associates with homologues of Leo1 and Cdc73; genes encoding homologues of Rtf1 and Paf1 were not found. RNAi targeting CTR9 resulted in at least ten-fold decreases in 131 essential mRNAs: they included several that are required for gene expression and its control, such as those encoding subunits of RNA polymerases, exoribonucleases that target mRNA, RNA helicases and RNA-binding proteins. Simultaneously, some genes from regions subject to chromatin silencing were derepressed, possibly as a secondary effect of the loss of two proteins that are required for silencing, ISWI and NLP1.

Highlights

The Paf complex has been implicated in several different aspects of gene expression within the nuclei of yeast and mammalian cells (Opisthokonts) and plants: defects have been shown to influence transcriptional elongation and termination, mRNA processing and export, and to result in epigenetic changes [1]
Trypanosome CTR9 is an Essential Protein which Copurifies with Homologues of Leo1 and Cdc73
The tagged CTR9 was subjected to two steps of affinity purification and the proteins present in the entire preparation were identified by mass spectrometry

Summary

Introduction

The Paf complex has been implicated in several different aspects of gene expression within the nuclei of yeast and mammalian cells (Opisthokonts) and plants: defects have been shown to influence transcriptional elongation and termination, mRNA processing and export, and to result in epigenetic changes [1]. In Saccharomyces cerevisiae, the Paf complex consists of five subunits: Paf, Cdc, Rtf, Ctr, and Leo; the human complex contains Ski8 [1]. Mutation in Rtf or Leo led to developmental defects [4,5], while in Arabidopisis, the complex is involved in transcriptional control of flowering [6]. The complex has been shown to bind to RNA polymerase II; mutations in either Cdc or Rtf abolished the binding and resulted in reduced phosphorylation of the C-terminal domain or the largest polymerase II subunit [7].

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Results

Conclusion