Searching for novel cell cycle regulators in Trypanosoma brucei with an RNA interference screen

Monnerat, S., Clucas, C. , Brown, E., Mottram, J. C. and Hammarton, T. C. (2009) Searching for novel cell cycle regulators in Trypanosoma brucei with an RNA interference screen. BMC Research Notes, 2(46), (doi:10.1186/1756-0500-2-46)

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Publisher's URL: http://dx.doi.org/10.1186/1756-0500-2-46

Abstract

<b>Background</b><br /> The protozoan parasite, Trypanosoma brucei, is spread by the tsetse fly and causes Human African Trypanosomiasis. Its cell cycle is complex and not fully understood at the molecular level. The T. brucei genome contains over 6000 protein coding genes with >50% having no predicted function. A small scale RNA interference (RNAi) screen was carried out in Trypanosoma brucei to evaluate the prospects for identifying novel cycle regulators.<p></p> <b>Results</b><br /> Procyclic form T. brucei were transfected with a genomic RNAi library and 204 clones isolated. However, only 76 RNAi clones were found to target a protein coding gene of potential interest. These clones were screened for defects in proliferation and cell cycle progression following RNAi induction. Sixteen clones exhibited proliferation defects upon RNAi induction, with eight clones displaying potential cell cycle defects. To confirm the phenotypes, new RNAi cell lines were generated and characterised for five genes targeted in these clones. While we confirmed that the targeted genes are essential for proliferation, we were unable to unambiguously classify them as cell cycle regulators.<p></p> <b>Conclusion</b><br /> Our study identified genes essential for proliferation, but did not, as hoped, identify novel cell cycle regulators. Screening of the RNAi library for essential genes was extremely labour-intensive, which was compounded by the suboptimal quality of the library. For such a screening method to be viable for a large scale or genome wide screen, a new, significantly improved RNAi library will be required, and automated phenotyping approaches will need to be incorporated.<p></p>

Item Type:Articles (Other)
Status:Published
Refereed:Yes
Glasgow Author(s) Enlighten ID:Clucas, Dr Caroline and Hammarton, Dr Tansy and Brown, Miss Elaine and Mottram, Professor Jeremy
Authors: Monnerat, S., Clucas, C., Brown, E., Mottram, J. C., and Hammarton, T. C.
Subjects:Q Science > QR Microbiology
College/School:College of Medical Veterinary and Life Sciences > Institute of Biodiversity Animal Health and Comparative Medicine
College of Medical Veterinary and Life Sciences > Institute of Infection Immunity and Inflammation
Journal Name:BMC Research Notes
Publisher:BioMed Central
ISSN:1756-0500
ISSN (Online):1756-0500
Copyright Holders:Copyright © 2009 The Authors
First Published:First published in BMC Research Notes 2(46)
Publisher Policy:Reproduced under a Creative Commons License
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Project CodeAward NoProject NamePrincipal InvestigatorFunder's NameFunder RefLead Dept
403591Regulation of cytokinesis in trypanosoma bruceiTansy HammartonMedical Research Council (MRC)G120/1001III - PARASITOLOGY