title: Characterization of RBP9 and RBP10, two developmentally regulated RNA-binding proteins inTrypanosoma brucei.
abstract: The fate of an mRNA is determined by its interaction with proteins and small RNAs within dynamic complexes called ribonucleoprotein complexes (mRNPs). InTrypanosoma bruceiand related kinetoplastids, responses to internal and external signals are mainly mediated by post-transcriptional processes. Here, we used proximity-dependent biotin identification (BioID) combined with RNA-seq to investigate the changes resulting from ectopic expression of RBP10 and RBP9, two developmentally regulated RNA-binding proteins (RBPs). Both RBPs have reduced expression in insect procyclic forms (PCFs) compared with bloodstream forms (BSFs). Upon overexpression in PCFs, both proteins were recruited to cytoplasmic foci, co-localizing with the processing body marker SCD6. Further, both RBPs altered the transcriptome from a PCF- to a BSF-like pattern. Notably, upon expression of BirA*-RBP9 and BirA*-RBP10, BioID yielded more than 200 high confidence protein interactors (more than 10-fold enriched); 45 (RBP9) and 31 (RBP10) were directly related to mRNA metabolism. This study validates the use of BioID for investigating mRNP components but also illustrates the complexity of mRNP function.
authors.affiliation: Department of Biochemistry, University of Cambridge, Cambridge CB2 1QW, UK.|Centre for Immunology and Infection (CII). Biology Dept., University of York, York YO10 5DD, UK.
authors.forename: Luis Miguel
authors.identifier: 0000-0001-8940-2919
authors.initials: LM
authors.lastname: De Pablos
authors.affiliation: Department of Plant Sciences, University of Oxford, South Parks Road, Oxford OX1 3RB, UK.
authors.forename: Steve
authors.initials: S
authors.lastname: Kelly
authors.affiliation: Department of Biochemistry, University of Cambridge, Cambridge CB2 1QW, UK.
authors.forename: Janaina
authors.initials: J
authors.lastname: de Freitas Nascimento
authors.affiliation: Sir William Dunn School of Pathology, University of Oxford, Oxford OX1 3RE, UK.
authors.forename: Jack
authors.initials: J
authors.lastname: Sunter
authors.affiliation: Department of Biochemistry, University of Cambridge, Cambridge CB2 1QW, UK mac115@cam.ac.uk.
authors.forename: Mark
authors.initials: M
authors.lastname: Carrington
mesh_terms: D002352:Carrier Proteins / Q000378:metabolism; D059447:Cell Cycle Checkpoints / Q000235:genetics; D019295:Computational Biology / Q000379:methods; D015870:Gene Expression; D011485:Protein Binding; D025941:Protein Interaction Mapping; D015800:Protozoan Proteins / Q000235:genetics / Q000378:metabolism*; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D016601:RNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D059467:Transcriptome; D014346:Trypanosoma brucei brucei / Q000235:genetics / Q000378:metabolism*
chemical_list: D002352:Carrier Proteins; D015800:Protozoan Proteins; D012333:RNA, Messenger; D016601:RNA-Binding Proteins
keywords: RNA; biotin identification; differentiation; gene expression; kinetoplastid