title: Reduction of Tubulin Expression in Angomonas deanei by RNAi Modifies the Ultrastructure of the Trypanosomatid Protozoan and Impairs Division of Its Endosymbiotic Bacterium.
abstract: In the last two decades, RNA interference pathways have been employed as a useful tool for reverse genetics in trypanosomatids. Angomonas deanei is a nonpathogenic trypanosomatid that maintains an obligatory endosymbiosis with a bacterium related to the Alcaligenaceae family. Studies of this symbiosis can help us to understand the origin of eukaryotic organelles. The recent elucidation of both the A. deanei and the bacterium symbiont genomes revealed that the host protozoan codes for the enzymes necessary for RNAi activity in trypanosomatids. Here, we tested the functionality of the RNAi machinery by transfecting cells with dsRNA to a reporter gene (green fluorescent protein), which had been previously expressed in the parasite and to α-tubulin, an endogenous gene. In both cases, protein expression was reduced by the presence of specific dsRNA, inducing, respectively, a decreased GFP fluorescence and the formation of enlarged cells with modified arrangement of subpellicular microtubules. Furthermore, symbiont division was impaired. These results indicate that the RNAi system is active in A. deanei and can be used to further explore gene function in symbiont-containing trypanosomatids and to clarify important aspects of symbiosis and cell evolution.
authors.affiliation: Laboratório de Ultraestrutura Celular Hertha Meyer, Instituto de Biofísica Carlos Chagas Filho, Universidade Federal do Rio de Janeiro, Av. Carlos Chagas Filho 373, bloco G/SS, 21949-900, Rio de Janeiro, RJ, Brazil.|Wellcome Trust Centre for Molecular Parasitology, Institute of Infection, Immunity and Inflammation, College of Medical, Veterinary and Life Sciences, University of Glasgow, Glasgow, G12 8TA, United Kingdom.|Department of Biology, Centre for Immunology and Infection, University of York, York, YO10 5DD, United Kingdom.
authors.forename: Carolina Moura Costa
authors.identifier: 0000-0001-8983-3487
authors.initials: CM
authors.lastname: Catta-Preta
authors.affiliation: Department of Microbiology, Immunology and Parasitology, Escola Paulista de Medicina, Universidade Federal de São Paulo, R. Pedro de Toledo 669, L6A, 04039-032, São Paulo, Brazil.
authors.forename: Bruno
authors.initials: B
authors.lastname: Dos Santos Pascoalino
authors.affiliation: Laboratório de Ultraestrutura Celular Hertha Meyer, Instituto de Biofísica Carlos Chagas Filho, Universidade Federal do Rio de Janeiro, Av. Carlos Chagas Filho 373, bloco G/SS, 21949-900, Rio de Janeiro, RJ, Brazil.
authors.forename: Wanderley
authors.initials: W
authors.lastname: de Souza
authors.affiliation: Wellcome Trust Centre for Molecular Parasitology, Institute of Infection, Immunity and Inflammation, College of Medical, Veterinary and Life Sciences, University of Glasgow, Glasgow, G12 8TA, United Kingdom.|Department of Biology, Centre for Immunology and Infection, University of York, York, YO10 5DD, United Kingdom.
authors.forename: Jeremy C
authors.initials: JC
authors.lastname: Mottram
authors.affiliation: Laboratório de Ultraestrutura Celular Hertha Meyer, Instituto de Biofísica Carlos Chagas Filho, Universidade Federal do Rio de Janeiro, Av. Carlos Chagas Filho 373, bloco G/SS, 21949-900, Rio de Janeiro, RJ, Brazil. motta@biof.ufrj.br.
authors.forename: Maria Cristina M
authors.initials: MC
authors.lastname: Motta
authors.affiliation: Department of Microbiology, Immunology and Parasitology, Escola Paulista de Medicina, Universidade Federal de São Paulo, R. Pedro de Toledo 669, L6A, 04039-032, São Paulo, Brazil. sschenkman@unifesp.br.
authors.forename: Sergio
authors.initials: S
authors.lastname: Schenkman
mesh_terms: D001419:Bacteria / Q000166:cytology* / Q000235:genetics; D002455:Cell Division; D015800:Protozoan Proteins / Q000235:genetics* / Q000378:metabolism; D034622:RNA Interference; D013559:Symbiosis; D014351:Trypanosomatina / Q000235:genetics / Q000378:metabolism / Q000382:microbiology* / Q000648:ultrastructure; D014404:Tubulin / Q000235:genetics / Q000378:metabolism
chemical_list: D015800:Protozoan Proteins; D014404:Tubulin
keywords: Cell division; cytoskeleton; endosymbiont