title: Replication Protein A Presents Canonical Functions and Is Also Involved in the Differentiation Capacity of Trypanosoma cruzi.
abstract: Replication Protein A (RPA), the major single stranded DNA binding protein in eukaryotes, is composed of three subunits and is a fundamental player in DNA metabolism, participating in replication, transcription, repair, and the DNA damage response. In human pathogenic trypanosomatids, only limited studies have been performed on RPA-1 from Leishmania. Here, we performed in silico, in vitro and in vivo analysis of Trypanosoma cruzi RPA-1 and RPA-2 subunits. Although computational analysis suggests similarities in DNA binding and Ob-fold structures of RPA from T. cruzi compared with mammalian and fungi RPA, the predicted tridimensional structures of T. cruzi RPA-1 and RPA-2 indicated that these molecules present a more flexible tertiary structure, suggesting that T. cruzi RPA could be involved in additional responses. Here, we demonstrate experimentally that the T. cruzi RPA complex interacts with DNA via RPA-1 and is directly related to canonical functions, such as DNA replication and DNA damage response. Accordingly, a reduction of TcRPA-2 expression by generating heterozygous knockout cells impaired cell growth, slowing down S-phase progression. Moreover, heterozygous knockout cells presented a better efficiency in differentiation from epimastigote to metacyclic trypomastigote forms and metacyclic trypomastigote infection. Taken together, these findings indicate the involvement of TcRPA in the metacyclogenesis process and suggest that a delay in cell cycle progression could be linked with differentiation in T. cruzi.
authors.affiliation: Laboratório Especial de Ciclo Celular, Instituto Butantan, São Paulo, São Paulo, Brazil.|Center of Toxins, Immune Response and Cell Signaling-CeTICS, Instituto Butantan, São Paulo, São Paulo, Brazil.
authors.forename: Raphael Souza
authors.initials: RS
authors.lastname: Pavani
authors.affiliation: Laboratório Especial de Ciclo Celular, Instituto Butantan, São Paulo, São Paulo, Brazil.|Center of Toxins, Immune Response and Cell Signaling-CeTICS, Instituto Butantan, São Paulo, São Paulo, Brazil.
authors.forename: Marcelo Santos
authors.initials: MS
authors.lastname: da Silva
authors.affiliation: Departamento de Física e Biofísica, Instituto de Biociências, Universidade Estadual Paulista Júlio de Mesquita Filho -UNESP, Botucatu, São Paulo, Brazil.
authors.forename: Carlos Alexandre Henrique
authors.initials: CA
authors.lastname: Fernandes
authors.affiliation: Instituto Carlos Chagas, Fiocruz-PR, Brazil.
authors.forename: Flavia Souza
authors.initials: FS
authors.lastname: Morini
authors.affiliation: Laboratório Especial de Ciclo Celular, Instituto Butantan, São Paulo, São Paulo, Brazil.|Center of Toxins, Immune Response and Cell Signaling-CeTICS, Instituto Butantan, São Paulo, São Paulo, Brazil.
authors.forename: Christiane Bezerra
authors.initials: CB
authors.lastname: Araujo
authors.affiliation: Departamento de Física e Biofísica, Instituto de Biociências, Universidade Estadual Paulista Júlio de Mesquita Filho -UNESP, Botucatu, São Paulo, Brazil.
authors.forename: Marcos Roberto de Mattos
authors.initials: MR
authors.lastname: Fontes
authors.affiliation: Center of Toxins, Immune Response and Cell Signaling-CeTICS, Instituto Butantan, São Paulo, São Paulo, Brazil.|Laboratório de Imunoquímica, Instituto Butantan, São Paulo, São Paulo, Brazil.
authors.forename: Osvaldo Augusto
authors.initials: OA
authors.lastname: Sant'Anna
authors.affiliation: Departamento de Bioquímica e Imunologia, ICB, Universidade Federal de Minas Gerais, Belo Horizonte, Minas Gerais, Brazil.
authors.forename: Carlos Renato
authors.initials: CR
authors.lastname: Machado
authors.affiliation: Departamento de Genética, Instituto de Biociências, Universidade Estadual Paulista Julio Mesquita Filho-UNESP, Botucatu, São Paulo, Brazil.
authors.forename: Maria Isabel
authors.initials: MI
authors.lastname: Cano
authors.affiliation: Instituto Carlos Chagas, Fiocruz-PR, Brazil.
authors.forename: Stenio Perdigão
authors.initials: SP
authors.lastname: Fragoso
authors.affiliation: Laboratório Especial de Ciclo Celular, Instituto Butantan, São Paulo, São Paulo, Brazil.|Center of Toxins, Immune Response and Cell Signaling-CeTICS, Instituto Butantan, São Paulo, São Paulo, Brazil.
authors.forename: Maria Carolina
authors.initials: MC
authors.lastname: Elias
mesh_terms: D000818:Animals; D002454:Cell Differentiation; D014355:Chagas Disease; D003198:Computer Simulation; D016054:DNA, Protozoan / Q000378:metabolism*; D004277:DNA, Single-Stranded / Q000378:metabolism; D006801:Humans; D056004:Molecular Dynamics Simulation; D011485:Protein Binding; D015800:Protozoan Proteins / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D051716:Replication Protein A / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D014349:Trypanosoma cruzi / Q000235:genetics / Q000502:physiology*
chemical_list: D016054:DNA, Protozoan; D004277:DNA, Single-Stranded; D015800:Protozoan Proteins; D051716:Replication Protein A