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Master's Dissertation
DOI
10.11606/D.42.2008.tde-01042009-102624
Document
Author
Full name
Simone Guedes Calderano
Institute/School/College
Knowledge Area
Date of Defense
Published
São Paulo, 2008
Supervisor
Committee
Sabbaga, Maria Carolina Quartim Barbosa Elias (President)
Faria, Marcella Barbosa
Winter, Carlos Eduardo
Title in Portuguese
Dinâmica molecular no núcleo de Trypanosoma cruzi.
Keywords in Portuguese
Trypanossoma cruzi
ORC
Organização nuclear
PCNA
Replicação
Sítios de replicação
Abstract in Portuguese
Em Trypanosoma cruzi os sítios de replicação estão localizados na periferia nuclear. A fim de entender a dinâmica das moléculas envolvidas na replicação, Orc/Cdc6 foi usado como marcador da maquinaria de pré-replicação e PCNA como marcador da maquinaria de replicação durante o ciclo celular da forma epimastigota. Ambas as moléculas apresentaram dois padrões de localização nuclear: padrão disperso e periférico. Em ensaio de dupla marcação três combinações dos padrões de Orc/Cdc6 e TcPCNA foram encontrados durante G1/S: Orc/Cdc6 periférico e PCNA disperso, ambos dispersos e ambos periféricos. Por meio destes resultados pudemos concluir que durante G1 ambas as moléculas se encontram dispersas. Ao final desta fase, Orc/Cdc6 migra para periferia nuclear enquanto PCNA permanece disperso, migrando para a periferia nuclear quando a célula entra em S, quando a replicação do DNA irá ocorrer. Assim, a replicação na periferia nuclear não se deve à localização prévia das moléculas de replicação nesta região, mas sim à migração destas moléculas para os sítios apropriados.
Title in English
Molecular dynamics at Trypanosoma cruzi nucleus.
Keywords in English
Trypanossoma cruzi
Nuclear organization
ORC
PCNA
Replication
Replication sites
Abstract in English
In Trypanosoma cruzi the replication sites are located at nuclear periphery. In order to analyse the dynamics of molecules involved in replication, Orc/Cdc6 was used as a marker of pre-replication machinery and PCNA as a marker of replication machinery during the cell cycle of epimastigote form. Both molecules presented two nuclear patterns: dispersed pattern and peripheral pattern. Double-labeling assay showed three different patterns of Orc/Cdc6 and PCNA in the nucleus: Orc/Cdc6 at nuclear periphery and PCNA dispersed, both dispersed and both at nuclear periphery. This data allowed us to conclude that during early G1 phase both molecules are dispersed in the nucleus and during late G1 Orc/Cdc6 goes to nuclear periphery while TcPCNA remains dispersed, moving to nuclear periphery, where DNA replication will take place, when S phase starts. Thus, the replication of DNA at nuclear periphery is not due to localization of replications factors at nuclear periphery; instead it depends on the movement of these factors to the appropriated sites.
 
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Publishing Date
2009-04-06
 
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