Functional characterization of giant protein in Trypanosoma brucei

• Grant number: 12/22129-2
• Support Opportunities: Scholarships in Brazil - Doctorate
• Start date: January 01, 2013
• End date: December 31, 2016
• Field of knowledge: Biological Sciences - Parasitology - Protozoology of Parasites
• Principal Investigator: Munira Muhammad Abdel Baqui
• Grantee: Bernardo Pereira Moreira
• Host Institution: Faculdade de Medicina de Ribeirão Preto (FMRP). Universidade de São Paulo (USP). Ribeirão Preto , SP, Brazil

Abstract

Trypanosoma brucei is a unicellular protozoa of great medical relevance since it is the etiological agent of sleeping sickness (African trypanosomiasis). Over the past decades, this organism has been used as experimental model for cellular, biochemical and molecular studies considering its particular structures as nucleus, cytoskeleton and the presence of unique organelles as kinetoplast and flagellum. In all genus of the Trypanosomatidae family, giant proteins were described as a novel class of high molecular mass phosphoproteins (1000-4000kDa) which, besides its structural function, might play a role in the organization and regulation of cytoskeleton and its constituents. However, factors that interact with these proteins in both the cytoskeleton and the host are still unknown and little is known about the giant proteins of the pathogenic trypanosomatids and their functions. Based on what has been described, this project aims to determine the functional role of T. brucei giant protein together with the biochemical and immunocytochemical characterization of this protein. The cell fraction containing the cytoskeleton proteins, including the giant protein, were resolved by SDS-PAGE. Then the slice relative to this protein was cut from the gel and injected in Balb/c mice to produce specific antibody. The polyclonal serum recognized the giant protein and immunofluorescence preliminary observations suggest this protein is localized along the flagellum and in the flagellar pocket. Moreover, mass spectrometry will be performed in order to determine the partial sequence, which will be used to identify the respective gene in T. brucei genome, what allow us to investigate the function of specific catalytic domains. Further, immunoprecipitation and pull-down assays will be performed in order to define which putative partners interact with the giant protein. Furthermore RNAi knockdown combined with confocal and electron microscopy will validate the giant protein function.

Scientific publications

(References retrieved automatically from Web of Science and SciELO through information on FAPESP grants and their corresponding numbers as mentioned in the publications by the authors)

MOREIRA, BERNARDO P.; FONSECA, CAROL K.; HAMMARTON, TANSY C.; BAQUI, MUNIRA M. A.. Giant FAZ10 is required for flagellum attachment zone stabilization and furrow positioning in Trypanosoma brucei. Journal of Cell Science, v. 130, n. 6, p. 1179-1193, MAR 15 2017. (10/19547-1, 12/22129-2)

Academic Publications

(References retrieved automatically from State of São Paulo Research Institutions)

MOREIRA, Bernardo Pereira. Cellular and molecular characterization of Trypanosoma brucei cytoskeletal giant proteins. 2016. Doctoral Thesis - Universidade de São Paulo (USP). Faculdade de Medicina de Ribeirão Preto (PCARP/BC) Ribeirão Preto.