Parasite signaling pathways involved in the interaction of Trypanosoma Cruzi with extracellular matrix isolated from distinct cell lines

• Grant number: 17/06717-5
• Support Opportunities: Scholarships in Brazil - Post-Doctoral
• Start date: June 01, 2017
• End date: May 31, 2019
• Field of knowledge: Biological Sciences - Biochemistry - Biochemistry of Microorganisms
• Agreement: Coordination of Improvement of Higher Education Personnel (CAPES)
• Principal Investigator: Maria Julia Manso Alves
• Grantee: Paloma Leão Sousa
• Host Institution: Instituto de Química (IQ). Universidade de São Paulo (USP). São Paulo , SP, Brazil

Abstract

S-nitrosylation is a reversible post-translational modification that has been implicated in the regulation of protein function. We described recently that S-nitrosylation of proteins occurs in T. cruzi trypomastigotes during their interaction with ECM, an obligatory step in the invasion process. Unpublished data (in collaboration with G. Palmisano) showed that proteins from the cytoskeleton (paraflagellar rod protein, alpha and beta-tubulin), phosphatases, kinases and ribosomal proteins, among others, are S-nitrosylated. This project has two main goals: to determine the influence of the ECM composition on the S-nitrosylation (SNO) of T. cruzi proteins and to search for the biological function of this modification. For that purposes, ECMs will be prepared from cell lines selected by their high and low capacity of adhesion to/ invasion by T. cruzi and the composition of both ECMs will be determined by proteomic analysis. To get mechanistic insights on the signaling pathway in ECMs- treated trypomastigotes, NO synthase activity, NO production, cGMP, cAMP, MAPK. PI3K and SNO and phosphorylation profiles in the presence or absence of NO synthase (L-NAME), phosphodiesterase (IBMX) and protein kinase (U0126, LY294002 or wortmanin) will be analyzed. In addition, total protein synthesis and the levels of S-nitrosylated proteins from the paraflagellar rod will be determined in ECMs- treated trypomastigotes under the same experimental conditions. Additionally, proteins associated with the cytoskeleton will be determined by pull down experiments and proteomic analysis.This project intends to understand better the signaling pathways activated in trypomastigotes during their adhesion to ECMs isolated from different cell lines, which may contribute to the understanding of T. cruzi invasion mechanisms. (AU)