Investigation of differential expression proteins and post-translational histones modifications during the Trypanosoma cruzi cell cycle and identification of possible candidates involved in the differentiation process

• Grant number: 23/03159-2
• Support Opportunities: Scholarships in Brazil - Post-Doctoral
• Start date until: November 01, 2023
• End date until: May 31, 2024
• Field of knowledge: Biological Sciences - Parasitology - Protozoology of Parasites
• Principal Investigator: Maria Carolina Quartim Barbosa Elias Sabbaga
• Grantee: Ana Paula de Jesus Menezes
• Host Institution: Instituto Butantan. Secretaria da Saúde (São Paulo - Estado). São Paulo , SP, Brazil
• Associated research grant: 20/00694-6 - How DNA replication contributes for the success of infection caused by Trypanosoma cruzi, AP.TEM

Abstract

In this proposal, we intend to improve the biological knowledge of Trypanosoma cruzi, which causes Chagas disease, focusing on the correlation between cell cycle, differentiation and epigenetics. Throughout the T.cruzi life cycle, different life forms are found in the invertebrate vector and the mammalian host. These forms adapted to distinct environmental conditions and thus differ in terms of morphology, gene expression, metabolic profile and replicative capacity. During the differentiation journey, the parasite undergoes significant changes in chromatin. These changes can be caused by epigenetic marks, such as post-translational modifications (PTMs) of histones, which enable cells to acquire new biological features for adapting to environmental changes. This peculiar connection is relevant for all organisms but underestimated in digenetic parasites such as T. cruzi. Thus, the main goal of this proposal is to analyze the data already generated from the nuclear proteome and histone PTMs in the different phases of the cell cycle. Next, we intend to confirm preliminary data that suggest a G1 differentiation window. Parasites were synchronized using 20 mM hydroxyurea to obtain cells in G1/S, S and G2/M in biological quadruplicates. We extracted the cell's nucleus and analyzed the nuclear proteome quantitatively by LC-MS/MS using TMT labelling. We identified more than 2,500 proteins, and 690 were quantified. The enrichment of the gene ontology (GO) terms associated with the nucleus, chromatin and chromosome confirms the nuclear enrichment. Regarding the analysis of histone PTMs, we found an increase in H4K14ac and H3K76me1 in G2/M, corroborating the data already described. Taken together, our results indicated the reliability and reproducibility of our proteomic assay. Nuclear proteins constitute a highly organized but complex network that plays diverse roles during biological processes such as the cell cycle. We hope, with this proposal, to improve the knowledge of nuclear proteins and histone PTMs differentially expressed during the cell cycle and, above all, to find proteins and histone PTMs involved in the differentiation process. (AU)