Generation of new tools to study the role of aurora kinases in chromatin dynamics in T. cruzi

FASSOLARI, M.; FLAWIÁ, M.M.; TORRES, H.N.; ALONSO, G.

Mar del Plata, Buenos Aires

Congreso; XLIII Reunión Anual de SAIB; 2007

SAIB

Aurora kinases (AUKs) have been implicated in regulating several cellular processes such as chromosome segregation and cell division. Three isoforms of these proteins, aurora A, B, and C, have been identified in mammals. Consistent with their localizations, AUK A regulates spindle assembly, AUK B controls chromosome segregation and cytokinesis initiation and AUK C was found in testis and certain tumor cell lines. Previously, we reported the finding of three AUK homologues (TcAUK1, 2 and 3) in Trypanosoma cruzi. The protein products were obtained by expression in E. coli but we failed to measure the enzymatic activity by using H2AS Histone mix from Sigma and Myelin Basic Protein as substrates. Taking into account the differences between the T. cruzi histones and their orthologs from other eukaryotes, we decided to perform the cloning and expression of the H3, H3 variant and H4 T. cruzi histones in bacterial systems. In addition, to investigate the AUK1 subcellular localization, we are using affinity purified recombinant protein to generate specific antiserum in mouse. Finally, to obtain new knowledge on the function of AUK1 in vivo we transfect epimastigote cells with the pTREX expression vector carrying the coding sequence of this enzyme. These evidences not only point out a conserved pathway along the evolution but also open new insights in drug therapy against Chagas’ disease.