tailieunhanh - Báo cáo khoa học: Protein and mRNA content of TcDHH1-containing mRNPs in Trypanosoma cruzi

In trypanosomatids, the regulation of gene expression occurs mainly at the post-transcriptional level. Previous studies have revealed nontranslated mRNA in theTrypanosoma cruzicytoplasm. Previously, we have identified and cloned the TcDHH1 protein, a DEAD box RNA helicase. It has been reported that Dhh1 is involved in multiple RNA-related processes in vari-ous eukaryotes. | Protein and mRNA content of TcDHHI-containing mRNPs in Trypanosoma cruzi Fabiola B. Holetz1 Lysangela R. Alves1 Christian M. Probst1 Bruno Dallagiovanna1 Fabricio K. Marchini1 Patricio Manque2 3 Gregory Buck2 Marco A. Krieger1 Alejandro Correa1 and Samuel Goldenberg1 1 Institute Carlos Chagas FIOCRUZ Curitiba Brazil 2 Center for the Study of BiologicalComplexity Virginia Commonwealth University Richmond VA USA 3 Instituto de Biotecnologia Universidad Mayor Santiago Chile Keywords differentiation P-bodies stress granules translation control Trypanosoma Correspondence S. Goldenberg Instituto Carlos Chagas FIOCRUZ Rua Professor Algacyr Munhoz Mader 3775 Curitiba 81350-010 PR Brazil Fax 55 41 3316 3230 Tel 55 41 3316 3230 E-mail sgoldenb@ Received 29 January 2010 revised 3 June 2010 accepted 23 June 2010 doi In trypanosomatids the regulation of gene expression occurs mainly at the post-transcriptional level. Previous studies have revealed nontranslated mRNA in the Trypanosoma cruzi cytoplasm. Previously we have identified and cloned the TcDHH1 protein a DEAD box RNA helicase. It has been reported that Dhh1 is involved in multiple RNA-related processes in various eukaryotes. It has also been reported to accumulate in stress granules and processing bodies of yeast animal cells Trypanosoma brucei and T. cruzi. TcDHH1 is localized to discrete cytoplasmic foci that vary depending on the life cycle status and nutritional conditions. To study the composition of mRNPs containing TcDHH1 we carried out immunoprecipitation assays with anti-TcDHH1 using epimastigote lysates. The protein content of mRNPs was determined by MS and pre-immune serum was used as control. We also carried out a ribonomic approach to identify the mRNAs present within the TcDHH1 immunoprecipitated complexes. For this purpose competitive microarray hybridizations were performed against negative controls the nonprecipitated fraction. Our results showed that mRNAs .