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TC TLF-1: A PUTATIVE TRANSCRIPTION REGULATION FACTOR IN TRYPANOSOMA CRUZI. Cribb, Pamela; Zelus, Dominique and Serra, Esteban. Instituto de Biología Molecular y Celular de Rosario. Suipacha 531. 2000. Rosario. pcribb@fbioyf.unr.edu.ar TATA-binding protein (TBP) is an ubiquitous transcription factor that takes part of Transcription Initiation Complex (IC) of all type-I, -II and -III eukariotic RNA polymerases. In the last years, a regulatory function of IC, via regulatory TAFs and alternative TBP-like factors (TLFs), was established in yeasts and metazoans. Using degenerated oligonucleotides a TLF from Trypanosoma cruzi was cloned by PCR (TcTLF-1). TcTLF-1 is coded by an unique gene in all T. cruzi strains assayed, being highly conserved at the sequence level among all them. Deduced TcTLF-1 sequence showed low, but significant identity with all TBPs, TLFs and archaeal TBPs. The coding sequence for TcTLF-1 was cloned in expression vectors (pGEX and pQE) providing recombinant proteins both as a fusion to GST and with Histidine tail. Polyclonal antibodies against recombinant protein were used in Western Blot assays indicating that the protein is present in both epimastigotes and trypomastigotes, although in different amounts. TcTLF-1 showed binding activity to the SL RNA gene promoter, the unique RNA pol II-dependent promoter identified in trypanosomatids in electrophoretic mobility shift assays. Moreover, Selex assays showed that recombinant TcTLF-1 binds preferably to a consensus sequence rich in G+C, similar to those present in the SL RNA gene promoter. Since no RNApol II-dependent transcriptional regulation mechanism has been described in trypanosomatids to date, we propose that gene expression may be regulated at the transcriptional level by TcTLF-1 through the IC. Area temática: Parasitología Comunicación Oral

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