Genome-wide Transcriptional Analysis of Tetrahymena thermophila Response to Exogenous Cholesterol

The ciliate Tetrahymena thermophila does not require sterols for growth and synthesizes pentacyclic triterpenoid alcohols, mainly tetrahymanol, as sterol surrogates. However, when sterols are present in the environment, T. thermophila efficiently incorporates and modifies them. These modifications consist of desaturation reactions at positions C5(6), C7(8), and C22(23), and de-ethylation at C24 of 29-carbon sterols (i.e. phytosterols). Three out of four of the enzymes involved in the sterol modification pathway have been previously identified. However, identification of the sterol C22 desaturase remained elusive, as did other basic aspects of this metabolism. To get more insights into this peculiar metabolism, we here perform a whole transcriptome analysis of T. thermophila in response to exogenous cholesterol. We found 356 T. thermophila genes to be differentially expressed after supplementation with cholesterol for 2 h. Among those that were upregulated, we found two genes belonging to the long spacing family of desaturases that we tentatively identified by RNAi analysis as sterol C22 desaturases. Additionally, we determined that the inhibition of tetrahymanol synthesis after supplementation with cholesterol occurs by a transcriptional downregulation of genes involved in squalene synthesis and cyclization. Finally, we identified several uncharacterized genes that are likely involved in sterols transport and signaling.Fil: Najle, Sebastián Rodrigo. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Rosario. Instituto de Biología Molecular y Celular de Rosario. Universidad Nacional de Rosario. Facultad de Ciencias Bioquímicas y Farmacéuticas. Instituto de Biología Molecular y Celular de Rosario; Argentina. Universitat Pompeu Fabra; España. Consejo Superior de Investigaciones Científicas; EspañaFil: Hernandez, Josefina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Rosario. Instituto de Biología Molecular y Celular de Rosario. Universidad Nacional de Rosario. Facultad de Ciencias Bioquímicas y Farmacéuticas. Instituto de Biología Molecular y Celular de Rosario; ArgentinaFil: Ocaña Pallarès, Eduard. Universitat Pompeu Fabra; España. Consejo Superior de Investigaciones Científicas; EspañaFil: García Siburu, Nicolás Pablo. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Rosario. Instituto de Biología Molecular y Celular de Rosario. Universidad Nacional de Rosario. Facultad de Ciencias Bioquímicas y Farmacéuticas. Instituto de Biología Molecular y Celular de Rosario; ArgentinaFil: Nusblat, Alejandro David. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Nanobiotecnología. Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica. Instituto de Nanobiotecnología; ArgentinaFil: Nudel, Berta Clara. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Nanobiotecnología. Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica. Instituto de Nanobiotecnología; ArgentinaFil: Slamovits, Claudio H.. Dalhousie University Halifax; CanadáFil: Uttaro, Antonio Domingo. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Rosario. Instituto de Biología Molecular y Celular de Rosario. Universidad Nacional de Rosario. Facultad de Ciencias Bioquímicas y Farmacéuticas. Instituto de Biología Molecular y Celular de Rosario; Argentin

CNBP modulates the transcription of Wnt signalling pathway components.

Background: Cellular nucleic acid binding protein (CNBP) is a small and highly conserved protein with nucleic acid chaperone activity that binds single-stranded nucleic acids. Data collected so far suggests that CNBP is required for proper craniofacial development. Despite the advances achieved in the last decade, the identity of the molecular targets of CNBP responsible for its role in rostral head development remains elusive. Methods: In this work we used the CNBP single-stranded DNA-consensus binding sequence to find out putative CNBP target genes present in the human, mouse, chicken, Xenopus and zebrafish genomes. Results: Most of the identified genes are associated with embryonic developmental processes, being three of them (cdk14, ptk7 and tcf7l2) members of the Wnt signaling pathway. This finding, along with previous one showing that CNBP down-regulates the transcription of Wnt5, aimed our work to address the role of CNBP on the WNT signaling players and pathway regulation. Experiments carried out in zebrafish developing embryos revealed that craniofacial morphology was more adversely affected as CNBP abundance decreased. Furthermore, we observed that CNBP up-regulated in a dose-dependent fashion the transcription of cdk14, ptk7 and tcf7l2, which in turn was reflected in c-myc, ccnd1 and axin2 expression. Conclusions: Results reveal a role of CNBP in transcriptional control of components of the Wnt signaling pathway, which might explain its requirement for proper craniofacial development.Fil: Margarit, Ezequiel. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Rosario. Instituto de Biología Molecular y Celular de Rosario; ArgentinaFil: Armas, Pablo. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Rosario. Instituto de Biología Molecular y Celular de Rosario; ArgentinaFil: García Siburu, Nicolás Pablo. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Rosario. Instituto de Biología Molecular y Celular de Rosario; ArgentinaFil: Calcaterra, Nora Beatriz. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Rosario. Instituto de Biología Molecular y Celular de Rosario; Argentin