title: Regulation of glycogen metabolism by the CRE-1, RCO-1 and RCM-1 proteins in Neurospora crassa. The role of CRE-1 as the central transcriptional regulator.
abstract: The transcription factor CreA/Mig1/CRE-1 is a repressor protein that regulates the use of alternative carbon sources via a mechanism known as Carbon Catabolite Repression (CCR). In Saccharomyces cerevisiae, Mig1 recruits the complex Ssn6-Tup1, the Neurospora crassa RCM-1 and RCO-1 orthologous proteins, respectively, to bind to promoters of glucose-repressible genes. We have been studying the regulation of glycogen metabolism in N. crassa and the identification of the RCO-1 corepressor as a regulator led us to investigate the regulatory role of CRE-1 in this process. Glycogen content is misregulated in the rco-1(KO), rcm-1(RIP) and cre-1(KO) strains, and the glycogen synthase phosphorylation is decreased in all strains, showing that CRE-1, RCO-1 and RCM-1 proteins are involved in glycogen accumulation and in the regulation of GSN activity by phosphorylation. We also confirmed the regulatory role of CRE-1 in CCR and its nuclear localization under repressing condition in N. crassa. The expression of all glycogenic genes is misregulated in the cre-1(KO) strain, suggesting that CRE-1 also controls glycogen metabolism by regulating gene expression. The existence of a high number of the Aspergillus nidulans CreA motif (5'-SYGGRG-3') in the glycogenic gene promoters led us to analyze the binding of CRE-1 to some DNA motifs both in vitro by DNA gel shift and in vivo by ChIP-qPCR analysis. CRE-1 bound in vivo to all motifs analyzed demonstrating that it down-regulates glycogen metabolism by controlling gene expression and GSN phosphorylation.
authors.affiliation: Departamento de Bioquímica e Tecnologia Química, Instituto de Química, Universidade Estadual Paulista, UNESP, 14800-060 Araraquara, SP, Brazil.
authors.forename: Fernanda Barbosa
authors.initials: FB
authors.lastname: Cupertino
authors.affiliation: Departamento de Bioquímica e Tecnologia Química, Instituto de Química, Universidade Estadual Paulista, UNESP, 14800-060 Araraquara, SP, Brazil.
authors.forename: Stela
authors.initials: S
authors.lastname: Virgilio
authors.affiliation: Departamento de Bioquímica e Tecnologia Química, Instituto de Química, Universidade Estadual Paulista, UNESP, 14800-060 Araraquara, SP, Brazil.
authors.forename: Fernanda Zanolli
authors.initials: FZ
authors.lastname: Freitas
authors.affiliation: Departamento de Bioquímica e Tecnologia Química, Instituto de Química, Universidade Estadual Paulista, UNESP, 14800-060 Araraquara, SP, Brazil.
authors.forename: Thiago de Souza
authors.initials: Tde S
authors.lastname: Candido
authors.affiliation: Departamento de Bioquímica e Tecnologia Química, Instituto de Química, Universidade Estadual Paulista, UNESP, 14800-060 Araraquara, SP, Brazil. Electronic address: mcbertol@iq.unesp.br.
authors.forename: Maria Célia
authors.initials: MC
authors.lastname: Bertolini
mesh_terms: D051698:Activating Transcription Factor 2 / Q000378:metabolism*; D002244:Carbon / Q000378:metabolism; D005656:Fungal Proteins / Q000378:metabolism*; D006003:Glycogen / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*; D006006:Glycogen Synthase / Q000378:metabolism; D009154:Mutation; D009492:Neurospora crassa / Q000235:genetics / Q000378:metabolism*; D010766:Phosphorylation; D011401:Promoter Regions, Genetic; D012097:Repressor Proteins / Q000378:metabolism*; D014157:Transcription Factors / Q000378:metabolism*
chemical_list: D051698:Activating Transcription Factor 2; D005656:Fungal Proteins; C102112:RCO-1 protein, Neurospora crassa; D012097:Repressor Proteins; D014157:Transcription Factors; D002244:Carbon; D006003:Glycogen; D006006:Glycogen Synthase
keywords: CRE-1; ChIP-qPCR; Gene expression; Glycogen; Neurospora crassa