title: Retina-derived POU domain factor 1 coordinates expression of genes relevant to renal and neuronal development.
abstract: Retina-derived POU domain Factor 1 (RPF-1), a member of POU transcription factor family, is encoded by POU6F2 gene, addressed by interstitial deletions at chromosome 7p14 in Wilms tumor (WT). Its expression has been detected in developing kidney and nervous system, suggesting an early role for this gene in regulating development of these organs. To investigate into its functions and determine its role in transcriptional regulation, we generated an inducible stable transfectant from HEK293 cells. RPF-1 showed nuclear localization, elevated stability, and transactivation of promoters featuring POU consensus sites, and led to reduced cell proliferation and in vivo tumor growth. By addressing the whole transcriptome regulated by its induction, we could detect a gross alteration of gene expression that is consistent with promoter occupancy predicted by genome-wide Chip-chip analysis. Comparison of bound regulatory regions with differentially expressed genes allowed identification of 217 candidate targets. Enrichment of divergent octamers in predicted regulatory regions revealed promiscuous binding to bipartite POUS and POUH consensus half-sites with intervening spacers. Gel-shift competition assay confirmed the specificity of RPF-1 binding to consensus motifs, and demonstrated that the Ser-rich region upstream of the POU domain is indispensable to achieve DNA-binding. Promoter-reporter activity addressing a few target genes indicated a dependence by RPF-1 on transcriptional response. In agreement with its expression in developing kidney and nervous system, the induced transcriptome appears to indicate a function for this protein in early renal differentiation and neuronal cell fate, providing a resource for understanding its role in the processes thereby regulated.
authors.affiliation: Department of Predictive & Preventive Medicine, Fondazione IRCCS Istituto Nazionale dei Tumori, Italy. Electronic address: antonio.fiorino@istitutotumori.mi.it.
authors.forename: Antonio
authors.initials: A
authors.lastname: Fiorino
authors.affiliation: Department of Predictive & Preventive Medicine, Fondazione IRCCS Istituto Nazionale dei Tumori, Italy.
authors.forename: Giacomo
authors.initials: G
authors.lastname: Manenti
authors.affiliation: Department of Predictive & Preventive Medicine, Fondazione IRCCS Istituto Nazionale dei Tumori, Italy.
authors.forename: Beatrice
authors.initials: B
authors.lastname: Gamba
authors.affiliation: Cogentech, Consortium for Genomic Technologies, IFOM-IEO Campus, Italy.
authors.forename: Gabriele
authors.initials: G
authors.lastname: Bucci
authors.affiliation: Functional Genomic Core Facility, Fondazione IRCCS Istituto Nazionale dei Tumori, Italy.
authors.forename: Loris
authors.initials: L
authors.lastname: De Cecco
authors.affiliation: Department of Predictive & Preventive Medicine, Fondazione IRCCS Istituto Nazionale dei Tumori, Italy.
authors.forename: Michele
authors.initials: M
authors.lastname: Sardella
authors.affiliation: Department of Bioscience, University of Milan, Milan, Italy.
authors.forename: Giacomo
authors.initials: G
authors.lastname: Buscemi
authors.affiliation: Department of Predictive & Preventive Medicine, Fondazione IRCCS Istituto Nazionale dei Tumori, Italy.
authors.forename: Sara
authors.initials: S
authors.lastname: Ciceri
authors.affiliation: Experimental Oncology & Molecular Medicine, Fondazione IRCCS Istituto Nazionale dei Tumori, Italy.
authors.forename: Maria T
authors.initials: MT
authors.lastname: Radice
authors.affiliation: Department of Predictive & Preventive Medicine, Fondazione IRCCS Istituto Nazionale dei Tumori, Italy.
authors.forename: Paolo
authors.initials: P
authors.lastname: Radice
authors.affiliation: Department of Predictive & Preventive Medicine, Fondazione IRCCS Istituto Nazionale dei Tumori, Italy.
authors.forename: Daniela
authors.initials: D
authors.lastname: Perotti
mesh_terms: D021581:Active Transport, Cell Nucleus; D020816:Amino Acid Motifs; D002467:Cell Nucleus / Q000378:metabolism; D049109:Cell Proliferation; D016384:Consensus Sequence; D018507:Gene Expression Regulation, Developmental; D057809:HEK293 Cells; D006801:Humans; D007668:Kidney / Q000254:growth & development*; D009474:Neurons / Q000166:cytology*; D050809:POU Domain Factors / Q000378:metabolism*; D014158:Transcription, Genetic
chemical_list: D050809:POU Domain Factors; C098906:POU6F2 protein, human
keywords: Chromatin immunoprecipitation (ChiP); DNA binding protein; Developmental factor; Gene expression; POU transcription factor; Wilms tumor