title: Minichromosome Maintenance Complex Is a Critical Node in the miR-183 Signaling Network of MYCN-Amplified Neuroblastoma Cells.
abstract: MYCN and HDAC2 jointly repress the transcription of tumor suppressive miR-183 in neuroblastoma. Enforced miR-183 expression induces neuroblastoma cell death and inhibits xenograft growth in mice. Here we aimed to focus more closely on the miR-183 signaling network using a label-free mass spectrometric approach. Analysis of neuroblastoma cells transfected with either control or miR-183 expression vectors identified 85 differentially expressed proteins. All six members of the minichromosome maintenance (MCM) complex, which is indispensable for initiation and elongation during DNA replication and transcriptionally activated by MYCN in neuroblastoma, emerged to be down-regulated by miR-183. Subsequent annotation category enrichment analysis revealed a ∼14-fold enrichment in the "MCM" protein module category, which highlighted this complex as a critical node in the miR-183 signaling network. Down-regulation was confirmed by Western blotting. MCMs 2-5 were predicted by in silico methods as direct miR-183 targets. Dual-luciferase reporter gene assays with 3'-UTR constructs of the randomly selected MCMs 3 and 5 experimentally confirmed them as direct targets of miR-183. Our results reveal the MCM complex to be a critical and directly regulated node within the miR-183 signaling network in MYCN-amplified neuroblastoma cells.
authors.affiliation: Department of Pediatric Hematology/Oncology/Stem Cell Transplantation Charité - Universitätsmedizin Berlin , Campus Virchow-Klinikum, Augustenburger Platz 1, 13353 Berlin, Germany.
authors.forename: Marco
authors.initials: M
authors.lastname: Lodrini
authors.affiliation: Molecular Proteomics Laboratory, Biological Medical Research Centre, Heinrich-Heine-University Düsseldorf , Universitätsstraße 1, 40225 Düsseldorf, Germany.
authors.forename: Gereon
authors.initials: G
authors.lastname: Poschmann
authors.affiliation: Clinical Cooperation Unit Pediatric Oncology, German Cancer Research Center (DKFZ) and German Consortium for Translational Cancer Research (DKTK), INF 280 , 69120 Heidelberg, Germany.
authors.forename: Victoria
authors.initials: V
authors.lastname: Schmidt
authors.affiliation: Department of Pediatric Hematology/Oncology/Stem Cell Transplantation Charité - Universitätsmedizin Berlin , Campus Virchow-Klinikum, Augustenburger Platz 1, 13353 Berlin, Germany.
authors.forename: Jasmin
authors.initials: J
authors.lastname: Wünschel
authors.affiliation: Division Neuroblastoma Genetics, DKFZ, INF 280 , 69120 Heidelberg, Germany.
authors.forename: Daniel
authors.initials: D
authors.lastname: Dreidax
authors.affiliation: Clinical Cooperation Unit Pediatric Oncology, German Cancer Research Center (DKFZ) and German Consortium for Translational Cancer Research (DKTK), INF 280 , 69120 Heidelberg, Germany.|Center for Individualized Pediatric Oncology (ZIPO) and Brain Tumors, Department of Pediatric Hematology/Oncology, University of Heidelberg and National Center for Tumor Diseases (NCT), INF 430 , 69120 Heidelberg, Germany.
authors.forename: Olaf
authors.initials: O
authors.lastname: Witt
authors.affiliation: Division of Theoretical Systems Biology, DKFZ, INF 280 , 69120 Heidelberg, Germany.
authors.forename: Thomas
authors.initials: T
authors.lastname: Höfer
authors.affiliation: Leibniz-Institut für Analytische Wissenschaften - ISAS - e.V., Bunsen-Kirchhoff-Str. 11, 44139 Dortmund, Germany.
authors.forename: Helmut E
authors.initials: HE
authors.lastname: Meyer
authors.affiliation: Molecular Proteomics Laboratory, Biological Medical Research Centre, Heinrich-Heine-University Düsseldorf , Universitätsstraße 1, 40225 Düsseldorf, Germany.|Institute for Molecular Medicine, University Hospital Düsseldorf , Universitätsstraße 1, 40225 Düsseldorf, Germany.
authors.forename: Kai
authors.initials: K
authors.lastname: Stühler
authors.affiliation: Department of Pediatric Hematology/Oncology/Stem Cell Transplantation Charité - Universitätsmedizin Berlin , Campus Virchow-Klinikum, Augustenburger Platz 1, 13353 Berlin, Germany.
authors.forename: Angelika
authors.initials: A
authors.lastname: Eggert
authors.affiliation: Department of Pediatric Hematology/Oncology/Stem Cell Transplantation Charité - Universitätsmedizin Berlin , Campus Virchow-Klinikum, Augustenburger Platz 1, 13353 Berlin, Germany.|Clinical Cooperation Unit Pediatric Oncology, German Cancer Research Center (DKFZ) and German Consortium for Translational Cancer Research (DKTK), INF 280 , 69120 Heidelberg, Germany.|Center for Individualized Pediatric Oncology (ZIPO) and Brain Tumors, Department of Pediatric Hematology/Oncology, University of Heidelberg and National Center for Tumor Diseases (NCT), INF 430 , 69120 Heidelberg, Germany.|Junior Neuroblastoma Research Group, Experimental and Clinical Research Center of the Max-Delbrück Center for Molecular Medicine and the Charité - Universitätsmedizin Berlin , Lindenberger Weg 80, 13125 Berlin, Germany.
authors.forename: Hedwig E
authors.initials: HE
authors.lastname: Deubzer
mesh_terms: D045744:Cell Line, Tumor; D015536:Down-Regulation; D006801:Humans; D013058:Mass Spectrometry; D035683:MicroRNAs / Q000378:metabolism / Q000494:pharmacology*; D064126:Minichromosome Maintenance Complex Component 3 / Q000187:drug effects / Q000378:metabolism; D064150:Minichromosome Maintenance Complex Component 5 / Q000187:drug effects / Q000378:metabolism; D064110:Minichromosome Maintenance Proteins / Q000378:metabolism*; D000071447:N-Myc Proto-Oncogene Protein / Q000502:physiology*; D009447:Neuroblastoma / Q000737:chemistry / Q000378:metabolism / Q000473:pathology*; D015398:Signal Transduction; D014162:Transfection; D015854:Up-Regulation
chemical_list: C533449:MIRN183 microRNA, human; C459801:MYCN protein, human; D035683:MicroRNAs; D000071447:N-Myc Proto-Oncogene Protein; D064126:Minichromosome Maintenance Complex Component 3; D064150:Minichromosome Maintenance Complex Component 5; D064110:Minichromosome Maintenance Proteins
keywords: cell cycle progression; dual-luciferase reporter gene assay; eukaryotic genome replication; genomic integrity; label-free proteomics; oncogene; systems biology