title: Reverse engineering a hierarchical regulatory network downstream of oncogenic KRAS.
abstract: RAS mutations are highly relevant for progression and therapy response of human tumours, but the genetic network that ultimately executes the oncogenic effects is poorly understood. Here, we used a reverse-engineering approach in an ovarian cancer model to reconstruct KRAS oncogene-dependent cytoplasmic and transcriptional networks from perturbation experiments based on gene silencing and pathway inhibitor treatments. We measured mRNA and protein levels in manipulated cells by microarray, RT-PCR and western blot analysis, respectively. The reconstructed model revealed complex interactions among the transcriptional and cytoplasmic components, some of which were confirmed by double pertubation experiments. Interestingly, the transcription factors decomposed into two hierarchically arranged groups. To validate the model predictions, we analysed growth parameters and transcriptional deregulation in the KRAS-transformed epithelial cells. As predicted by the model, we found two functional groups among the selected transcription factors. The experiments thus confirmed the predicted hierarchical transcription factor regulation and showed that the hierarchy manifests itself in downstream gene expression patterns and phenotype.
authors.affiliation: Laboratory of Molecular Tumour Pathology, Institute of Pathology, Charité Universitätsmedizin Berlin, Berlin, Germany.
authors.forename: Iwona
authors.initials: I
authors.lastname: Stelniec-Klotz
authors.forename: Stefan
authors.initials: S
authors.lastname: Legewie
authors.forename: Oleg
authors.initials: O
authors.lastname: Tchernitsa
authors.forename: Franziska
authors.initials: F
authors.lastname: Witzel
authors.forename: Bertram
authors.initials: B
authors.lastname: Klinger
authors.forename: Christine
authors.initials: C
authors.lastname: Sers
authors.forename: Hanspeter
authors.initials: H
authors.lastname: Herzel
authors.forename: Nils
authors.initials: N
authors.lastname: Blüthgen
authors.forename: Reinhold
authors.initials: R
authors.lastname: Schäfer
mesh_terms: D000704:Analysis of Variance; D000818:Animals; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D002471:Cell Transformation, Neoplastic / Q000235:genetics / Q000378:metabolism; D004847:Epithelial Cells / Q000187:drug effects / Q000378:metabolism / Q000473:pathology; D005260:Female; D015972:Gene Expression Regulation, Neoplastic / Q000502:physiology*; D053263:Gene Regulatory Networks / Q000502:physiology*; D011905:Genes, ras; D025743:HMGA2 Protein / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D006801:Humans; D000075723:Kruppel-Like Factor 6; D051741:Kruppel-Like Transcription Factors / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D046228:Microarray Analysis; D008954:Models, Biological; D010051:Ovarian Neoplasms / Q000235:genetics* / Q000378:metabolism; D010053:Ovary / Q000187:drug effects / Q000473:pathology; D011518:Proto-Oncogene Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D016760:Proto-Oncogene Proteins c-fos / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D016283:Proto-Oncogene Proteins p21(ras); D034741:RNA, Small Interfering / Q000378:metabolism / Q000494:pharmacology; D051381:Rats; D020133:Reverse Transcriptase Polymerase Chain Reaction; D015398:Signal Transduction; D014157:Transcription Factors / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D018631:ras Proteins / Q000235:genetics / Q000378:metabolism*; D000099169:Fos-Related Antigen 1
chemical_list: D025743:HMGA2 Protein; D000075723:Kruppel-Like Factor 6; D051741:Kruppel-Like Transcription Factors; D011518:Proto-Oncogene Proteins; D016760:Proto-Oncogene Proteins c-fos; D016283:Proto-Oncogene Proteins p21(ras); D034741:RNA, Small Interfering; D014157:Transcription Factors; D018631:ras Proteins; D000099169:Fos-Related Antigen 1; C105465:KLF6 protein, human; C117307:KRAS protein, human