The Action Mechanism of Omomyc expression was analyzed via an oligonucleotide array containing probes for approximately 7,000 full-length sequences and 1,000 EST (expressed sequence tags) clusters. Data are accessible in the GEO database with the following accession number: GSE25039. Relative mRNA expression values (Fold Changes) were computed by taking as reference Rat1-control cells at time 0; only genes showing a Fold Change of at least +3 (upregulated genes) or 23 (downregulated genes) were taken into account. At the time of serum stimulation, Omomer expressing and control Rat1 cells displayed a virtually identical gene expression profile (Figure 3, top). At 90 min of serum induction 111 sequences were upregulated and 96 downregulated in Rat1-control cells. Strik- ingly, a pronounced downregulation was found in Rat1-Omomer cells: as many of 516 gene sequences were downregulated and 143 upregulated (Table S1). Overall, the total number of sequences whose expression was up or downregulated in the presence of Omomyc after 90 min of serum stimulation amounted to 8.2% of the probes on the array. Among the sequences downregulated in the presence of Omomyc, 475 represented genes with a known GeneID (gene identifier) and 41 other transcribed regions. To find out whether genes downregulated in the presence of Omomyc were also validated Myc targets we compared them to the set of genes listed in the Myc Cancer Gene database (www.myccancer- gene.org), a collection of Myc responsive genes identified in Figure 3. Omomyc impact on the transcriptional response of Rat1 fibroblasts to serum stimulation. Omomyc promotes downregulation of numerous genes. Top. Many more genes were downregulated in Rat1 cells expressing Omomer (Omomer) than in cells that do not (Control) at 909 following serum stimulation of Rat1 fibroblasts in the presence of tamoxifen. The number of upregulated genes was similar. Rat1-control cells at time 0 were taken as reference. Middle. Distribution in different Biological Process classes – based on Gene Ontology classification – of genes downregulated at 909 following serum stimulation in Omomer and control cells. Bottom. Genes related to nucleotide and nucleic acid metabolism are the most significantly over-represented – as computed by the Panther software – among the genes downregulated in the presence of Omomyc (Omomer cells in the presence of tamoxifen). doi:10.1371/journal.pone.0022284.g003 PLoS ONE | www.plosone.org 5 July 2011 | Volume 6 | Issue 7 | e22284