Three immunoblots were performed with anti-Fra-1 sera and lysates of Scp-2 cells treated with the indicated siRNAs as with figure 5B. ideals were divided S100A4 from the Renilla luciferase ideals (from cotransfected plasmid pRLSV40P). In (C) the ideals were scaled to an average of 1.0 for the Scp-2 cells and the -819 to +71 MMP1 reporter, except for CMV which was scaled to its own ideals for Scp-2 cells. In (E) the ideals were scaled in each replicate to 1 1.0 for Scp-2 cells and the -172/-27 MMP1 reporter. Data file 3. Data for numbers 3B and D. Relative luciferase activity is definitely demonstrated for three replicates. Each replicate is the average of duplicate determinations (i.e. six total points). The firefly luciferase ideals were divided from the Renilla luciferase ideals (from cotransfected plasmid pRLSV40P). The ideals were scaled in each replicate to 1 1.0 for Scp-2 cells and the wild type reporter. For LDC000067 the synthetic reporter, the ideals were scaled to that of the 3X-AP1 site reporter. Data file 4. Data for numbers 4A and C. A) Relative mRNA expression from your indicated cell lines and genes is definitely demonstrated for three replicates from self-employed mRNA isolations. The figures reflect qPCR Ct figures using 18S rRNA Ct ideals. The ideals were scaled to a value of 1 1.0 for Scp-2 cells. The primers used are demonstrated in Materials and Methods. C) Relative Fra-1 protein expression. Three immunoblots were performed with anti-Fra-1 sera and lysates of the indicated cell lines as with number 4B. The Fra-1 band intensities were quantified using the Odyssey infrared imager (Li-Cor) and the LDC000067 software provided by the manufacturer. The intensities were scaled to the average ideals for Fra-1 protein in Scp-2 cells. Data file 5. Data for numbers 5A, C and D. A) Relative Fra-1 mRNA manifestation in Scp-2 cells treated with the indicated siRNAs. Manifestation determined by qPCR is demonstrated for three replicates from self-employed mRNA isolations. The figures reflect qPCR delta Ct figures using 18S rRNA Ct ideals. The ideals were scaled to an average value of 1 1.0 for untreated Scp-2 cells. The primers used are demonstrated in Materials and Methods. B) Relative Fra-1 protein manifestation. Three immunoblots were performed with anti-Fra-1 sera and lysates of Scp-2 cells treated with the indicated siRNAs as with number 5B. The Fra-1 band intensities were quantified using the Odyssey infrared imager (Li-Cor) and the software provided by the manufacturer. The ideals were scaled to the average intensities of Fra-1 protein in Scp-2 cells. C) As with (A) exept that MMP-1 and GAPDH mRNA manifestation were measured by qPCR. The primers used are demonstrated LDC000067 in materials and methods. Data file 6. Data for number 6C. Chromatin immunoprecipitation from your indicated cell lines and using anti-Fra1 or control (no antibody) was quantified by qPCR using the indicated primers. The binding ideals were divided by those of input DNA and are demonstrated as percent of input. Ideals from three replicate chromatin immunoprecipitations are demonstrated. Data file 7. Data for numbers 7B and D. B) Scp-2 or Scp-21 cells were treated with the protein synthesis inhibitor cycloheximide for the indictate occasions. The cell lysates were then immunoblotted with anti-Fra-1 antibodies as with number 7A. The Fra-1 protein band intensities were quantified within the Odyssey infrared imager (Li-Cor). The ideals for three self-employed experiments are demonstrated. The ideals.