Sci. STKE, 30 October 2007
Gene Silencing Ras Versus Fas
John F. Foley
Sciences STKE, AAAS, Washington, DC 20005, USA
Cancer cells in which Ras oncogenes are activated often exhibit epigenetic silencing of Fas, leading to the prevention of Fas-ligand-mediated apoptosis. Inactivation of Fas is usually due to hypermethylation at its promoter, but how this silencing is directed is unknown. Gazin et al. transduced K-ras-transformed NIH 3T3 cells that did not express Fas on the cell surface with a library of retroviruses containing small hairpin RNAs (shRNAs) and used immunomagnetic beads to isolate cells that regained surface expression of Fas. The shRNAs that promoted expression of Fas on the cell surface were sequenced, thus identifying 28 target genes, which encoded signaling molecules, regulators of gene expression, and proteins not previously associated with Ras. The abundance of these genes was higher in K-ras-transformed NIH 3T3 cells than in NIH 3T3 cells, as measured by real-time reverse transcription polymerase chain reaction assays. Methylated DNA immunoprecipitation assays showed that although the Fas promoter was hypermethylated in K-ras-transformed NIH 3T3 cells, it was not hypermethylated in any of the 28 cell lines, each of which expressed a single shRNA targeting one of the identified genes. Chromatin immunoprecipitation assays established that the DNA methyltransferase DNMT1 was associated with the Fas promoter in K-ras-transformed NIH-3T3 cells but that this association was decreased in all 28 shRNA-containing cell lines. At least 21 of the 28 genes identified by the screen were also required for the hypermethylation of 5 other genes that are known to be silenced in Ras-transformed cells. Tumors in mice resulting from injection of any of 9 of the K-ras-transformed NIH 3T3 shRNA-containing cell lines were smaller than those in mice injected with K-ras-transformed NIH 3T3 cells. These results suggest that oncogenic Ras does indeed direct epigenetic silencing of Fas (and other genes) through a number of effectors, which might serve as targets for new therapies.
C. Gazin, N. Wajapeyee, S. Gobeil, C.-M. Virbasius, M. R. Green, An elaborate pathway required for Ras-mediated epigenetic silencing. Nature 449, 1073-1077 (2007). [PubMed]
Citation: J. F. Foley, Ras Versus Fas. Sci. STKE 2007, tw390 (2007).
Science Signaling. ISSN 1937-9145 (online), 1945-0877 (print). Pre-2008: Science's STKE. ISSN 1525-8882