Fluorescent detection of SignalSilence® Control siRNA (Fluorescein Conjugate) #6201 in living HeLa cells 24 hours post-transfection, demonstrating nearly 100% transfection efficiency.
Western blot analysis of extracts from HeLa cells, untransfected or transfected with either nonspecific control siRNA or eIF4E siRNA. eIF4E was detected using eIF4E Antibody #9742, and eIF4B was detected using eIF4B Antibody #3592. The eIF4E Antibody confirms silencing of eIF4E expression, and the eIF4B Antibody is used to control for loading and siRNA specificity.
CST recommends transfection with 100 nM SignalSilence® eIF4E siRNA I 48-72 hours prior to cell lysis.
SignalSilence® siRNA is supplied in RNAse-free water. Aliquot and store at -20ºC.
SignalSilence® eIF4E siRNA I from Cell Signaling Technology (CST) allows the researcher to specifically inhibit eIF4E expression using RNA interference, a method whereby gene expression can be selectively silenced through the delivery of double stranded RNA molecules into the cell. All SignalSilence® siRNA products are rigorously tested in-house and have been shown to reduce target protein expression by western analysis.
Eukaryotic initiation factor 4E (eIF4E) binds to the mRNA cap structure to mediate the initiation of translation (1,2). eIF4E interacts with eIF4G, a scaffold protein that promotes assembly of eIF4E and eIF4A into the eIF4F complex (2). eIF4B is thought to assist the eIF4F complex in translation initiation. Upon activation by mitogenic and/or stress stimuli mediated by Erk and p38 MAPK, Mnk1 phosphorylates eIF4E at Ser209 in vivo (3,4). Two Erk and p38 MAPK phosphorylation sites in mouse Mnk1 (Thr197 and Thr202) are essential for Mnk1 kinase activity (3). The carboxy-terminal region of eIF4G also contains serum-stimulated phosphorylation sites, including Ser1108, Ser1148, and Ser1192 (5). Phosphorylation at these sites is blocked by the PI3 kinase inhibitor LY294002 and by the FRAP/mTOR inhibitor rapamycin.
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