Confocal immunofluorescent analysis of HeLa cells, mock-transfected (left) or transfected with SignalSilence® GSK-3α/β siRNA (right), using GSK-3-alpha/beta (D75D3) XP® Rabbit mAb #5676 (green). Actin filaments have been labeled wth DY-554 phalloidin (red). Blue pseudocolor = DRAQ5® #4084 (fluorescent DNA dye).
Western blot analysis of extracts from HeLa cells, untransfected or transfected with GSK-3a/β siRNA. GSK-3β was detected using GSK-3β Antibody #9332, and Bcl-xL was detected using Bcl-xL Antibody #2762. The GSK-3β Antibody confirms silencing of GSK-3β expression, and the Bcl-xL Antibody is used to control for loading and siRNA specificity.
Western blot analysis of extracts from HeLa cells, untransfected or transfected with either GSK-3alpha/beta siRNA or GSK-3alpha siRNA. GSK-3alpha and GSK-3beta were detected using a GSK-3alpha/beta antibody, and eIF4B was detected using eIF4B Antibody #3592. The GSK-3alpha/beta antibody confirms silencing of GSK-3alpha and beta expression, and the eIF4B Antibody is used to control for loading and siRNA specificity.
CST recommends transfection with 50 nM GSK-3alpha/beta siRNA 48 hours prior to cell lysis. See Protocol for transfection procedure.
GSK-3α/β siRNA is supplied in RNAse-free water. Aliquot and store at -20ºC.
SignalSilence® GSK-3α/β siRNA from Cell Signaling Technology allows the researcher to specifically inhibit GSK-3aα and GSK-3bβ expression using RNA interference, a method in which 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 protein expression in specified cell lines.
Glycogen synthase kinase-3 (GSK-3) was initially identified as an enzyme that regulates glycogen synthesis in response to insulin (1). GSK-3 is a ubiquitously expressed serine/threonine protein kinase that phosphorylates and inactivates glycogen synthase. GSK-3 is a critical downstream element of the PI3K/Akt cell survival pathway whose activity can be inhibited by Akt-mediated phosphorylation at Ser21 of GSK-3α and Ser9 of GSK-3β (2,3). GSK-3 has been implicated in the regulation of cell fate in Dictyostelium and is a component of the Wnt signaling pathway required for Drosophila, Xenopus, and mammalian development (4). GSK-3 has been shown to regulate cyclin D1 proteolysis and subcellular localization (5).
Small Interfering RNA (siRNA) has been used to specifically silence GSK-3alpha and beta expression, causing an upregulation of beta-catenin protein levels (6).
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Cell Signaling Technology is a trademark of Cell Signaling Technology, Inc.
SignalSilence is a registered trademark of Cell Signaling Technology, Inc.
DRAQ5 is a registered trademark of Biostatus Limited.
Limited Use Label License, RNA interference: This product is licensed under European Patent 1144623 and foreign equivalents from Ribopharma AG, Kulmbach, Germany and is provided only for use in non-commercial research specifically excluding use (a) in drug discovery or drug development, including target identification or target validation, by or on behalf of a commercial entity, (b) for contract research or commercial screening services, (c) for the production or manufacture of siRNA-related products for sale, or (d) for the generation of commercial databases for sale to Third Parties. Information about licenses for these and other commercial uses is available from Ribopharma AG, Fritz-Hornschuch-Str. 9, D-95326 Kulmbach, Germany.