Product Pathways - Cytoskeletal Signaling
SignalSilence® PAK1 siRNA I (Mouse Specific) #8093
Species cross-reactivity is determined by western blot. Species enclosed in parentheses are predicted to react based on 100% sequence homology.
Western blot analysis of extracts from NIH/3T3 cells, transfected with 100 nM SignalSilence® Control siRNA (Unconjugated) #6568 (-) or SignalSilence® PAK1 siRNA I (Mouse Specific) (+), using PAK1 Antibody #2602 (upper) or α-Tubulin (11H10) Rabbit mAb #2125 (lower). The PAK1 Antibody confirms silencing of PAK1 expression, while the α-Tubulin (11H10) Rabbit mAb is used as a loading control.
SignalSilence® PAK1 siRNA I (Mouse Specific) from Cell Signaling Technology (CST) allows the researcher to specifically inhibit PAK1 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 from CST are rigorously tested in-house and have been shown to reduce target protein expression by western analysis.
Oligonucleotide synthesis is monitored base by base through trityl analysis to ensure appropriate coupling efficiency. The oligo is subsequently purified by affinity-solid phase extraction. The annealed RNA duplex is further analyzed by mass spectrometry to verify the exact composition of the duplex. Each lot is compared to the previous lot by mass spectrometry to ensure maximum lot-to-lot consistency.
Directions for Use
CST recommends transfection with 100 nM SignalSilence® PAK1 siRNA I (Mouse Specific) 48 to 72 hours prior to cell lysis. For transfection procedure, follow protocol provided by the transfection reagent manufacturer. Please feel free to contact CST with any questions on use.
Each vial contains the equivalent of 100 transfections, which corresponds to a final siRNA concentration of 100 nM per transfection in a 24-well plate with a total volume of 300 μl per well.
The p21-activated kinase (PAK) family of serine/threonine kinases is engaged in multiple cellular processes, including cytoskeletal reorganization, MAPK signaling, apoptotic signaling, control of phagocyte NADPH oxidase, and growth factor-induced neurite outgrowth (1,2). Several mechanisms that induce PAK activity have been reported. Binding of Rac/Cdc42 to the CRIB (or PBD) domain near the amino terminus of PAK causes autophosphorylation and conformational changes in PAK (1). Phosphorylation of PAK1 at Thr423 by PDK induces activation of PAK1 (3). Several autophosphorylation sites have been identified, including Ser199 and Ser204 of PAK1 and Ser192 and Ser197 of PAK2 (4,5). Because the autophosphorylation sites are located in the amino-terminal inhibitory domain, it has been hypothesized that modification in this region prevents the kinase from reverting to an inactive conformation (6). Research indicates that phosphorylation at Ser144 of PAK1 or Ser139 of PAK3 (located in the kinase inhibitory domain) affects kinase activity (7). Phosphorylation at Ser21 of PAK1 or Ser20 of PAK2 regulates binding with the adaptor protein Nck (8). PAK4, PAK5, and PAK6 have lower sequence similarity with PAK1-3 in the amino-terminal regulatory region (9). Phosphorylation at Ser474 of PAK4, a site analogous to Thr423 of PAK1, may play a pivotal role in regulating the activity and function of PAK4 (10).
- Knaus, U.G. and Bokoch, G.M. (1998) Int. J. Biochem. Cell Biol. 30, 857-862.
- Daniels, R.H. et al. (1998) EMBO J. 17, 754-764.
- King, C.C. et al. (2000) J. Biol. Chem. 275, 41201-41209.
- Manser, E. et al. (1997) Mol. Cell. Biol. 17, 1129-1143.
- Gatti, A. et al. (1999) J. Biol. Chem. 274, 8022-8028.
- Lei, M. et al. (2000) Cell 102, 387-397.
- Chong, C. et al. (2001) J. Biol. Chem. 276, 17347-17353.
- Zhao, Z. et al. (2000) Mol. Cell. Biol. 20, 3906-3917.
- Abo, A. et al. (1998) EMBO J. 17, 6527-6540.
- Qu, J. et al. (2001) Mol. Cell. Biol. 21, 3523-3533.
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- 6568 SignalSilence® Control siRNA (Unconjugated)
- 6201 SignalSilence® Control siRNA (Fluorescein Conjugate)
- 2602 PAK1 Antibody
- 2601 Phospho-PAK1 (Thr423)/PAK2 (Thr402) Antibody
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.
For Research Use Only. Not For Use In Diagnostic Procedures.