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Image Search Results
Journal: Molecular Biology of the Cell
Article Title: A Specific Form of Phospho Protein Phosphatase 2 Regulates Anaphase-promoting Complex/Cyclosome Association with Spindle Poles
doi: 10.1091/mbc.E09-07-0598
Figure Lengend Snippet: Depletion of the PPP2 subunits CA, R1A, or R2B leads to defective spindle formation and chromosome congression. (A) HeLa cells were transfected with siRNA targeting CA, R1A, or R2B for 48 h. The extent of gene knockdown was assessed by immunoblotting with antibodies directed against CA, R1A, R2B, and actin or GAPDH as loading controls. (B) Depletion of CA, R1A, or R2B leads to centrosome, mitotic spindle assembly, and chromosome congression defects. RNAi-treated cells in A were fixed and stained with Hoechst 33342, anti-α-tubulin, anti-pericentrin, and anti-centrin antibodies to visualize DNA, the mitotic spindle, the pericentriolar material, and the centrioles, respectively. Three groups of cell phenotypes were observed upon depletion of PPP2 subunits and are designated as groups 1 through 3. Group 1, has centrosome amplification and chromosome congression defects along with multipolar spindle formation. Group 2, contains chromosome cohesion defects and defective spindles. Group 3, exhibits defective spindle formation, spindle pole fragmentation, and a failure to congress chromosomes properly. Bar, 5 μm.
Article Snippet: The following antibodies were used: mouse anti-Cdc27 (Western blot), Eg5 (BD Transduction Laboratories, Lexington, KY), mouse anti-Cdh1 (Neomarkers, Fremont, CA), rabbit anti-Cdc20, cyclin D, cyclin B (Santa Cruz Biotechnology, Santa Cruz, CA), rabbit anti-PPP1CA, PPP2CA, PPP2R1A, and PPP3CA (Upstate), rabbit anti-PPP2R2B (Bethyl Labs, Montgomery, TX), rabbit anti-PPP4C (Novus Biologicals, Littleton, CO), mouse anti-PPP5C (BD Transduction Laboratories), rabbit anti-PPP6C (Chemicon, Temecula, CA), rat anti-α-tubulin (Serotec), goat anti-GAPDH (Novus Biologicals),
Techniques: Transfection, Knockdown, Western Blot, Staining, Amplification