cip2a antibody  (Bio-Techne corporation)

Journal: Journal of Cellular and Molecular Medicine

Article Title: CIP2A facilitates the G1/S cell cycle transition via B‐Myb in human papillomavirus 16 oncoprotein E6‐expressing cells

doi: 10.1111/jcmm.13693

Figure Lengend Snippet: Induction of CIP2A mRNA and protein expression by HPV‐16E6 in PHKs. A, mRNA expression of HPV‐16E6 in PHKs expressing 16E6 and F2V using β‐actin as a loading control. B, Protein levels of HPV‐16E6, p53 and p21 in PHKs expressing 16E6 and F2V. Expression of GAPDH was used as a loading control. A representative of 2 independent experiments is shown. C, HPV‐16E6 expression leads to increased protein expression of CIP2A in PHKs. Data from a representative of 3 experiments are shown. D, Data from 3 experiments are summarized. E, Relative CIP2A mRNA expression was determined by qRT‐PCR in the above cells. Data from 3 experiments are summarized. The mean and standard deviation (SD) of 3 independent experiments are shown. Babe, pBabe‐puromycin vector. *, P < .05; **, P < .01; and ***, P < .001

Article Snippet: Membranes were probed with primary antibodies specific for the following proteins: Cdk1 (BD Biosciences, 610038), p21 (BD Biosciences, 610233); CIP2A (Novus Biologicals, NB100‐68264); B‐Myb (sc‐725), Cdk2 (sc‐6248), Cdk4 (sc‐260), Cdk6 (sc‐177), cyclin A2 (sc‐751), cyclin B1 (sc‐752), cyclin D1 (sc‐718), cyclin E1 (sc‐198), c‐Myc (sc‐764), p53 (sc‐126) and GAPDH (all from Santa Cruz Biotechnology, sc‐25778); and phospho‐c‐Myc (phospho S62, Abcam, ab51156), HPV‐16/18E6 (Abcam 70) and β‐tubulin (Sigma‐Aldrich, T4026).

Techniques: Expressing, Control, Quantitative RT-PCR, Standard Deviation, Plasmid Preparation

Journal: Journal of Cellular and Molecular Medicine

Article Title: CIP2A facilitates the G1/S cell cycle transition via B‐Myb in human papillomavirus 16 oncoprotein E6‐expressing cells

doi: 10.1111/jcmm.13693

Figure Lengend Snippet: Inhibition of CIP2A by siRNA impeded cell viability and DNA synthesis in HPV‐16E6–expressing cells. A, Elevated expression of CIP2A protein in 16E6‐expressing RPE1 cells. B, Western blot analysis of CIP2A and p53 proteins after transfection with scrambled siRNA (siCon) or CIP2A siRNA (siCIP2A) for 48 h. A representative of 3 independent experiments is shown. C, Cell viability assay of RPE1‐16E6 cells with CIP2A knockdown. D, Representative flow cytometry of BrdU staining profiles is shown. E, The mean and SD of BrdU‐positive cells from 3 experiments are summarized. **, P < .01.

Article Snippet: Membranes were probed with primary antibodies specific for the following proteins: Cdk1 (BD Biosciences, 610038), p21 (BD Biosciences, 610233); CIP2A (Novus Biologicals, NB100‐68264); B‐Myb (sc‐725), Cdk2 (sc‐6248), Cdk4 (sc‐260), Cdk6 (sc‐177), cyclin A2 (sc‐751), cyclin B1 (sc‐752), cyclin D1 (sc‐718), cyclin E1 (sc‐198), c‐Myc (sc‐764), p53 (sc‐126) and GAPDH (all from Santa Cruz Biotechnology, sc‐25778); and phospho‐c‐Myc (phospho S62, Abcam, ab51156), HPV‐16/18E6 (Abcam 70) and β‐tubulin (Sigma‐Aldrich, T4026).

Techniques: Inhibition, DNA Synthesis, Expressing, Western Blot, Transfection, Viability Assay, Knockdown, Flow Cytometry, BrdU Staining

Journal: Journal of Cellular and Molecular Medicine

Article Title: CIP2A facilitates the G1/S cell cycle transition via B‐Myb in human papillomavirus 16 oncoprotein E6‐expressing cells

doi: 10.1111/jcmm.13693

Figure Lengend Snippet: Silencing CIP2A caused G1 arrest in 16E6‐expressing cells. A, Flow cytometric analysis of 16E6‐expressing cells transfected with CIP2A siRNA for 36 h, treated with PBS or 10 μg/mL bleomycin for 24 h and then stained with PI. G1, S and G2 phases are indicated. Data from a representative of 4 experiments are shown. B, Quantification of percentages G1 phase and S phase cells. Data from 4 experiments are summarized. *, P < .05; **, P < .01

Article Snippet: Membranes were probed with primary antibodies specific for the following proteins: Cdk1 (BD Biosciences, 610038), p21 (BD Biosciences, 610233); CIP2A (Novus Biologicals, NB100‐68264); B‐Myb (sc‐725), Cdk2 (sc‐6248), Cdk4 (sc‐260), Cdk6 (sc‐177), cyclin A2 (sc‐751), cyclin B1 (sc‐752), cyclin D1 (sc‐718), cyclin E1 (sc‐198), c‐Myc (sc‐764), p53 (sc‐126) and GAPDH (all from Santa Cruz Biotechnology, sc‐25778); and phospho‐c‐Myc (phospho S62, Abcam, ab51156), HPV‐16/18E6 (Abcam 70) and β‐tubulin (Sigma‐Aldrich, T4026).

Techniques: Expressing, Transfection, Staining

Journal: Journal of Cellular and Molecular Medicine

Article Title: CIP2A facilitates the G1/S cell cycle transition via B‐Myb in human papillomavirus 16 oncoprotein E6‐expressing cells

doi: 10.1111/jcmm.13693

Figure Lengend Snippet: Silencing CIP2A caused decreased Cdk1 and Cdk2 proteins in 16E6‐expressing cells. A, Western blot analysis of CIP2A, Cdk4, Cdk6, cyclin D1, Cdk1, Cdk2, cyclin B1, cyclin A2 and cyclin E1 protein levels in cells expressing HPV‐16E6 transfected with CIP2A siRNA and then treated with PBS or 10 μg/mL bleomycin for 24 h. A representative of 3 independent experiments is shown. B, Quantification of all cell cycle‐related proteins. Data from 3 experiments are summarized. C, Relative mRNA levels of all cell cycle‐related genes determined by qRT‐PCR. Data from 3 experiments are summarized. *, P < .05; **, P < .01

Article Snippet: Membranes were probed with primary antibodies specific for the following proteins: Cdk1 (BD Biosciences, 610038), p21 (BD Biosciences, 610233); CIP2A (Novus Biologicals, NB100‐68264); B‐Myb (sc‐725), Cdk2 (sc‐6248), Cdk4 (sc‐260), Cdk6 (sc‐177), cyclin A2 (sc‐751), cyclin B1 (sc‐752), cyclin D1 (sc‐718), cyclin E1 (sc‐198), c‐Myc (sc‐764), p53 (sc‐126) and GAPDH (all from Santa Cruz Biotechnology, sc‐25778); and phospho‐c‐Myc (phospho S62, Abcam, ab51156), HPV‐16/18E6 (Abcam 70) and β‐tubulin (Sigma‐Aldrich, T4026).

Techniques: Expressing, Western Blot, Transfection, Quantitative RT-PCR

Journal: Journal of Cellular and Molecular Medicine

Article Title: CIP2A facilitates the G1/S cell cycle transition via B‐Myb in human papillomavirus 16 oncoprotein E6‐expressing cells

doi: 10.1111/jcmm.13693

Figure Lengend Snippet: Regulation of Cdk1 and Cdk2 by CIP2A is dependent on B‐Myb rather than c‐Myc. A, Western blot analysis of CIP2A, c‐Myc, phospho‐S62‐Myc and B‐Myb protein levels in 16E6‐expressing cells after CIP2A knockdown. β‐Tubulin was used as a loading control. B, Protein levels of B‐Myb, c‐Myc and phospho‐S62‐Myc in 16E6‐expressing PHKs and (C) RPE1 cells. D, Protein levels of B‐Myb, Cdk1 and Cdk2, CIP2A and p53 in 16E6‐expressing cells after B‐Myb knockdown with siRNA. Data from a representative of 3 experiments are shown. E, Knockdown of B‐Myb down‐regulates Cdk1 and Cdk2 luciferase reporter activities. RPE1 cells were cotransfected with the Cdk1 or Cdk2 promoter‐luciferase constructs and renilla luciferase control plasmid together with B‐Myb siRNA plasmid. Cells were harvested after 48 h, and lysates were assayed for luciferase activity. F, Flow cytometric analysis of 16E6‐expressing cells transfected with B‐Myb siRNA treated with PBS or bleomycin. G1, S and G2 phases are indicated. A representative flow cytometry of 3 independent experiments is shown. G, Quantification of percentages G1 phase cells. Data from 3 experiments are summarized. H, Western blot analysis of B‐Myb, Cdk1 and Cdk2 in B‐Myb–overexpressing CIP2A knockdown cells. Data from a representative of 3 experiments are shown. *, P < .05; ***, P < .001

Article Snippet: Membranes were probed with primary antibodies specific for the following proteins: Cdk1 (BD Biosciences, 610038), p21 (BD Biosciences, 610233); CIP2A (Novus Biologicals, NB100‐68264); B‐Myb (sc‐725), Cdk2 (sc‐6248), Cdk4 (sc‐260), Cdk6 (sc‐177), cyclin A2 (sc‐751), cyclin B1 (sc‐752), cyclin D1 (sc‐718), cyclin E1 (sc‐198), c‐Myc (sc‐764), p53 (sc‐126) and GAPDH (all from Santa Cruz Biotechnology, sc‐25778); and phospho‐c‐Myc (phospho S62, Abcam, ab51156), HPV‐16/18E6 (Abcam 70) and β‐tubulin (Sigma‐Aldrich, T4026).

Techniques: Western Blot, Expressing, Knockdown, Control, Luciferase, Construct, Plasmid Preparation, Activity Assay, Transfection, Flow Cytometry

Journal: Journal of Cellular and Molecular Medicine

Article Title: CIP2A facilitates the G1/S cell cycle transition via B‐Myb in human papillomavirus 16 oncoprotein E6‐expressing cells

doi: 10.1111/jcmm.13693

Figure Lengend Snippet: Inhibition of Cdk1 and Cdk2 by CIP2A knockdown in cervical cancer SiHa cells caused G1 arrest. A, Western blot analysis of 16E6, p53 and CIP2A after HPV‐16E6 knockdown in cervical cancer SiHa cells. B, Protein expression of CIP2A, B‐Myb, Cdk1 and Cdk2 in SiHa cells after CIP2A knockdown. Data from a representative of 3 experiments are shown. C, Flow cytometric analysis of SiHa cells with CIP2A knockdown treated with PBS or bleomycin. A representative flow cytometry of 3 independent experiments is shown. D, Quantification of percentages G1 phase cells. Data from 3 experiments are summarized. *, P < .05

Article Snippet: Membranes were probed with primary antibodies specific for the following proteins: Cdk1 (BD Biosciences, 610038), p21 (BD Biosciences, 610233); CIP2A (Novus Biologicals, NB100‐68264); B‐Myb (sc‐725), Cdk2 (sc‐6248), Cdk4 (sc‐260), Cdk6 (sc‐177), cyclin A2 (sc‐751), cyclin B1 (sc‐752), cyclin D1 (sc‐718), cyclin E1 (sc‐198), c‐Myc (sc‐764), p53 (sc‐126) and GAPDH (all from Santa Cruz Biotechnology, sc‐25778); and phospho‐c‐Myc (phospho S62, Abcam, ab51156), HPV‐16/18E6 (Abcam 70) and β‐tubulin (Sigma‐Aldrich, T4026).

Techniques: Inhibition, Knockdown, Western Blot, Expressing, Flow Cytometry