Journal: Circulation Research

Article Title: Peptide-Mediated Disruption of Calmodulin–Cyclin E Interactions Inhibits Proliferation of Vascular Smooth Muscle Cells and Neointima Formation

doi: 10.1161/circresaha.110.239483

Figure Lengend Snippet: Figure 1. A synthetic peptide corresponding to the CaM- binding sequence of cyclin E inhibits CaM–cyclin E interac- tions and calcium-sensitive CDK2 activity in VSMCs. A, Sequences of the CaM binding region of mouse and human cyclin E1 and of synthesized peptides. Hydrophobic residues are bold and underlined. 5A indicates control peptide with 5 alanine substi- tutions at every hydrophobic residue; NC, negative control peptide of identical length but randomly generated sequence. B, Histone H1 in vitro kinase assay on G1/S-synchronized mouse VSMC extracts at 500 nmol/L [Ca2] in the presence of peptides (each 100 mol/L). C, Co-IP analysis. Cyclin E/CDK2 complex formation and CaM–cyclin E interaction were examined in the presence of each peptide (100 mol/L). A representative blot is shown (N2). D, Histone H1 in vitro kinase assay on G1/S-synchronized mouse VSMC extracts at 0 or 500 nmol/L [Ca2] in the presence of CBS or NC (100 mol/L). For all in vitro kinase assay experiments, N3 for each condition, with experiments repeated at least twice. Results were normalized to the kinase activity of the untreated group (B) and the untreated group at 0 nmol/L [Ca2] (D). (**P0.01 vs no peptide by 1-way ANOVA and post hoc Student’s t test for multiple comparisons.)

Article Snippet: Blots were blocked with 5% non-fat dry milk in TBS-T (10 mmol/L Tris (pH 8.0), 150 mmol/L NaCl, and 0.05 % Tween 20) overnight at 4°C and incubated for 3 h at 25°C in TBS-T plus 3% non-fat dry milk containing primary antibodies: anti-CDK2 (07-631, Upstate Biotech, Billerica, MA), -cyclin E (ab7959, Abcam, Cambridge, MA), -phosphorylated CDK2 on Thr160 (2561, Cell Signaling Technology, Danvers, MA), -CaM (sc-1989, Santa Cruz Biotechnology, Santa Cruz, CA), -phosphorylated CDK2 on Thr14/Tyr15 (sc-28435-R, Santa Cruz Biotechnology), and -actin (sc-7210, Santa Cruz Biotechnology).

Techniques: Binding Assay, Sequencing, Activity Assay, Synthesized, Control, Residue, Negative Control, Generated, In Vitro, Kinase Assay, Co-Immunoprecipitation Assay

Journal: Circulation Research

Article Title: Peptide-Mediated Disruption of Calmodulin–Cyclin E Interactions Inhibits Proliferation of Vascular Smooth Muscle Cells and Neointima Formation

doi: 10.1161/circresaha.110.239483

Figure Lengend Snippet: Figure 3. CBS inhibits CDK2 phosphor- ylation and S-phase entry of serum- stimulated VSMCs. Primary mouse aortic SMCs were starved for 72 hours, nucleo- fected with peptides (1 mmol/L), serum- stimulated for 24 hours, and harvested for: Western blotting, representative blots shown with normalized band intensities (N3 for each experiment shown) (A); and FACS analysis with 10 000 cells per group (N3) (B) (*P0.05 vs no peptide by 1-way ANOVA and post hoc Student’s t test).

Article Snippet: Blots were blocked with 5% non-fat dry milk in TBS-T (10 mmol/L Tris (pH 8.0), 150 mmol/L NaCl, and 0.05 % Tween 20) overnight at 4°C and incubated for 3 h at 25°C in TBS-T plus 3% non-fat dry milk containing primary antibodies: anti-CDK2 (07-631, Upstate Biotech, Billerica, MA), -cyclin E (ab7959, Abcam, Cambridge, MA), -phosphorylated CDK2 on Thr160 (2561, Cell Signaling Technology, Danvers, MA), -CaM (sc-1989, Santa Cruz Biotechnology, Santa Cruz, CA), -phosphorylated CDK2 on Thr14/Tyr15 (sc-28435-R, Santa Cruz Biotechnology), and -actin (sc-7210, Santa Cruz Biotechnology).

Techniques: Western Blot

Journal: Cancer medicine

Article Title: The expression and role of SUZ12 in lung adenocarcinoma.

doi: 10.1002/cam4.70190

Figure Lengend Snippet: FIGURE 7 The effect of SUZ12 on CDKs and cyclins expression was tested by qRT-PCR and western blotting. sh-SUZ12 decreased CDK3 mRNA (A), CDK2/3 (C), and cyclin D1 (F) protein expression, while increased cyclin E1 protein expression (F), without significantly effected the cyclins mRNA (E). oe-SUZ12 increased CDK3/6 mRNA (B) and CDK3 protein expression (D). *p < 0.05, **p < 0.001.

Article Snippet: The list of primary antibodies: SUZ12 (1 μg/mL, Abcam Cambridge, cat no: ab12073), CDK2 (1:1000; Proteintech, USA, cat. no. 10122- 1- AP), CDK3 (1:2000; Proteintech, USA, cat. no. 55103- 1- AP), CDK6 (1:2000; Proteintech, USA, cat. no. 14052- 1- AP), cyclin D1 (1:5000; Proteintech, USA, cat. no. 26939- 1- AP), cyclin E1 (1:1000; Proteintech, USA, cat. no. 11554- 1- AP), p18 (1:1000, BOSTER China, cat. no. M03299- 1), p19 (1:1000, BOSTER China, cat. no. MA1075), p53 (1:5000; Proteintech, USA, cat. no. 60283- 2- Ig), p- p53 (1:2000; Proteintech, USA, cat. no. 28961- 1- AP), p57 (1:1000, BOSTER China, cat. no. BM4129), Rb (1:1000, BOSTER China, cat. no. BM4500), pRb (1:1000, BOSTER China, cat. no. BM4338), Bcl- 2 (1:1000; Proteintech, USA, cat. no. 26593- 1- AP), Bax (1:2000; Proteintech, USA, cat. no. 50599- 2- lg), Ecadherin (1:5000; Proteintech, USA, cat. no. 20874- 1- AP), N- cadherin (1:3000; Proteintech, USA, cat. no. 22018- 1- AP), vimentin (1:4000; Proteintech, USA, cat. no. 10366- 1- AP), MMP1 (1:1000, BOSTER China, cat. no. A00733- 1), MMP2 (1:500, BOSTER China, cat. no. BM4075), MMP9 (1:1000, BOSTER China, cat. no. PB0709), MMP14 (1:1000, BOSTER China, cat. no. BM4119), TIMP1 (1:1000, Bioss China, cat. no. bs0415R), TIMP2 (1:1000, Bioss China, cat. no. bs- 10395R), TIMP3 (1:1000; Proteintech, USA, cat. no. 10858- 1- AP), ITGB1 (1:1000, BOSTER China, cat. no. BM4308), ITGB3 (1:1000, BOSTER China, cat. no. BA1670), ITGB5 (1:1000, BOSTER China, cat. no. A04201- 1), nm23 (1:1000, BOSTER China, cat. no. BA3787), PD- L1 (1:3000; Proteintech, USA, cat. no. 66248- 1- Ig), and βactin (1:5000; Proteintech, USA, cat. no. 66009- 1- Ig).

Techniques: Expressing, Quantitative RT-PCR, Western Blot

Journal: Biomedicine & pharmacotherapy = Biomedecine & pharmacotherapie

Article Title: NT157 inhibits cell proliferation and sensitizes glioma cells to TRAIL-induced apoptosis by up-regulating DR5 expression.

doi: 10.1016/j.biopha.2022.113502

Figure Lengend Snippet: Fig. 8. Proposed signal pathways. NT157 caused ROS generation and DNA damage, which activated p21 and p27, and subsequently lunched S-phase and M-phase cell cycle arrest through regulating cyclin A1, CDK2, cyclin B1 and CDK1. NT157 also dysfunctioned MAPKs, PI3K/AKT and EGFR-STAT3 pathways, and enhanced TRAIL-induced glioma cells apoptosis by up-regulating DR5 expression.

Article Snippet: CyclinA (sc-271682), anti-p21(sc-6246), CDK1 (sc-53219) antibodies were purchased from Santa Cruz Biotechnology (Shanghai, China). β-actin antibody and IgG were bought from Beijing zhongshan Jinqiao Biotechnology (Beijing, China). p27 (#3686), CDK2 (#18048), Cyclin B1 (#12231), PARP (#9542), Caspase-3 (#9662), PCNA (#13110), Ku70 (#4588), Ku80 (#2753), γ-H2AX (#80312), p-ERK (#4370), ERK (#4695), p-p38 (#4511), p38 (#8690), p-JNK (#9255), JNK (#9252), Akt (#4685), p-Akt (#4060), EGFR (#8083), p-Stat3 (#9145), Stat3 (#9139), DR5(#69400), DR4(#42533) were all purchased from Cell Signaling Technology (CST, USA).

Techniques: Expressing