Skip to main content
. Author manuscript; available in PMC: 2014 Oct 1.
Published in final edited form as: Cell Signal. 2013 May 23;25(10):2047–2059. doi: 10.1016/j.cellsig.2013.05.012

Figure 1. Ectopic expression of Fbxl12 disrupts the cdk4/cyclin D1 complex by mediating CaMKI degradation.

Figure 1

(A) MLE cells were transfected with control plasmid LacZ or V5-tagged F-box proteins belonging to the L family. Cells were lysed and analyzed by immunoblotting using V5, CaMKI, PKCα, p38α/β and β-actin antibodies. (B) MLE cells transfected with increasing amounts of Fbxl12 plasmid were analyzed by immunoblotting with CaMKI, CaMKII, V5 and β-actin antibodies. (C) Flow cytometric analysis of the cells transfected with LacZ and Fbxl12 plasmids. (D) MLE cells transfected with control plasmid LacZ and Fbxl12 plasmid were processed for immunoprecipitation using cdk4 antibody. The cdk4/cyclin D1 complex was then analyzed by immunoblotting using cyclin D1 antibody. (E) MLE cells were transfected with control shRNA and CaMKI shRNA. Cells were then collected followed by immunoprecipitation with cyclin D1 antibody and analyzed by immunoblotting with cdk4 antibody. (F) MLE cells were exposed to inducers of G1 arrest for 24 hours: LPS (100ng), RA (10 mM), 2% DMSO, CHX (1-0.5mg/ml, 2–2 mg/ml), and ethanol (100 mM). Cells were then harvested and processed for immunoblot analysis with Fbxl12 and CaMKI antibodies using β-actin as a control. The data from each panel represents n=3 separate experiments.