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. Author manuscript; available in PMC: 2010 Aug 15.
Published in final edited form as: Cancer Res. 2009 Aug 15;69(16):6668–6675. doi: 10.1158/0008-5472.CAN-09-1284

Figure 6. CUL1 regulates PLK4 protein stability.

Figure 6

(A) Immunofluorescence microscopic analysis of U-2 OS/centrin-GFP cells for endogenous PLK4 at 48 h after transfection of empty vector (control) or DN-CUL1 (left panels). Note centriole multiplication together with an excessive amount of endogenous PLK4 in DN-CUL1-transfected cells (bottom). Quantification of the fold-changes of PLK4 integrated signal intensities at maternal centrioles in U-2 OS/centrin-GFP cells transfected with empty vector or DN-CUL1 for 48 h or treated with 1 μM IO or 0.1% DMSO (control) for 48 h (right panel).

(B) Immunoblot analysis of whole cell extracts from U-2 OS/centrin-GFP cells after transient transfection (48 h) with empty vectors (control) or PLK4 in combination with either cyclin E/CDK2 alone or cyclin E/CDK2 and DN-CUL1. Note the increase of PLK4 protein in the presence of DN-CUL1 in the last lane.

(C) Immunoblot analysis of whole cell extracts from U-2 OS/centrin-GFP cells after transient transfection with empty vectors (control) or dominant-negative CUL1 (DN-CUL1) for 48 h followed by transfection with Myc-PLK4 for 24 h and treatment with 30 μg/ml cycloheximide (CHX) for the indicated time intervals. Note the increased protein stability of Myc-PLK4 in the presence of DN-CUL1 (6 h CHX, last lane).

(D) Tentative model of centriole multiplication triggered by oncogenic stimuli such as c-MYC, E2F-1 or HPV-16 E7. These stimuli are known to deregulate cyclin E/CDK2 complexes and we show here that this leads to an aberrant recruitment of PLK4 to maternal centrioles. However, degradation of enzymatically active PLK4 (and low baseline protein expression) by CUL1-based SCF E3 ubiquitin ligase activity prevents the formation of supernumerary daughter centrioles. Only when PLK4 is overexpressed or its CUL1-mediated degradation is altered, centriole multiplication occurs. Hence, it is likely that oncogene-induced centriole multiplication, for example by HPV-16 E7, also involves impairment of CUL1-mediated protein degradation through mechanisms that remain to be determined. Genetic alterations such as deletion of the CUL1 locus in certain human malignancies are likewise to promote centriole duplication.