Cyclin-dependent kinase inhibitor indirubin-3′-oxime selectively inhibits human papillomavirus type 16 E7-induced numerical centrosome anomalies

Abstract

Dysregulation of the centrosome duplication cycle has been implicated in tumorigenesis. Our previous work has shown that the human papillomavirus type 16 (HPV-16) E7 oncoprotein rapidly induces aberrant centrosome and centriole duplication in normal human cells. We report here that HPV E7-induced abnormal centriole duplication is specifically abrogated by a small molecule CDK inhibitor, indirubin-3 0 -oxime (IO), but not a kinase-inactive derivative. Importantly, normal centriole duplication was not markedly affected by IO, and the inhibitory effects were observed at concentrations that did not affect the G1/S transition of the cell division cycle. Depletion of CDK2 by siRNA similarly abrogated HPV E7-induced abnormal centrosome duplication and ectopic expression of CDK2 in combination with cyclin E or cyclin A could rescue the inhibitory effect of IO. IO treatment also reduced the steady-state level of aneuploid cells in HPV-16 E7-expressing cell populations. Our results suggest that cyclin/CDK2 activity is critically involved in abnormal centrosome duplication induced by HPV-16 E7 oncoprotein expression, but may be dispensable for normal centrosome duplication and cell cycle progression.

Centriole overduplication through the concurrent formation of multiple daughter centrioles at single maternal templates

“…Overexpression of cyclin E/CDK2 and PLK4 stimulates the concurrent formation of multiple daughter centrioles at single maternal centrioles We asked whether ectopic expression of cyclin E/CDK2 and/or PLK4 could aberrant daughter centriole formation in the absence of a proteasome inhibitor. Transient transfection of U-2 OS/centrin-GFP cells with cyclin E/CDK2 did not lead to a significant increase in the percentage of cells with excessive daughter centriole formation at maternal templates (Figure 4a), although in line with previous experiments (Duensing et al, 2004), a 2.2-fold increase in cells with more than four centrioles arranged in a random fashion was detected (data not shown). Overexpression of PLK4, however, caused a significant 17.9-fold increase of abnormal centriole formation at maternal centrioles from 1 to 17.9% (Pp0.0005) in agreement with previous results (Habedanck et al, 2005).…”

“…The human papillomavirus type 16 (HPV-16) E7 oncoprotein has been found to rapidly stimulate supernumerary centrosomes in primary human cells and tumor cell lines (Duensing et al, 2000(Duensing et al, , 2004(Duensing et al, , 2006a. HPV-16 E7 binds and degrades the retinoblastoma tumor suppressor protein (pRB), inactivates cyclindependent kinase inhibitors such as p21…”

“…How these activities can trigger centriole overduplication within a time period that corresponds to approximately a single cell-division cycle (Duensing et al, 2004(Duensing et al, , 2006a in currently unknown.…”

RNA polymerase II transcription is required for human papillomavirus type 16 E7- and hydroxyurea-induced centriole overduplication

“…HPV-16 E7-induced centriole overduplication requires CDK2 activity (Duensing et al, 2004;Duensing et al, 2006) and, as shown here, ongoing RNA pol II transcription. These findings highlight that the HPV-16 E7-induced generation of immature daughter centrioles (Guarguaglini et al, 2005) is an active process with specific molecular requirements and not simply a consequence of cytokinesis defects, ploidy alterations or aberrant centriole splitting .…”

“…We have previously shown that CDK2 activity is critical for centriole overduplication induced by the HPV-16 E7 oncoprotein (Duensing et al, 2004;Duensing et al, 2006). CDK2 is activated by cyclin E in late G1 and by cyclin A during S phase (Malumbres and Barbacid, 2001).…”