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

Abstract

Aberrant centrosome numbers are detected in virtually all human cancers where they can contribute to chromosomal instability by promoting mitotic spindle abnormalities. Despite their widespread occurrence, the molecular mechanisms that underlie centrosome amplification are only beginning to emerge. Here, we present evidence for a novel regulatory circuit involved in centrosome overduplication that centers on RNA polymerase II (pol II). We found that human papillomavirus type 16 E7 (HPV-16 E7)-and hydroxyurea (HU)-induced centriole overduplication are abrogated by a-amanitin, a potent and specific RNA pol II inhibitor. In contrast, normal centriole duplication proceeded undisturbed in a-amanitin-treated cells. Centriole overduplication was significantly reduced by siRNA-mediated knock down of CREB-binding protein (CBP), a transcriptional co-activator. We identified cyclin A2 as a key transcriptional target of RNA pol II during HU-induced centriole overduplication. Collectively, our results show that ongoing RNA pol II transcription is required for centriole overduplication whereas it may be dispensable for normal centriole duplication. Given that many chemotherapeutic agents function through inhibition of transcription, our results may help to develop strategies to target centrosomemediated chromosomal instability for cancer therapy and prevention.

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

“…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.…”

“…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…”

p21Waf1/Cip1 Deficiency Stimulates Centriole Overduplication

“…(C) Immunofluorescence analysis of control or p21-depleted 32D cells for centrin (antibody kindly provided by Jeffrey L. Salisbury, Mayo Clinic, Rochester, MN, USA) as described previously. 41 Coimmunofluorescence analysis of p21-depleted 32D cells for centrin and Cep170. 31 Briefly, cells were stained for centrin as previously described 41 followed by an anti-Cep170 antibody (kindly provided by Erich A. Nigg, Max Planck Institute for Biochemistry, Martinsried, Germany) and a Rhodamine Red-conjugated anti-rabbit secondary antibody (Jackson Immunoresearch).…”

“…41 Coimmunofluorescence analysis of p21-depleted 32D cells for centrin and Cep170. 31 Briefly, cells were stained for centrin as previously described 41 followed by an anti-Cep170 antibody (kindly provided by Erich A. Nigg, Max Planck Institute for Biochemistry, Martinsried, Germany) and a Rhodamine Red-conjugated anti-rabbit secondary antibody (Jackson Immunoresearch). All antibody incubations following the first primary antibody were for 2 h at 37˚C.…”

Abrogation of p53 function leads to metastatic transcriptome networks that typify tumor progression in human breast cancer xenografts

“…This notwithstanding, breast tumors are thought to become refractory to conventional anti-cancer therapies due in part to tumor cell heterogeneity, which confers manifold options for resistance to a given treatment modality (9)(10)(11). The tumor suppressor p53, whose function is lost in 50% of human cancers, plays a key role in the maintenance of chromosomal stability through tight coordination of DNA replication with centrosome duplication, activation of cell cycle checkpoints following genotoxic stress, and initiation of programmed cell death if damaged DNA can not be repaired properly (5,7,12,13). P53 operates as a tumor suppressor mainly as a transcription factor that directly binds to the promoter region of target genes and activates or suppresses their transcription (14).…”