Human papillomavirus 16 E6 increases the radiosensitivity of p53-mutated cervical cancer cells, associated with up-regulation of aurora A.
OBJECTIVE
To examine the effect of the human papillomavirus (HPV) type 16-E6 (HPV 'early' gene) oncoprotein on in vitro radiosensitivity of HPV-negative/p53 mutant C33a cervical cancer cells.


METHODS
The human cervical cancer cell line C33a was stably transfected with either the HPV16 E6 cDNA cloned into the vector pcDNA3.0 (C33aE6) or the empty-vector control (C33aV). Radiosensitivity, DNA damage, and cell cycle measurements were made using standard clonogenic assays, immunofluorescent assessment of nuclear histone H2AX phosphorylated on serine-139 (gamma-H2AX) foci, and flow cytometry. Western immunoblotting and fluorescence confocal microscopy were used to analyse the changes in cellular proteins. Real-time polymerase chain reaction (PCR) was used to compare levels of aurora A mRNA.


RESULTS
Compared to C33aV cells, C33aE6 cells showed enhanced radiation cell killing. This was associated with a large amount of polyploidy which was followed by late cell death in C33aE6 cells. Aurora A was highly expressed in C33aE6 cells at pre- and post-irradiation times compared to C33aV cells. Silencing aurora A resulted in a reduced amount of residual gamma-H2AX foci in C33aE6 cells, and diminished the difference in radiosensitivity between the C33aE6 and C33aV cells.


CONCLUSIONS
Our in vitro results indicate that genetic instability could be augmented in the HPV-infected cancer cells by up-regulation of aurora A, especially against a background of dysfunctional p53. Further studies are needed to examine whether aurora A could be a viable therapeutic target in HPV-related tumours.
Shin
Hye-Jin
HJ
Research Institute and Hospital, National Cancer Center, Goyang, Korea.
Kim
Joo-Young
JY
Hampson
Lynne
L
Pyo
Hongryull
H
Baek
Hye Jung
HJ
Roberts
Stephen A
SA
Hendry
Jolyon H
JH
Hampson
Ian N
IN
D064127:Aurora Kinases; D045744:Cell Line, Tumor; D005260:Female; D015971:Gene Expression Regulation, Enzymologic; D016158:Genes, p53; D006657:Histones / Q000032:analysis; D006801:Humans; D009154:Mutation; D009856:Oncogene Proteins, Viral / Q000502:physiology*; D011123:Polyploidy; D017346:Protein Serine-Threonine Kinases / Q000037:antagonists & inhibitors / Q000235:genetics / Q000502:physiology*; D034741:RNA, Small Interfering / Q000235:genetics; D011836:Radiation Tolerance; D012097:Repressor Proteins / Q000502:physiology*; D015854:Up-Regulation; D002583:Uterine Cervical Neoplasms / Q000235:genetics / Q000473:pathology / Q000532:radiotherapy* / Q000821:virology
C068651:E6 protein, Human papillomavirus type 16; C468783:H2AX protein, human; D006657:Histones; D009856:Oncogene Proteins, Viral; D034741:RNA, Small Interfering; D012097:Repressor Proteins; D064127:Aurora Kinases; D017346:Protein Serine-Threonine Kinases