Silencing of integrated human papillomavirus-16 oncogenes by small interfering RNA-mediated heterochromatization.
Double-stranded RNAs or small interfering RNAs (siRNA) targeting the promoters of genes are known to cause gene knockdown by a process known as transcriptional gene silencing (TGS). We screened multiple siRNAs homologous to one of the NF-1 binding sites in the human papillomavirus-16 (HPV-16) enhancer and identified one siRNA which causes specific TGS of the HPV-16 oncogenes E6 and E7 when transfected into two HPV-16-positive cell lines siHa and CaSki. This phenomenon was specific to the HPV-16 enhancer with no effect on the HPV-18 enhancer. TGS was associated with heterochromatization of the targeted region of the enhancer but no DNA methylation was noted during the time period studied. The choice of target in the enhancer was important as siRNAs differing by one or two bases showed no suppression of downstream gene expression. A low copy number enhancer-associated transcript was detected in the cell lines studied and its level decreased significantly after treatment with the siRNA that caused TGS. This supports the RNA:RNA model described previously for TGS. This siRNA which causes simultaneous silencing of E6 as well as E7 oncogenes by an epigenetic mechanism might be useful as a therapeutic modality for HPV-16-positive cervical and other epithelial cancers.
Palanichamy
Jayanth Kumar
JK
Department of Biochemistry, All India Institute of Medical Sciences, New Delhi 110029, India.
Mehndiratta
Mohit
M
Bhagat
Mohita
M
Ramalingam
Pradeep
P
Das
Brati
B
Das
Prerna
P
Sinha
Subrata
S
Chattopadhyay
Parthaprasad
P
D017209:Apoptosis; D001483:Base Sequence; D001665:Binding Sites / Q000235:genetics; D015153:Blotting, Western; D045744:Cell Line, Tumor; D049109:Cell Proliferation; D002843:Chromatin / Q000235:genetics / Q000378:metabolism; D004742:Enhancer Elements, Genetic / Q000235:genetics; D006367:HeLa Cells; D006570:Heterochromatin / Q000235:genetics / Q000378:metabolism; D006657:Histones / Q000378:metabolism; D052162:Human papillomavirus 16 / Q000235:genetics; D006801:Humans; D008239:Lysine / Q000378:metabolism; D008745:Methylation; D025542:Neurofibromin 1 / Q000378:metabolism; D009856:Oncogene Proteins, Viral / Q000235:genetics*; D050725:Papillomavirus E7 Proteins / Q000235:genetics*; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000235:genetics*; D012097:Repressor Proteins / Q000235:genetics*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014158:Transcription, Genetic; D014162:Transfection; D016159:Tumor Suppressor Protein p53 / Q000378:metabolism
D002843:Chromatin; C068651:E6 protein, Human papillomavirus type 16; D006570:Heterochromatin; D006657:Histones; D025542:Neurofibromin 1; D009856:Oncogene Proteins, Viral; D050725:Papillomavirus E7 Proteins; D034741:RNA, Small Interfering; D012097:Repressor Proteins; D016159:Tumor Suppressor Protein p53; C059731:oncogene protein E7, Human papillomavirus type 16; D008239:Lysine