Over-expression of Id-1 induces cell proliferation in hepatocellular carcinoma through inactivation of p16INK4a/RB pathway.
Inhibitors of differentiation and DNA binding-1 (Id-1) have been demonstrated to oppose Ets-mediated activation of p16INK4a. As p16INK4a protein is inactivated in hepatocellular carcinoma (HCC), we aimed to investigate the role of Id-1 in regulating p16INK4a expression during the development of HCC in HCC patients and direct ectopic Id-1 introduction into the PLC/PRF/5 HCC cell line. Sixty-two HCC samples were recruited for evaluation of Id-1 and proliferating cell nuclear antigen (PCNA) protein expression. The messenger RNA (mRNA) expression of Id-1 and p16INK4a was detected by quantitative reverse transcription-polymerase chain reaction. For in vitro Id-1 transfection, five Id-1 transfected clones were isolated and the effect of ectopic Id-1 introduction was investigated by 3-(4,5-cimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide assay, flow cytometry, immunostaining and western blot. Our results showed that Id-1 was over-expressed in HCC specimens both at mRNA and protein levels. Over-expression of Id-1 protein was correlated with PCNA (r = 0.334, P = 0.033). HCC samples showing low Id-1 protein expression had a lower Id-1 mRNA level (340.2 versus 1467%, P = 0.039) and higher p16INK4a expression (195 versus -78.6%, P = 0.039) than samples with high Id-1 protein expression. In the PLC/PRF/5 HCC cell line study, ectopic Id-1 expression resulted in proliferation of HCC cells and an increased percentage of S phase cells and PCNA expression. The results showed that over-expression of Id-1 induces cell proliferation in HCC through inactivation of p16INK4a/retinoblastoma pathway. In conclusion, the results provided an insight for the understanding of the role of Id-1 in functional inactivation of p16INK4a in HCC.
Lee
Terence Kin-Wah
TK
Centre for the Study of Liver Disease and Department of Surgery, The University of Hong Kong, Pokfulam, Hong Kong, China.
Man
Kwan
K
Ling
Ming-Tat
MT
Wang
Xiang-Hong
XH
Wong
Yong-Chuan
YC
Lo
Chung-Mau
CM
Poon
Ronnie Tung-Ping
RT
Ng
Irene Oi-Lin
IO
Fan
Sheung-Tat
ST
D001483:Base Sequence; D006528:Carcinoma, Hepatocellular / Q000378:metabolism / Q000473:pathology*; D002455:Cell Division / Q000502:physiology*; D019941:Cyclin-Dependent Kinase Inhibitor p16 / Q000235:genetics / Q000378:metabolism*; D017931:DNA Primers; D006801:Humans; D051798:Inhibitor of Differentiation Protein 1; D008113:Liver Neoplasms / Q000378:metabolism / Q000473:pathology*; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D012097:Repressor Proteins; D016160:Retinoblastoma Protein / Q000235:genetics / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism / Q000502:physiology*
D019941:Cyclin-Dependent Kinase Inhibitor p16; D017931:DNA Primers; C497453:ID1 protein, human; D051798:Inhibitor of Differentiation Protein 1; D012333:RNA, Messenger; D012097:Repressor Proteins; D016160:Retinoblastoma Protein; D014157:Transcription Factors