The Study On Role And Mechanism Of JMJD5 In Hepatocellular Carcinoma

Research shows that JmjC family contains more than thirty members,all of them have the same Jumonji C domain which owns demethylase activity.This family can erase three methyl from methylated histones.It is known that many members of this family are dysregulated in many human cancers and can regultate the proliferation and invasion of tumor cells,but the role of this family in the initiation and progression of hepatocellular carcinoma(HCC)is still not clear.In this study,we first evaluated the tissue expression profile of this family in HCC by mining public database and found that jumonji domain containing 5(JMJD5)decreased significantly in HCC.Meanwhile downregulation of JMJD5 has clinical significance,HCC patients with low JMJD5 expression have shorter survival time compared to those with high JMJD5 expression.Some cellular and molecular experiments including Western blotting,Real-time PCR and Chomatin Immunoprecipitation confirmed that JMJD5 was indeed obviously decreased in HCC tissues and the downregulation of JMJD5 is caused by altered epigenetic histone modifications.In HCC specimens,some transcriptional active markers,including acetylation of H3K9 and H3K27 and methylation of H3K4,decreased on the JMJD5 promoter while the transcriptional repressive markers such as trimethylation of H3K9 and H3K27 increased.Functional experiments revealed that knockdown of JMJD5 promoted HCC cell growth curve,plate colony formation,soft agar colony formation and in vivo tumorigenicity.On the contrary,ectopic JMJD5 expression had the opposite effects.Cell Cytometry assay showed that JMJD5 knockdown accelerated cell cycle from G1 phase to S phase,but overexpression of JMJD5 inhibited the S phase entrance.At molecular mechanism,we found that JMJD5 inhibited HCC cell ploliferation mainly by activating CDKN1 A transcription.JMJD5 knockdown led to the down-regulation of CDKN1 A and ectopic expression of JMJD5 not only increased but also rescued CDKN1 A transcription.Moreover,CDKN1 A knockdown could abrogate the effect of JMJD5 knockdown or overexpression on cell proliferation.Meanwhile in TP53-null Hep3 B cells,JMJD5 knckdown also reduced CDKN1 A transcription suggesting that JMJD5 regulated CDKN1 A transcription independent on TP53.ChIP and luciferase reporter experiments proved that JMJD5 could bind to CKDN1 A promoter and regulated its activity,moreover JMJD5 did not affect dimethylation of H3K36 on CDKN1 A gene locus.In addition,Western blotting and plate coloy formation assays exhibited that mutant JMJD5 which abrogated its enzyme activity not only activated CDKN1 A transcription but also inhibited HCC cell proliferation suggesting that JMJD5 regulated CDKN1 A transcription and HCC cell proliferation independent on its enzyme activity.All in all,our results prove that JMJD5 is a potential tumor suppressor gene in HCC,epigenetic silencing of JMJD5 promotes the proliferation of HCC cells by downregulating the transcription of CDKN1 A.