RRC ID 44120
Author Venkatesan N, Krishnakumar S, Deepa PR, Deepa M, Khetan V, Reddy MA.
Title Molecular deregulation induced by silencing of the high mobility group protein A2 gene in retinoblastoma cells.
Journal Mol Vis
Abstract AIM:To explore the molecular mechanisms deregulated by high mobility group protein A2 (HMGA2) gene silencing in retinoblastoma (RB) cells.
METHODS:Synthetic anti-HMGA2 short interfering RNA (siRNA) was used to silence the HMGA2 gene in cultured Y79 RB cells that were subjected to whole genome microarray analysis. The expression of differentially regulated key genes was confirmed with quantitative reverse-transcriptase polymerase chain reaction (qRT-PCR) in post-silenced RB cell lines (Y79 and WERI Rb1). These deregulated genes were compared for their constitutive expression in primary RB tumors (n=10). Zymographic determination of matrix metalloproteinase (MMP) activity was performed in RB cells. A cell cycle assay and a proliferation assay were performed in post-transfected RB cells.
RESULTS:HMGA2 gene silencing in cultured RB cells results in reduced cell proliferation and transition in the G1/S phase. The whole genome microarray analysis of HMGA2 silenced Y79 cells revealed overall upregulation of 1,132 genes (≥ 1.0 fold) and downregulation of 1,562 genes (≤ -1.0 fold). Specific quantitative pathway analysis of the deregulated genes (using Biointerpreter) revealed 150 upregulated genes and 77 downregulated genes (≥ 1.0 fold) involved in vital pathways, namely, mitogen-activated protein kinase, Janus kinase/signal transducers and activators of transcription, Ras pathway, Ras-induced extracellular signal-regulated protein kinases 1 and 2, and tumor protein p53. The differential expression of genes obtained from microarray analysis (Homo sapiens ELK1, member of ETS oncogene family [ELK1], Homo sapiens cyclin-dependent kinase 6 [CDK6], Homo sapiens E2F transcription factor 4, p107/p130-binding [E2F4], Homo sapiens G-2 and S-phase expressed 1 [GTSE1], Damage-regulated autophagy modulator [DRAM], Homo sapiens cadherin 1, type 1,E-cadherin (epithelial) [CDH1], Homo sapiens snail homolog 1 (Drosophila) [SNAI1], Homo sapiens matrix metallopeptidase 2 [MMP2], and Homo sapiens matrix metallopeptidase 9 [MMP9]) was confirmed with quantitative reverse-transcriptase polymerase chain reaction in post-silenced RB cells. Zymographic analysis revealed that the increase in MMP mRNA expression in the post-silenced RB cells did not correlate with corresponding enzyme activity.
CONCLUSIONS:Our study revealed molecular regulatory changes induced by HMGA2 silencing in RB cancer cells, offering mechanistic insights into the anticancer potential. HMGA2 may be considered a promising candidate for gene silencing therapy in RB.
Volume 18
Pages 2420-37
Published 2012
PMID 23077401
PMC PMC3472926
MeSH Cell Cycle Cell Line, Tumor Cell Proliferation Eye Proteins / genetics* Gene Expression Profiling Gene Expression Regulation, Neoplastic Gene Silencing HMGA2 Protein / antagonists & inhibitors* HMGA2 Protein / genetics* Humans Matrix Metalloproteinases / genetics Molecular Targeted Therapy Neoplasm Proteins / genetics* Oligonucleotide Array Sequence Analysis RNA, Small Interfering / genetics* Retinoblastoma / genetics* Retinoblastoma / metabolism Retinoblastoma / pathology Signal Transduction / genetics
IF 2.174
Times Cited 14
Human and Animal Cells