Project description:Transcriptional profiling of LNCaP prostate cancer cells comparing control siRNA-treated LNCaP cells with LNCaP cells treated with siRNAs targeting Prostate Cancer Associated Transcript-1 (PCAT1), an uncharacterized long non-coding RNA. High-throughput sequencing of polyA+ RNA (RNA-Seq) in human cancer shows remarkable potential to identify both novel disease-specific markers for clinical uses and uncharacterized aspects of tumor biology, particularly non-coding RNA (ncRNA) species. To illustrate this approach, we employed RNA-Seq on a cohort of 102 prostate tissues and cells lines and found that aberrant expression profiles of novel tissue-specific ncRNAs distinguished benign, cancerous, and metastatic tumors. Among these, a novel prostate-cancer specific ncRNA (termed PCAT-1) defined a subset of aggressive cancers with low expression of the epigenetic regulator EZH2, a component of the Polycomb Repressive Complex 2 (PRC2) commonly upregulated in metastatic cancers. In vitro assays for core PRC2 genes indicated that the PRC2 complex directly binds and represses PCAT-1, and that the PCAT-1 transcript reciprocally binds PRC2, suggesting a regulatory feedback mechanism. Importantly, knockdown of PCAT-1 in cells with high levels of endogenous PCAT-1 transcript showed changes in cell proliferation and transcriptional regulation of several key biological processes, including cell cycle. Finally, we showed that ncRNA expression signatures, including PCAT-1, were effective for the non-invasive detection of prostate cancer, and that high ncRNA expression signature values correlate with high-grade histology. The findings presented herein establish the utility of RNA-Seq to comprehensively identify unannotated ncRNAs that define human disease states and characterize PCAT-1 as a novel regulator of cell proliferation mechanistically linked to PRC2 and contributory to translational clinical tests for prostate cancer. Two-condition experiment: Control-siRNA-treated versus PCAT1-siRNA-treated LNCaP cells. Biological replicates: 3 control replicates, 3 treatment replicates.

Project description:BACKGROUND. Human prostate cancer LNCaP and PC-3 cell lines have been extensively used as prostate cancer cell models to study prostate cancer progression and to develop therapeutic agents. Although LNCaP and PC-3 cells are generally assumed to represent early and late stages of prostate cancer development, respectively, there is limited information regarding comprehensive gene expression patterns between these two cells lines and relating these cells to prostate cancer progression based on their gene expression. METHODS. Comprehensive gene expression analysis was performed in LNCaP and PC-3 cells. Total RNA was isolated from cultured cells and hybridized to Illumina human Ref-8 version 3 BeadChips representing 24,526 transcripts. Bioinformatics approach was applied to identify genes, their functional roles and interaction networks that are unique in either LNCaP or PC-3 cells. RESULTS. We observed large differences in gene expression between LNCaP and PC-3 cells.Using robust statistical analysis and very high significance criteria to identify tractable number of genes 115 and 188 genes were identified uniquely expressed in LNCaP and PC-3 cells, respectively. Genes uniquely expressed in LNCaP cells contained UDP-glucosyltransferases as a signature for this cell line. This cell line demonstrated upregulation of various metabolic pathways on gene expression level. Talα/β, GATA-1 and c-Myc/Max were identified by in silico analysis as possible transcription factors regulating unique LNCaP genes. PC-3 cells were characterized by cytosceleton-related genes, keratins in particular. Several other well known genes (VEGFC, IL8, TGFβ2 and others) scattered throughout literature were identified and summarized in the discussion. CONCLUSIONS. This study demonstrated that LNCaP and PC-3 cells represent two distinct prostate cancer cell lineages. LNCaP cells retain many prostate cell specific properties, whereas PC-3 cells have acquired more aggressive bone-like characteristics following bone metastasis and show little resemblance to prostate cells. Microarray studies confirmed previously published results and provided more information between these two prostate cancer cell lines. Future studies need to consider their similarities and differences in gene expression between localized and metastasized prostate cancer.

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LNCaP prostate cancer cells

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