Metabolomics,Unknown,Transcriptomics,Genomics,Proteomics

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Microarray screening of RNA samples from low-, intermediate- and high-passage human prostate LNCaP cells

ABSTRACT: This laboratory studies the structures, biosynthesis, and functions of mucin glycans The low-(<33) and high-(>80)passage human prostate LNCaP cells developed by Dr. Ming-Fong Lin (Lin et al, Urology 166:1943, 2001) have been considered as the best in vitro model that reflects the progression of prostate cancer from androgen-dependent to androgen-independent stage (Parajuli et al, Cancer Res. 61:8227, 2001; Demeade et al, The Prostate 54:249, 2003; Unni et al, Cancer Res. 64:7156, 2004). The high-passage cells are metastatic while the low-passage cells are not. RT-PCR analysis of these two cell clones for 25 glycogenes among 8 different groups of glycogenes showed that only high-passage cells expressed two (isozymes 1 and 5) of the five known GlcNAc6ST genes. In addition, P-selectin blot analysis showed qualitative and quantitative differences between these clones. They included 3 higher intensity bands and 3 lower intensity bands in high-passage cells as compared to low-passage cells. The difference in P-selectin-specific glycoconjugates between low and high-passage LNCaP cells should help identify the glycans that distinguish high- from low-passage LNCaP cells. RNA preparations from low-, intermediate-, and high-passage human prostate LNCaP cells were sent to Microarray Core (E). *Concurrent analysis of glycan profiles in these cells was performed in Core C*. Three replicate samples from each condition were used in the study. The RNA was amplified, labeled, and hybridized to the GLYCOv3 microarrays. Data was analyzed to identify the glycogenes, glycans, and/or carbohydrate binding proteins responsible for the difference in metastatic property between low- and high-passage LNCaP cells. The intermediate-passage cells were used for identifying changes in glycogene, glycan, and/or carbohydrate binding protein expression that may be responsible for the earliest change of the metastatic property.

ORGANISM(S): Homo sapiens

SUBMITTER: Steven Head

PROVIDER: E-GEOD-28927 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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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.

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Microarray screening of RNA samples from low-, intermediate- and high-passage human prostate LNCaP cells

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