Computational protocol: Targeting ACLY sensitizes castration-resistant prostate cancer cells to AR antagonism by impinging on an ACLY-AMPK-AR feedback mechanism

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Protocol publication

[…] After isolation of RNA using Trizol, Bioanalysis was conducted to confirm quality, using 0.8 RIN value as a cut-off for library preparation. RNA libraries were prepared according to manufacturer instruction using Illumina's TruSeq RNA sample preparation v2 kit using ‘Low Sample’ and the adapters provided in the kit were used.All the RNA-seq libraries were sequenced by the Functional Genomics Core at the University of Pennsylvania. Sequencing reads were aligned to the UCSC hg19 using RUM pipeline and differential gene expression analysis were performed using edgeR [, ]. As an unbiased interrogation of gene regulation, differentially expressed genes (FDR < 0.0001, no fold-change cut off) between any pair of conditions were selected for downstream analysis. Hierarchical clustering was performed based on the average log2-scaled gene expression levels using 1 – “Pearson correlation coefficient” as a distance measure under Ward's linkage criterion. Optimal leaf ordering was also performed for better visualization []. After the clustering, four most distinct groups of genes were defined among which two major clusters displayed increasing & gradual response to the combination of ACLYi and ENZ. Gene ontology analysis was performed using Homer and top 10 most significant terms were selected for presentation from each of “Biological Process” and “KEGG Pathway” categories []. These data have been deposited in NCBI's Gene Expression Omnibus and are accessible through GSE81796 ( […]

Pipeline specifications

Software tools RUM, edgeR, HOMER
Application RNA-seq analysis
Organisms Homo sapiens
Diseases Prostatic Neoplasms
Chemicals Fatty Acids