GSE124657
PTEN modulates gene transcription by redistributing genome-wide RNA polymerase II occupancy [ChIP-seq]
Control of gene expression is one of the most complex yet continuous physiological processes impacting cellular homeostasis. RNA polymerase II (Pol II) transcription is tightly regulated at promoter-proximal regions by intricate dynamic processes including Pol II pausing, release into elongation, and premature termination. Here, we identify PTEN interacting with the Pol II transcription machinery and dephosphorylating Pol II C-terminal domain in vitro. PTEN alters the expression of hundreds of genes, and its restoration establishes Pol II promoter-proximal pausing in PTEN null cells. Furthermore, PTEN re-distributes genome-wide Pol II occupancy and possibly impacts Pol II pause duration, release, and elongation rate in order to enable precise gene regulation at genome-wide scale. Our observations demonstrate a direct and imperative role of PTEN in global transcriptional regulation. Overall design: Nuclei were isolated from formaldehyde (1% w/v, methanol-free) crosslinked cells by using NEXSON method followed by chromatin preparation using a Covaris sonicator. Chromatin was immunoprecipitated using an appropriate antibody, washed, reverse-crosslinked, and DNA isolated using the phenol-chloroform method. 10ng precipitated DNA was used to prepare library using Illumina TruSeq ChIP library preparation kit. Libraries were quantified for cluster generation using KAPA Library Quantification Kit. Sequencing was performed using Illumina HiSeq2500 using 50-bp paired end rapid run format. Each experiment was performed using two independent biological replicates (each containing two technical replicates).