Project description:Analysis of acute effects of ligand-treatment on vitamin D receptor binding genome-wide using ChIP-seq. THP-1 monocytic leucemia cells were treated with 1?,25(OH)2D3 (1,25D) or left unstimulated to investigate the acute effects of VDR chromatin occupancy. We identified in total 2340 VDR binding sites with and without the ligand. Without the ligand, there is a considerable presence of VDR already on the chromatin. However, upon a short (40 min) ligand treatment VDR shifts from sites that rarely contain a DR3 type element to sites that frequently contain one or more DR3-type element. Genome-wide identification of VDR binding in THP-1 cells at the unstimulated state and after 40 min ligand (10 nM 1?,25(OH)2D3 (1,25D, calcitriol)) treatment.
Project description:The RNA polymerase II (POLII) driven transcription cycle is tightly regulated at distinct checkpoints through cyclin dependent kinases (CDKs) and their cognate Cyclins. The molecular events underpinning transcriptional elongation and processivity and CDK-Cyclins involved remain poorly understood. Using CRISPR-CAS9 homology-directed-repair we generated analog-sensitive-kinase variants of CDK12 and CDK13 to probe their individual and shared biological and molecular roles. Single inhibition of CDK12 or CDK13 induced transcriptional responses associated with DNA-damage and cellular growth signaling pathways respectively, with minimal effects on cell viability. In contrast, dual-kinase inhibition potently induced cell death, which was associated with extensive genome-wide transcriptional changes including wide-spread use of alternative 3’ polyadenylation sites. At the molecular level dual-kinase inhibition resulted in the loss of POLII CTD phosphorylation and greatly reduced POLII elongation rates and processivity. These data define significant redundancy between CDK12 and CDK13, and identify both as fundamental regulators of global POLII processivity and transcription elongation.