Project description:Macrophages adopt dynamic functional states to maintain tissue homeostasis or respond to environmental challenges. Here we performed a genome-wide CRISPR screen in primary BMDMs identifiying negative regulators of iNOS expression in response to IFNg stimulation.
Project description:WT BMDMs were nucleofected with sgRNA/Cas9 RNPs to induce knockouts of genes encoding lysosomal proteins of interest, and bulk RNA-seq was performed at day 4 and d13 post-CRISPR.
Project description:Previous single-cell RNA sequencing of microglia from aged WT and Grn KO mice revealed a microglial subset with transcriptomic signatures that were highly specific to Grn KO. Further epigenetic analysis demonstrated that these signatures were likely regulated by Mitf/Tfe family transcription factors. To investigate the necessity of these transcription factors in mediating the Grn KO transcriptomic response, we turned to an experimental in vitro validation. A multiplexed CRISPR approach was used to knockout Mitf, Tfe3 and Tfeb in BMDMs from WT or Grn KO mice. 10 days post-CRISPR, bulk RNA sequencing was performed to analyze the transcriptional profiles. These profiles identified the expected enrichment of Grn KO-specific genes in BMDMs from Grn KO mouse background. Strikingly, the levels of many of these genes were reversed with knockout of Mitf, Tfe3 and Tfeb.
Project description:Haploid human embryonic stem cells harboring TP53 or MLH1 knockout (KO) were subjected to a genome-wide CRISPR-Cas9 knockout screen to identify synthetic lethal interactions associated with the mentioned genes.