Chemogenomic profiling of endogenous PARK2 expression using a genome-edited coincidence reporter.

Paradigms and Technologies
Therapeutic Approaches

Abstract

Parkin, an E3 ubiquitin ligase, is a central mediator of mitochondrial quality control and is linked to familial forms of Parkinson's disease (PD). Removal of dysfunctional mitochondria from the cell by Parkin is thought to be neuroprotective, and pharmacologically increasing Parkin levels may be a novel therapeutic approach. We used genome-editing to integrate a coincidence reporter into the PARK2 gene locus of a neuroblastoma-derived cell line and developed a quantitative high-throughput screening (qHTS) assay capable of accurately detecting subtle compound-mediated increases in endogenous PARK2 expression. Interrogation of a chemogenomic library revealed diverse chemical classes that up-regulate the PARK2 transcript, including epigenetic agents, drugs controlling cholesterol biosynthesis, and JNK inhibitors. Use of the coincidence reporter eliminated wasted time pursuing reporter-biased false positives accounting for ∼2/3 of the actives and, coupled with titration-based screening, greatly improves the efficiency of compound selection. This approach represents a strategy to revitalize reporter-gene assays for drug discovery.

Authors

Hasson, Samuel A; Fogel, Adam I; Wang, Chunxin; MacArthur, Ryan; Guha, Rajarshi; Heman-Ackah, Sabrina; Martin, Scott; Youle, Richard J; Inglese, James;

Keywords

  • Cell Line, Tumor
  • Cholesterol/ biosynthesis
  • Epigenesis, Genetic
  • Gene Expression Profiling
  • Genes, Reporter
  • Genomics
  • High-Throughput Screening Assays
  • Humans
  • MAP Kinase Kinase 4/ antagonists & inhibitors
  • Ubiquitin-Protein Ligases/ genetics

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