Dose-dependent activation of gene expression is achieved using CRISPR and small molecules that recruit endogenous chromatin machinery.

Anna M Chiarella1, Kyle V Butler2, Berkley E Gryder3, Dongbo Lu1, Tiffany A Wang1, Xufen Yu2, Silvia Pomella3,4, Javed Khan3, Jian Jin5, Nathaniel A Hathaway6

  1. Division of Chemical Biology and Medicinal Chemistry, Center for Integrative Chemical Biology and Drug Discovery, UNC Eshelman School of Pharmacy, Chapel Hill, NC, USA.
  2. Mount Sinai Center for Therapeutics Discovery, Departments of Pharmacological Sciences and Oncological Sciences, Tisch Cancer Institute, Icahn School of Medicine at Mount Sinai, New York, NY, USA.
  3. Oncogenomics Section, Genetics Branch, National Cancer Institute, National Institutes of Health (NIH), Bethesda, MD, USA.
  4. Department of Pediatric Hematology and Oncology, Bambino Gesu' Children's Hospital, Rome, Italy.
  5. Mount Sinai Center for Therapeutics Discovery, Departments of Pharmacological Sciences and Oncological Sciences, Tisch Cancer Institute, Icahn School of Medicine at Mount Sinai, New York, NY, USA. jian.jin@mssm.edu.
  6. Division of Chemical Biology and Medicinal Chemistry, Center for Integrative Chemical Biology and Drug Discovery, UNC Eshelman School of Pharmacy, Chapel Hill, NC, USA. hathaway@unc.edu.

Abstract

Gene expression can be activated or suppressed using CRISPR--Cas9 systems. However, tools that enable dose-dependent activation of gene expression without the use of exogenous transcription regulatory proteins are lacking. Here we describe chemical epigenetic modifiers (CEMs) designed to activate the expression of target genes by recruiting components of the endogenous chromatin-activating machinery, eliminating the need for exogenous transcriptional activators. The system has two parts: catalytically inactive Cas9 (dCas9) in complex with FK506-binding protein (FKBP) and a CEM consisting of FK506 linked to a molecule that interacts with cellular epigenetic machinery. We show that CEMs upregulate gene expression at target endogenous loci up to 20-fold or more depending on the gene. We also demonstrate dose-dependent control of transcriptional activation, function across multiple diverse genes, reversibility of CEM activity and specificity of our best-in-class CEM across the genome.

Presented by Anna Chiarella

Genomics, Biochemistry, Genetics & Genome BiologyNicholas Liao2020-01 (Jan)