A Chemical Biology Approach to Model Pontocerebellar Hypoplasia Type 1B (PCH1B)

  • ACS Chem Biol. 2018 Oct 19;13(10):3000-3010. doi: 10.1021/acschembio.8b00745.
Liberty François-Moutal  1  2 ,  Shahriyar Jahanbakhsh  3 ,  Andrew D L Nelson  4 ,  Debashish Ray  5 ,  David D Scott  1  2 ,  Matthew R Hennefarth  3 ,  Aubin Moutal  1 ,  Samantha Perez-Miller  1  2 ,  Andrew J Ambrose  6 ,  Ahmed Al-Shamari  1 ,  Philippe Coursodon  1 ,  Bessie Meechoovet  7 ,  Rebecca Reiman  7 ,  Eric Lyons  4 ,  Mark Beilstein  4 ,  Eli Chapman  6 ,  Quaid D Morris  5  8  9  10 ,  Kendall Van Keuren-Jensen  7 ,  Timothy R Hughes  5  8 ,  Rajesh Khanna  1  2 ,  Carla Koehler  3 ,  Joanna Jen  11 ,  Vijay Gokhale  12 ,  May Khanna  1  2
Affiliations
  • 1. Department of Pharmacology , College of Medicine, University of Arizona , Tucson , Arizona 85724 , United States.
  • 2. Center for Innovation in Brain Science , Tucson , Arizona 85721 , United States.
  • 3. Department of Chemistry and Biochemistry , University of California , Los Angeles , California 90095 , United States.
  • 4. School of Plant Sciences , University of Arizona , Tucson , Arizona 85721 , United States.
  • 5. Donnelly Centre , University of Toronto , Toronto , Canada M5S 3E1.
  • 6. Pharmacology and Toxicology, School of Pharmacy , University of Arizona , Tucson , Arizona 85724 , United States.
  • 7. Neurogenomics Division , TGen , Phoenix , Arizona 85004 , United States.
  • 8. Department of Molecular Genetics , University of Toronto , Toronto , Canada M5S 1A8.
  • 9. Department of Computer Science , University of Toronto , Toronto , Canada M5S 2E4.
  • 10. Department of Electrical and Computer Engineering , University of Toronto , Toronto , Canada M5S3G4.
  • 11. Mount Sinai , New York , New York 10029 , United States.
  • 12. Bio5 Institute , University of Arizona , Tucson , Arizona , United States.
PMID: 30141626 DOI: 10.1021/acschembio.8b00745
Abstract

Mutations of EXOSC3 have been linked to the rare neurological disorder known as Pontocerebellar Hypoplasia type 1B (PCH1B). EXOSC3 is one of three putative RNA-binding structural cap proteins that guide RNA into the RNA exosome, the cellular machinery that degrades RNA. Using RNAcompete, we identified a G-rich RNA motif binding to EXOSC3. Surface plasmon resonance (SPR) and microscale thermophoresis (MST) indicated an affinity in the low micromolar range of EXOSC3 for long and short G-rich RNA sequences. Although several PCH1B-causing mutations in EXOSC3 did not engage a specific RNA motif as shown by RNAcompete, they exhibited lower binding affinity to G-rich RNA as demonstrated by MST. To test the hypothesis that modification of the RNA-protein interface in EXOSC3 mutants may be phenocopied by small molecules, we performed an in-silico screen of 50 000 small molecules and used enzyme-linked immunosorbant assays (ELISAs) and MST to assess the ability of the molecules to inhibit RNA-binding by EXOSC3. We identified a small molecule, EXOSC3-RNA disrupting (ERD) compound 3 (ERD03), which ( i) bound specifically to EXOSC3 in saturation transfer difference nuclear magnetic resonance (STD-NMR), ( ii) disrupted the EXOSC3-RNA interaction in a concentration-dependent manner, and ( iii) produced a PCH1B-like phenotype with a 50% reduction in the cerebellum and an abnormally curved spine in zebrafish embryos. This compound also induced modification of zebrafish RNA expression levels similar to that observed with a morpholino against EXOSC3. To our knowledge, this is the first example of a small molecule obtained by rational design that models the abnormal developmental effects of a neurodegenerative disease in a whole organism.

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