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Author Notes:

Eric J. Sorscher, Emory University School of Medicine, Health Sciences Research Building, Suite E-280, 1760 Haygood Drive Northeast, Atlanta, Georgia 30322, USA. Phone: 404.727.3293; Email: esorscher@emory.edu

Zoya Ignatova, Institute for Biochemistry and Molecular Biology, University of Hamburg, Martin-Luther-King-Platz 6, Hamburg, Germany 20146. Phone: 49.40.42838.2332; Email: zoya.ignatova@chemie.uni-hamburg.de

KEO, RR, MM, RAK, KLK, AR, IB, JSH, JLH, ZI, and EJS designed studies.

KEO, RR, MM, WW, MJW, and JM conducted experiments.

KEO, RR, MM, WW, JM, CMS, RAK, IB, JLH, ZI, and EJS acquired and analyzed data.

KEO, RR, MM, IB, JLH, ZI, and EJS wrote the manuscript.

We thank David Bedwell (UAB) and David Schneider (UAB) for helpful discussions and technical guidance.

JLH is founder of and holds equity in Spectrum PhenomX LLC, which aims to commercialize quantitative high-throughput cell array phenotyping (Q-HTCP) technology.

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Research Funding:

Funding was provided by the NIH (F31HL131231, R01HL136414, P30DK072482), the Cystic Fibrosis Foundation (OLIVER17F0, BRAAKM14XXO, HARTMA15G0, IGNATO17XXO, SORSCH13XXO, SORSCH14XXO), the Burroughs Wellcome Fund (Collaborative Research Travel Grant, 1018774), Deutsche Forschungsgemeinschaft FOR1805 (IG74/14-1, IG74/14-2), the German Cystic Fibrosis Foundation (muko e.V. 1603), and the Netherlands Science Organization (graduate school program grant).

Keywords:

  • Science & Technology
  • Life Sciences & Biomedicine
  • Medicine, Research & Experimental
  • Research & Experimental Medicine
  • TRANSMEMBRANE CONDUCTANCE REGULATOR
  • CYSTIC-FIBROSIS
  • DELTA-F508 CFTR
  • MESSENGER-RNA
  • IN-VIVO
  • PROTEIN
  • TRANSLATION
  • MUTATION
  • RESCUE
  • DOMAIN

Slowing ribosome velocity restores folding and function of mutant CFTR

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Journal Title:

JOURNAL OF CLINICAL INVESTIGATION

Volume:

Volume 129, Number 12

Publisher:

, Pages 5236-5253

Type of Work:

Article | Final Publisher PDF

Abstract:

Cystic fibrosis (CF) is caused by mutations in the CF transmembrane conductance regulator (CFTR), with approximately 90% of patients harboring at least one copy of the disease-associated variant F508del. We utilized a yeast phenomic system to identify genetic modifiers of F508del-CFTR biogenesis, from which ribosomal protein L12 (RPL12/uL11) emerged as a molecular target. In the present study, we investigated mechanism(s) by which suppression of RPL12 rescues F508del protein synthesis and activity. Using ribosome profiling, we found that rates of translation initiation and elongation were markedly slowed by RPL12 silencing. However, proteolytic stability and patch-clamp assays revealed RPL12 depletion significantly increased F508del-CFTR steady-state expression, interdomain assembly, and baseline open-channel probability. We next evaluated whether Rpl12-corrected F508del-CFTR could be further enhanced with concomitant pharmacologic repair (e.g., using clinically approved modulators lumacaftor and tezacaftor) and demonstrated additivity of these treatments. Rpl12 knockdown also partially restored maturation of specific CFTR variants in addition to F508del, and WT Cftr biogenesis was enhanced in the pancreas, colon, and ileum of Rpl12 haplosufficient mice. Modulation of ribosome velocity therefore represents a robust method for understanding both CF pathogenesis and therapeutic response.

Copyright information:

© 2019, American Society for Clinical Investigation.

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