Identification of Full-Length Wild-Type and Mutant Huntingtin Interacting Proteins by Crosslinking Immunoprecipitation in Mice Brain Cortex

Karen A. Sap, Arzu Tugce Guler, Aleksandra Bury, Dick Dekkers, Jeroen A. A. Demmers, Eric A. Reits

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14 Citations (Scopus)


Background: Huntington's disease is a neurodegenerative disorder caused by a CAG expansion in the huntingtin gene, resulting in a polyglutamine expansion in the ubiquitously expressed mutant huntingtin protein. Objective: Here we set out to identify proteins interacting with the full-length wild-type and mutant huntingtin protein in the mice cortex brain region to understand affected biological processes in Huntington's disease pathology. Methods: Full-length huntingtin with 20 and 140 polyQ repeats were formaldehyde-crosslinked and isolated via their N-terminal Flag-tag from 2-month-old mice brain cortex. Interacting proteins were identified and quantified by label-free liquid chromatography-mass spectrometry (LC-MS/MS). Results: We identified 30 interactors specific for wild-type huntingtin, 14 interactors specific for mutant huntingtin and 14 shared interactors that interacted with both wild-type and mutant huntingtin, including known interactors such as F8a1/Hap40. Syt1, Ykt6, and Snap47, involved in vesicle transport and exocytosis, were among the proteins that interacted specifically with wild-type huntingtin. Various other proteins involved in energy metabolism and mitochondria were also found to associate predominantly with wild-type huntingtin, whereas mutant huntingtin interacted with proteins involved in translation including Mapk3, Eif3h and Eef1a2. Conclusion: Here we identified both shared and specific interactors of wild-type and mutant huntingtin, which are involved in different biological processes including exocytosis, vesicle transport, translation and metabolism. These findings contribute to the understanding of the roles that wild-type and mutant huntingtin play in a variety of cellular processes both in healthy conditions and Huntington's disease pathology.
Original languageEnglish
Pages (from-to)335-347
Number of pages13
JournalJournal of Huntington's Disease
Issue number3
Publication statusPublished - 2021


  • Huntington's disease
  • cerebral cortex
  • cross-linking reagents
  • huntingtin protein
  • immunoprecipitation
  • protein interaction mapping
  • proteomics

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