Graduate Student Wake Forest School of Medicine, Wake Forest School of Medicine
Erin Hughes (Wake Forest School of Medicine, Wake Forest School of Medicine)| William Booth (Wake Forest School of Medicine)| Jill Clodfelter (Wake Forest School of Medicine)| Sandra Leone-Kabler (Wake Forest School of Medicine)| Allyn Howlett (Wake Forest School of Medicine, Wake Forest School of Medicine)| W. Lowther (Wake Forest School of Medicine, Wake Forest School of Medicine)
Cannabinoid receptor interacting protein 1a (CRIP1a) is an 18 kDa protein that regulates the CB1 cannabinoid receptor by modifying G-protein-mediated signaling and competing for β-arrestin interactions with the receptor (Booth et al., Molecules. 2019; 24: 3672). Physiological functions associated with CRIP1a include involvement in certain cancers, embryonic development, and functions of the retina (reviewed in Oliver et al., Biomolecules. 2020, 10: 1609). We determined the CRIP1a structure in an effort to elucidate CRIP1a functions. The 1.55 Å resolution crystal structure shows that CRIP1a exhibits a 10-stranded, antiparallel b-barrel with an interior comprised of conserved hydrophobic residues and loops at the bottom and a short helical cap at the top that serve to exclude solvent. The DALI server identified CRIP1a as homologous to a family of lipidated protein-carriers. Based upon this structural knowledge, we tested the hypothesis that CRIP1a could serve as a carrier of lipidated proteins from cytosol to membranes. Using a mouse neuroblastoma N18TG2 cultured cell model, homogenates were sedimented, and cytosolic and membrane fractions were subjected to SDS-page electrophoresis and western analysis. We determined that CRIP1a is found in both cytosolic and membrane fractions. Immunoprecipitations using a CRIP1a antibody in the absence of detergents showed an apparent molecular weight shift for CRIP1a upon western blot analysis compared with purified, recombinant CRIP1a or CRIP1a in detergent extracts. The molecular weight shift in CRIP1a suggests the formation of a complex or interaction with other cellular proteins consistent with a role for CRIP1a in trafficking of cargo proteins.
Support or Funding Information
This work was supported by NIH grants R01-DA042157 and K12-GM102773.