Biophysical characterisation of calumenin as a charged F508del-CFTR folding modulator

PLoS One. 2014 Aug 13;9(8):e104970. doi: 10.1371/journal.pone.0104970. eCollection 2014.

Abstract

The cystic fibrosis transmembrane regulator (CFTR) is a cyclic-AMP dependent chloride channel expressed at the apical surface of epithelial cells lining various organs such as the respiratory tract. Defective processing and functioning of this protein caused by mutations in the CFTR gene results in loss of ionic balance, defective mucus clearance, increased proliferation of biofilms and inflammation of human airways observed in cystic fibrosis (CF) patients. The process by which CFTR folds and matures under the influence of various chaperones in the secretory pathway remains incompletely understood. Recently, calumenin, a secretory protein, belonging to the CREC family of low affinity calcium binding proteins has been identified as a putative CFTR chaperone whose biophysical properties and functions remain uncharacterized. We compared hydropathy, instability, charge, unfoldability, disorder and aggregation propensity of calumenin and other CREC family members with CFTR associated chaperones and calcium binding proteins, wild-type and mutant CFTR proteins and intrinsically disordered proteins (IDPs). We observed that calumenin, along with other CREC proteins, was significantly more charged and less folded compared to CFTR associated chaperones. Moreover like IDPs, calumenin and other CREC proteins were found to be less hydrophobic and aggregation prone. Phylogenetic analysis revealed a close link between calumenin and other CREC proteins indicating how evolution might have shaped their similar biophysical properties. Experimentally, calumenin was observed to significantly reduce F508del-CFTR aggregation in a manner similar to AavLEA1, a well-characterized IDP. Fluorescence microscopy based imaging analysis also revealed altered trafficking of calumenin in bronchial cells expressing F508del-CFTR, indicating its direct role in the pathophysiology of CF. In conclusion, calumenin is characterized as a charged protein exhibiting close similarity with IDPs and is hypothesized to regulate F508del-CFTR folding by electrostatic effects. This work provides useful insights for designing optimized synthetic structural correctors of CFTR mutant proteins in the future.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Calcium-Binding Proteins / chemistry
  • Calcium-Binding Proteins / genetics
  • Calcium-Binding Proteins / metabolism*
  • Cystic Fibrosis / genetics
  • Cystic Fibrosis / metabolism*
  • Cystic Fibrosis Transmembrane Conductance Regulator / genetics
  • Cystic Fibrosis Transmembrane Conductance Regulator / metabolism*
  • Humans
  • Mutation
  • Phylogeny
  • Protein Aggregation, Pathological / genetics
  • Protein Aggregation, Pathological / metabolism
  • Protein Conformation
  • Protein Folding*
  • Protein Transport
  • Static Electricity

Substances

  • CALU protein, human
  • Calcium-Binding Proteins
  • Cystic Fibrosis Transmembrane Conductance Regulator

Grants and funding

The authors thank the Association Gaétan Saleün, the French foundation Vaincre la Mucoviscidose, Conseil Général de Bretagne (http://www.conseil-general.com/conseil-regional/conseil-regional-bretagne.htm) and Inserm (http://www.inserm.fr) for their financial support. BC was funded by an Irish Research Council EMBARK post-graduate fellowship. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.