Cystic Fibrosis (CF) is principally caused by deletion of Phe 508 in the cystic fibrosis transmembrane conductance regulator (CFTR) protein (?F508), a residue that is critical for both intra-domain and inter-domain contacts controlling thermodynamic stability and channel activity. We now appreciate that folding in the cell is not only about the primary sequence;rather it is managed by an extensive protein homeostasis or 'proteostasis'biology that generates, maintains and destroys the protein fold when it is damaged or no longer required (Science (2008), 319: 916;Science (2010) 329, 766). Because the ?F508 structure is unstable, it is degraded, leading to loss-of-function disease. The Riordan and Balch groups have made considerable progress during the previous funding period to develop structural, biochemical, molecular and morphological insights into CFTR structure and dynamics as well as the proteostasis biology that impacts both wild-type (WT) and ?F508 folding and function. We now hypothesize that the Phe508 deletion is a networking problem at two levels: (1) loss of the energetically stabilizing Phe 508 triggers the loss of key structure-based network interactions within the molecule required to maintain the appropriate balance of dynamics and stability;(2) changes in folding network management by proteostasis biology leading to degradation. We now need to understand these network-based pathways in depth. We further hypothesize that the fold itself and the proteostasis biology managing the fold can be modified to 'repair'?F508 function. This renewal application will systematically study the impact of the structure and proteostasis biology based networks on WT and ?F508 folding to understand why it fails in ?F508 disease. We propose two Aims to address these central issues in CF.
In Aim 1, the Balch lab hypothesizes that understanding the proteostasis biology of WT and ?F508 CFTR folding is an important key to understanding the defect and that this biology is fully accessible for use in its correction.
In Aim 2, the Riordan laboratory hypothesizes that application of computational, biochemical and physiological measures of channel activity and stability for understanding the network of internal structural interactions dictating the WT and ?F508 CFTR folds will provide critical insight into the organization of the fold and the mechanisms by which it can be stabilized by small molecules in ?F508 disease. Integration of the Riordan and Balch efforts will lead to a new understanding of how structure and proteostasis biology work together to achieve biological function. A pathway/network-based approach provides a new opportunity to understand the variables confounding ?F508 function in disease and identify the key nodes in the networks amendable to retune function to improve human healthspan. We hypothesize the general concept that therapeutic management of the emergent properties of folding and proteostasis biology can be used to control the energetics of the CFTR folding landscape and that this new understanding will have a major benefit in the clinic.

Public Health Relevance

Misfolding diseases such as Cystic Fibrosis are a consequence of the failure of protein folding. We will characterize both the structural and proteostasis biology networks required for folding, and function of WT and ?F508 CFTR to restore activity in human disease.

Agency
National Institute of Health (NIH)
Institute
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Type
Research Project (R01)
Project #
2R01DK051870-15
Application #
8238972
Study Section
Lung Cellular, Molecular, and Immunobiology Study Section (LCMI)
Program Officer
Mckeon, Catherine T
Project Start
1996-09-30
Project End
2016-11-30
Budget Start
2011-12-10
Budget End
2012-11-30
Support Year
15
Fiscal Year
2012
Total Cost
$463,527
Indirect Cost
$111,433
Name
University of North Carolina Chapel Hill
Department
Biochemistry
Type
Schools of Medicine
DUNS #
608195277
City
Chapel Hill
State
NC
Country
United States
Zip Code
27599
He, Lihua; Aleksandrov, Andrei A; An, Jianli et al. (2015) Restoration of NBD1 thermal stability is necessary and sufficient to correct ?F508 CFTR folding and assembly. J Mol Biol 427:106-20
Yang, Zhengrong; Wang, Chi; Zhou, Qingxian et al. (2014) Membrane protein stability can be compromised by detergent interactions with the extramembranous soluble domains. Protein Sci 23:769-89
Hamvas, Aaron; Deterding, Robin; Balch, William E et al. (2014) Diffuse lung disease in children: summary of a scientific conference. Pediatr Pulmonol 49:400-9
Balch, William E; Sznajder, Jacob I; Budinger, Scott et al. (2014) Malfolded protein structure and proteostasis in lung diseases. Am J Respir Crit Care Med 189:96-103
Roth, Daniela Martino; Balch, William E (2013) Q-bodies monitor the quinary state of the protein fold. Nat Cell Biol 15:1137-9
Clunes, Lucy A; Davies, Catrin M; Coakley, Raymond D et al. (2012) Cigarette smoke exposure induces CFTR internalization and insolubility, leading to airway surface liquid dehydration. FASEB J 26:533-45
Aleksandrov, Andrei A; Kota, Pradeep; Cui, Liying et al. (2012) Allosteric modulation balances thermodynamic stability and restores function of ?F508 CFTR. J Mol Biol 419:41-60
Bouchecareilh, Marion; Balch, William E (2011) Proteostasis: a new therapeutic paradigm for pulmonary disease. Proc Am Thorac Soc 8:189-95
Cholon, Deborah M; O'Neal, Wanda K; Randell, Scott H et al. (2010) Modulation of endocytic trafficking and apical stability of CFTR in primary human airway epithelial cultures. Am J Physiol Lung Cell Mol Physiol 298:L304-14
He, Lihua; Aleksandrov, Luba A; Cui, Liying et al. (2010) Restoration of domain folding and interdomain assembly by second-site suppressors of the DeltaF508 mutation in CFTR. FASEB J 24:3103-12

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