The most common mutation of the Cystic Fibrosis Transmembrane Conductance Regulator (CFTR), deltaF508-CFTR, is a trafficking mutant that is retained in the endoplasmic reticulum (ER) and targeted for rapid intracellular degradation, at least in part by the ubiquitin/proteasome system. We have previously demonstrated that this trafficking defect can be repaired in vitro, and partially in vivo, by the pharmaceutical agent Sodium 4- Phenylbutyrate (4PBA). However, the mechanism by which 4PBA repairs deltaF508-CFTR trafficking is not known, 4PBA is a known regulator of gene transcription, although, at effective concentrations in vitro, it does not significantly alter CFTR mRNA levels. Given this evidence, the overall hypothesis of this proposal is that 4PBA repairs the intracellular trafficking of deltaF508-CFTR by regulation of a protein or proteins important in the regulation of folding of nascent proteins and targeting of misfolded proteins for intracellular degradation. Specifically, we will test the hypothesis that 4PBA repairs deltaF508-CFTR intracellular trafficking by down-regulating the expression of Hsc70, the constitutively expressed member of the 70 kDa heat shock protein family that is necessary for the ubiquitination and proteasome degradation of a number of cellular proteins. This specific hypothesis is addressed in the present proposal in the studies directed at the following principal specific aims. (1) To determine whether specific modulation of Hsc70 expression in vitro by means other than 4PBA treatment results in alterations of deltaF508-CFTR and wild type CFTR intracellular trafficking. (2) To determine the mechanism by which 4PBA regulates Hsc70 at the protein and mRNA level by examining the influence of 4PBA on the kinetics of synthesis and degradation of Hsc70 protein and mRNA.

Agency
National Institute of Health (NIH)
Institute
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Type
Research Project (R01)
Project #
5R01DK058046-04
Application #
6614007
Study Section
Medical Biochemistry Study Section (MEDB)
Program Officer
Mckeon, Catherine T
Project Start
2000-09-01
Project End
2004-07-31
Budget Start
2003-08-01
Budget End
2004-07-31
Support Year
4
Fiscal Year
2003
Total Cost
$255,000
Indirect Cost
Name
Children's Hospital of Philadelphia
Department
Type
DUNS #
073757627
City
Philadelphia
State
PA
Country
United States
Zip Code
19104
Grumbach, Yael; Bikard, Yann; Suaud, Laurence et al. (2014) ERp29 regulates epithelial sodium channel functional expression by promoting channel cleavage. Am J Physiol Cell Physiol 307:C701-9
Chanoux, Rebecca A; Shubin, Calla B; Robay, Amal et al. (2013) Hsc70 negatively regulates epithelial sodium channel trafficking at multiple sites in epithelial cells. Am J Physiol Cell Physiol 305:C776-87
Chanoux, Rebecca A; Robay, Amal; Shubin, Calla B et al. (2012) Hsp70 promotes epithelial sodium channel functional expression by increasing its association with coat complex II and its exit from endoplasmic reticulum. J Biol Chem 287:19255-65
Liu, Ji; Lu, Wennan; Guha, Sonia et al. (2012) Cystic fibrosis transmembrane conductance regulator contributes to reacidification of alkalinized lysosomes in RPE cells. Am J Physiol Cell Physiol 303:C160-9
Mueller, Gunhild M; Yan, Wusheng; Copelovitch, Lawrence et al. (2012) Multiple residues in the distal C terminus of the ?-subunit have roles in modulating human epithelial sodium channel activity. Am J Physiol Renal Physiol 303:F220-8
Suaud, Laurence; Miller, Katelyn; Alvey, Lora et al. (2011) ERp29 regulates DeltaF508 and wild-type cystic fibrosis transmembrane conductance regulator (CFTR) trafficking to the plasma membrane in cystic fibrosis (CF) and non-CF epithelial cells. J Biol Chem 286:21239-53
Suaud, Laurence; Miller, Katelyn; Panichelli, Ashley E et al. (2011) 4-Phenylbutyrate stimulates Hsp70 expression through the Elp2 component of elongator and STAT-3 in cystic fibrosis epithelial cells. J Biol Chem 286:45083-92
Rubenstein, Ronald C; Lockwood, Shannon R; Lide, Ellen et al. (2011) Regulation of endogenous ENaC functional expression by CFTR and ýýF508-CFTR in airway epithelial cells. Am J Physiol Lung Cell Mol Physiol 300:L88-L101
Das, Shamie; Smith, Tekla D; Sarma, Jayasri Das et al. (2009) ERp29 restricts Connexin43 oligomerization in the endoplasmic reticulum. Mol Biol Cell 20:2593-604
Yan, Wusheng; Spruce, Lynn; Rosenblatt, Michael M et al. (2007) Intracellular trafficking of a polymorphism in the COOH terminus of the alpha-subunit of the human epithelial sodium channel is modulated by casein kinase 1. Am J Physiol Renal Physiol 293:F868-76

Showing the most recent 10 out of 20 publications