Abstract
In cystic fibrosis, deletion of phenylalanine 508 (F508del) in the cystic fibrosis transmembrane conductance regulator (CFTR) anion channel causes misfolding and premature degradation. One possible approach to reducing the detrimental health effects of cystic fibrosis could be the identification of proteins whose suppression rescues F508del-CFTR function in bronchial epithelial cells. However, searches for these potential targets have not yet been conducted, particularly in a relevant airway background using a functional readout. To identify proteins associated with F508del-CFTR processing, we used a highthroughput functional assay to screen an siRNA library targeting 6,650 different cellular proteins. We identified 37 proteins whose silencing significantly rescued F508del-CFTR activity, as indicated by enhanced anion transport through the plasma membrane. These proteins included FAU, UBE2I, UBA52, MLLT6, UBA2, CHD4, PLXNA1, and TRIM24, among others. We focused our attention on FAU, a poorly characterized protein with unknown function. FAU knockdown increased the plasma membrane targeting and function of F508del-CFTR, but not of wildtype CFTR. Investigation into the mechanism of action revealed a preferential physical interaction of FAU with mutant CFTR, leading to its degradation. FAU and other proteins identified in our screening may offer a therapeutically relevant panel of drug targets to correct basic defects in F508del-CFTR processing.
| Original language | English |
|---|---|
| Pages (from-to) | 1203-1217 |
| Number of pages | 15 |
| Journal | Journal of Biological Chemistry |
| Volume | 293 |
| Issue number | 4 |
| DOIs | |
| Publication status | Published - Jan 1 2018 |
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ASJC Scopus subject areas
- Biochemistry
- Molecular Biology
- Cell Biology
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High-throughput screening identifies FAU protein as a regulator of mutant cystic fibrosis transmembrane conductance regulator channel. / Tomati, Valeria; Pesce, Emanuela; Caci, Emanuela; Sondo, Elvira; Scudieri, Paolo; Marini, Monica; Amato, Felice; Castaldo, Giuseppe; Ravazzolo, Roberto; Galietta, Luis J.V.; Pedemonte, Nicoletta.
In: Journal of Biological Chemistry, Vol. 293, No. 4, 01.01.2018, p. 1203-1217.Research output: Contribution to journal › Article
}
TY - JOUR
T1 - High-throughput screening identifies FAU protein as a regulator of mutant cystic fibrosis transmembrane conductance regulator channel
AU - Tomati, Valeria
AU - Pesce, Emanuela
AU - Caci, Emanuela
AU - Sondo, Elvira
AU - Scudieri, Paolo
AU - Marini, Monica
AU - Amato, Felice
AU - Castaldo, Giuseppe
AU - Ravazzolo, Roberto
AU - Galietta, Luis J.V.
AU - Pedemonte, Nicoletta
PY - 2018/1/1
Y1 - 2018/1/1
N2 - In cystic fibrosis, deletion of phenylalanine 508 (F508del) in the cystic fibrosis transmembrane conductance regulator (CFTR) anion channel causes misfolding and premature degradation. One possible approach to reducing the detrimental health effects of cystic fibrosis could be the identification of proteins whose suppression rescues F508del-CFTR function in bronchial epithelial cells. However, searches for these potential targets have not yet been conducted, particularly in a relevant airway background using a functional readout. To identify proteins associated with F508del-CFTR processing, we used a highthroughput functional assay to screen an siRNA library targeting 6,650 different cellular proteins. We identified 37 proteins whose silencing significantly rescued F508del-CFTR activity, as indicated by enhanced anion transport through the plasma membrane. These proteins included FAU, UBE2I, UBA52, MLLT6, UBA2, CHD4, PLXNA1, and TRIM24, among others. We focused our attention on FAU, a poorly characterized protein with unknown function. FAU knockdown increased the plasma membrane targeting and function of F508del-CFTR, but not of wildtype CFTR. Investigation into the mechanism of action revealed a preferential physical interaction of FAU with mutant CFTR, leading to its degradation. FAU and other proteins identified in our screening may offer a therapeutically relevant panel of drug targets to correct basic defects in F508del-CFTR processing.
AB - In cystic fibrosis, deletion of phenylalanine 508 (F508del) in the cystic fibrosis transmembrane conductance regulator (CFTR) anion channel causes misfolding and premature degradation. One possible approach to reducing the detrimental health effects of cystic fibrosis could be the identification of proteins whose suppression rescues F508del-CFTR function in bronchial epithelial cells. However, searches for these potential targets have not yet been conducted, particularly in a relevant airway background using a functional readout. To identify proteins associated with F508del-CFTR processing, we used a highthroughput functional assay to screen an siRNA library targeting 6,650 different cellular proteins. We identified 37 proteins whose silencing significantly rescued F508del-CFTR activity, as indicated by enhanced anion transport through the plasma membrane. These proteins included FAU, UBE2I, UBA52, MLLT6, UBA2, CHD4, PLXNA1, and TRIM24, among others. We focused our attention on FAU, a poorly characterized protein with unknown function. FAU knockdown increased the plasma membrane targeting and function of F508del-CFTR, but not of wildtype CFTR. Investigation into the mechanism of action revealed a preferential physical interaction of FAU with mutant CFTR, leading to its degradation. FAU and other proteins identified in our screening may offer a therapeutically relevant panel of drug targets to correct basic defects in F508del-CFTR processing.
UR - http://www.scopus.com/inward/record.url?scp=85041203560&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=85041203560&partnerID=8YFLogxK
U2 - 10.1074/jbc.M117.816595
DO - 10.1074/jbc.M117.816595
M3 - Article
C2 - 29158263
AN - SCOPUS:85041203560
VL - 293
SP - 1203
EP - 1217
JO - Journal of Biological Chemistry
JF - Journal of Biological Chemistry
SN - 0021-9258
IS - 4
ER -