Protein kinase A phosphorylation potentiates cystic fibrosis transmembrane conductance regulator gating by relieving autoinhibition on the stimulatory C terminus of the regulatory domain.
J Biol Chem
; 295(14): 4577-4590, 2020 04 03.
Article
em En
| MEDLINE
| ID: mdl-32102849
ABSTRACT
Cystic fibrosis transmembrane conductance regulator (CFTR) is a chloride channel activated by protein kinase A (PKA) phosphorylation on the regulatory (R) domain. Phosphorylation at several R domain residues stimulates ATP-dependent channel openings and closings, termed channel gating. To explore the protein segment responsible for channel potentiation and PKA-dependent activation, deletion mutations were constructed by removing one to three protein segments of the R domain including residues 708-759 (ΔR708-759), R760-783, and R784-835, each of which contains one or two PKA phosphorylation sites. Deletion of R708-759 or R760-783 had little effect on CFTR gating, whereas all mutations lacking R784-835 reduced CFTR activity by decreasing the mean burst duration and increasing the interburst interval (IBI). The data suggest that R784-835 plays a major role in stimulating CFTR gating. For ATP-associated regulation, ΔR784-835 had minor impact on gating potentiation by 2'dATP, CaATP, and pyrophosphate. Interestingly, introducing a phosphorylated peptide matching R809-835 shortened the IBI of ΔR708-835-CFTR. Consistently, ΔR815-835, but not ΔR784-814, enhanced IBI, whereas both reduced mean burst duration. These data suggest that the entirety of R784-835 is required for stabilizing the open state of CFTR; however, R815-835, through interactions with the channel, is dominant for enhancing the opening rate. Of note, PKA markedly decreased the IBI of ΔR708-783-CFTR. Conversely, the IBI of ΔR708-814-CFTR was short and PKA-independent. These data reveal that for stimulating CFTR gating, PKA phosphorylation may relieve R784-814-mediated autoinhibition that prevents IBI shortening by R815-835 This mechanism may elucidate how the R domain potentiates channel gating and may unveil CFTR stimulation by other protein kinases.
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Texto completo:
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Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Proteínas Quinases Dependentes de AMP Cíclico
/
Regulador de Condutância Transmembrana em Fibrose Cística
Limite:
Humans
Idioma:
En
Revista:
J Biol Chem
Ano de publicação:
2020
Tipo de documento:
Article
País de afiliação:
China