Cystic fibrosis transmembrane conductance regulator (CFTR) channel regulatory (R) domain  -  Cystic fibrosis transmembrane conductance regulator (CFTR) channel

Biological function
CFTR is a multifunctional protein, which provides the pore of a linear conductance chloride channel and also functions to regulate other membrane proteins. CFTR is regulated by phosphorylation at the intrinsically disordered regulatory (R) region. The R region has multiple effects on CFTR: it can contribute to activation or inhibition of the channel depending on the position and number of phosphorylation. The nucleotide binding domain (NBD1) preferably interacts with the non-phosphorylated R region. Binding occurs in a dynamic fashion with multiple, exchanging R sites.

Structural evidence
Assignments were obtained for 97% of the 1H, 15N, 13Cα, 13Cβ and C’ resonances of the nonphosphorylated R-region sequence and 99% of the resonances of the phosphorylated sequence, using a variety of triple-resonance experiments. Residues 718-722 in the nonphosphorylated R region lack assignments because of resonance broadening (loss of intensity accompanied by an increase in NMR resonance linewidth), probably resulting from millisecond- to microsecond-timescale sampling of a small population of stabilized conformations for these residues. Distinct segments of the R region have fractionally populated local helical structure, reflecting a bias of these residues toward helical conformations in the pool of conformers. In the nonphosphorylated R region, residues 654-668, 759- 764, 766-776 and 801-817 all have a greater than 5% a-helical population. Phosphorylation produces a global decrease in helical content, consistent with that observed in circular dichroism experiments. Phosphorylation generally decreases R2 relaxation rates. Upon adding NBD1, ratios of peak intensities varied from 0 to 1.3, with broad rather than sharp minima, reflective of longer stretches of residues interacting with varying affinities and no global disorder-to-order transition. Several segments of the R region appear to bind NBD1, to various degrees, implying dynamic exchange of several R-region binding segments on and off NBD1. The presence of fractional helical structure in the free state of the R region and changes in resonance intensity over several stretches of 10-15 residues upon binding to NBD1 provide evidence that R- region interactions are mediated by stabilization of fluctuating helical structural elements.

Biochemical evidence
Phosphorylation reduces helical propensity and reduces NBD1 binding.

Transient helices of R are stabilized upon the interaction. The interactions established by multiple regions could explain the lack of a specific phosphorylation site to regulate the interaction. Transient interactions between multiple R segments and the NBD1 also modulate the equilibrium of the NBD1 - NBD2 dimerization, which ultimately results in channel opening.

Mechanism category
conformational selection

Posttranslational modification
CFTR activity is regulated by R region phosphorylation, which reduces the propensity of transient helices. The R regions behavior is a ’rheostat’, which gradually tunes channel activity.

Isoforms, context-dependence
Interactions between NBD1 and R are sensitive to various interaction partners.

Dynamic interactions modulated by phosphorylation of the fuzzy region regulate channel opening via variable interactions with the NBD1 domains.

Medical relevance
Mutations in CFTR leading to defective regulation or transport of chloride ions across the apical surface of epithelial cells are the primary cause of the genetic disease cystic fibrosis.