Cystic fibrosis transmembrane conductance regulator Cl- channels with R domain deletions and translocations show phosphorylation-dependent and -independent activity
O. Baldursson et al., Cystic fibrosis transmembrane conductance regulator Cl- channels with R domain deletions and translocations show phosphorylation-dependent and -independent activity, J BIOL CHEM, 276(3), 2001, pp. 1904-1910
Phosphorylation of the R domain regulates cystic fibrosis transmembrane con
ductance regulator Cl- channel activity. Earlier studies suggested that the
R domain controls activity via more than one mechanism; a phosphorylated R
domain may stimulate activity, and an unphosphorylated R domain may preven
t constitutive activity, i.e, opening with ATP alone. However, the mechanis
ms responsible for these two regulatory properties are not understood. In t
his study we asked whether the two effects are dependent on its position in
the protein and whether smaller regions from the R domain mediate the effe
cts. We found that several portions of the R domain conferred phosphorylati
on-stimulated activity. This was true whether the R domain sequences were p
resent in their normal location or were translocated to the C terminus. We
also found that some parts of the R domain could be deleted without inducin
g constitutive activity. However, when residues 760-783 were deleted, chann
els opened without phosphorylation, Translocation of the R domain to the C
terminus did not prevent constitutive activity, These results suggest that
different parts of the phosphorylated R domain can stimulate activity and t
hat their location within the protein is not critical. In contrast, prevent
ion of constitutive activity required a short specific sequence that could
not be moved to the C terminus. These results are consistent with a recent
model of an R domain composed primarily of random coil in which more than o
ne phosphorylation site is capable of stimulating channel activity, and net
activity reflects interactions between multiple sites in the R domain and
the rest of the channel.