Journal
MOLECULAR BIOLOGY OF THE CELL
Volume 17, Issue 12, Pages 4937-4945Publisher
AMER SOC CELL BIOLOGY
DOI: 10.1091/mbc.E06-08-0670
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Funding
- NEI NIH HHS [R01 EY013574, EY13574] Funding Source: Medline
- NHLBI NIH HHS [R01 HL059198, HL-73856, HL-59198, R01 HL073856] Funding Source: Medline
- NIBIB NIH HHS [R37 EB000415, EB-00415, R01 EB000415] Funding Source: Medline
- NIDDK NIH HHS [DK-72517, P30 DK072517, R37 DK035124, R01 DK035124, DK-35124] Funding Source: Medline
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Mutations in cystic fibrosis transmembrane conductance regulator (CFTR), a cAMP-regulated chloride channel, cause cystic fibrosis. To investigate interactions of CFTR in living cells, we measured the diffusion of quantum dot-labeled CFTR molecules by single particle tracking. In multiple cell lines, including airway epithelia, CFTR diffused little in the plasma membrane, generally not moving beyond 100-200 nm. However, CFTR became mobile over micrometer distances after 1) truncations of the carboxy terminus, which contains a C-terminal PDZ (PSD95/Dlg/ZO-1) binding motif; 2) blocking PDZ binding by C-terminal green fluorescent protein fusion, 3) disrupting CFTR association with actin by expression of a mutant EBP50/NHERF1 lacking its ezrin binding domain; or 4) skeletal disruption by latrunculin. CFTR also became mobile when the cytoskeletal adaptor protein binding capacity was saturated by overexpressing CFTR or its C terminus. Our data demonstrate remarkable and previously unrecognized immobilization of CFTR in the plasma membrane and provide direct evidence that C-terminal coupling to the actin skeleton via EBP50/ezrin is responsible for its immobility.
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