Comparison of the gating behaviour of human and murine cystic fibrosis transmembrane conductance regulator Cl- channels expressed in mammalian cells.
1. To investigate the function of the murine cystic fibrosis transmembrane conductance regulator (CFTR), a full-length cDNA encoding wild-type murine CFTR was assembled and stably expressed in Chinese hamster ovary (CHO) cells. 2. Like human CFTR, murine CFTR formed Cl- channels that were regulated by cAMP-dependent phosphorylation and intracellular ATP. However, murine CFTR Cl- channels had a reduced single-channel conductance and decreased open probability (Po) compared with those of human CFTR. 3. Analysis of the dwell time distributions of single channels suggested that the reduced Po of murine CFTR was caused by both decreased residence in the open state and transitions to a new closed state, described by an intermediate closed time constant. 4. For both human and murine CFTR, ATP and ADP regulated the rate of exit from the long-lived closed state. 5. 5'-Adenylylimidodiphosphate (AMP-PNP) and pyrophosphate, two compounds that disrupt cycles of ATP hydrolysis, stabilized the open state of human CFTR. However, neither agent locked murine CFTR Cl- channels open, although AMP-PNP increased the Po of murine CFTR. 6. The data indicate that although human and murine CFTR have many properties in common, some important differences in function are observed. These differences could be exploited in future studies to provide new understanding about CFTR.
Lansdell
K A
KA
Human Genetics Unit, Department of Medicine, University of Edinburgh, Molecular Medicine Centre, Western General Hospital, Edinburgh EH4 2XU, UK.
Delaney
S J
SJ
Lunn
D P
DP
Thomson
S A
SA
Sheppard
D N
DN
Wainwright
B J
BJ
D000244:Adenosine Diphosphate / Q000378:metabolism; D000255:Adenosine Triphosphate / Q000378:metabolism; D000266:Adenylyl Imidodiphosphate / Q000494:pharmacology; D000818:Animals; D002478:Cells, Cultured; D018118:Chloride Channels / Q000187:drug effects / Q000378:metabolism*; D006224:Cricetinae; D000242:Cyclic AMP / Q000378:metabolism; D019005:Cystic Fibrosis Transmembrane Conductance Regulator / Q000235:genetics / Q000378:metabolism*; D004247:DNA / Q000235:genetics; D011756:Diphosphates / Q000494:pharmacology; D004594:Electrophysiology; D006801:Humans; D015640:Ion Channel Gating / Q000187:drug effects / Q000502:physiology*; D008564:Membrane Potentials / Q000187:drug effects / Q000502:physiology; D051379:Mice; D018408:Patch-Clamp Techniques
C505032:CFTR protein, human; D018118:Chloride Channels; D011756:Diphosphates; D019005:Cystic Fibrosis Transmembrane Conductance Regulator; D000266:Adenylyl Imidodiphosphate; D000244:Adenosine Diphosphate; D000255:Adenosine Triphosphate; D004247:DNA; D000242:Cyclic AMP