ATP-regulated chloride conductance in endoplasmic reticulum (ER)-enriched pig pancreas microsomes.
The Cl- conductance of endoplasmic reticulum-enriched pancreatic microsomes was identified. Its regulation by nucleotides was investigated by measuring the rate of cation ionophore-induced microsome swelling in the presence of an inward Cl- gradient. The conductance was solubilized and reconstituted into liposomes. The Cl- conductance in intact microsomes was inhibited by stilbene (10(-4) M) and indanyloxyacetic acid (10(-5) M) derivatives. ATP increased Cl- conductance with half-maximal stimulation at 8 x 10(-6) M. Other trinucleotides (GTP, CTP and UTP) were without effect at 10(-4) M. The non-hydrolysable analogue of ATP, adenosine 5'-[beta gamma-methylene]triphosphate (AppCH2p) increased Cl- conductance with a potency similar to that of ATP. The same concentration of adenosine 5'-[gamma-thio]triphosphate (ATP gamma S) which is a substrate for kinases, had no effect. ATP stimulation of Cl- conductance was inhibited by stilbene derivatives. The data suggest the presence of at least one ATP-binding site, and show that the ATP does not need to be hydrolyzed and that its spatial conformation is important for activating the Cl- conductance. Solubilized microsomal proteins reconstituted into liposomes retained their stilbene-inhibited, ATP-stimulated Cl- conductance. A 167 kDa protein was detected by anti-CFTR antibodies in the intact microsomes, but not in the solubilized proteins. The 64 kDa protein (a component of a ubiquitous Cl- channel) was detected in the both intact and solubilized microsomes. These results suggest that this Cl- conductance is not a CFTR protein.
Bégault
B
B
INSERUM U.323 CHU Necker, Paris, France.
Anagnostopoulos
T
T
Edelman
A
A
D000255:Adenosine Triphosphate / Q000031:analogs & derivatives*; D000818:Animals; D001665:Binding Sites; D018118:Chloride Channels / Q000378:metabolism; D002712:Chlorides / Q000378:metabolism*; D004553:Electric Conductivity / Q000187:drug effects; D004721:Endoplasmic Reticulum / Q000378:metabolism*; D006016:Glycolates / Q000494:pharmacology; D008081:Liposomes / Q000737:chemistry; D008861:Microsomes / Q000378:metabolism; D010179:Pancreas / Q000378:metabolism* / Q000648:ultrastructure; D013267:Stilbenes / Q000494:pharmacology; D013552:Swine
D018118:Chloride Channels; D002712:Chlorides; D006016:Glycolates; D008081:Liposomes; D013267:Stilbenes; C015765:MK 473; D000255:Adenosine Triphosphate