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Am J Physiol Cell Physiol 264: C925-C931, 1993;
0363-6143/93 $5.00
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AJP - Cell Physiology, Vol 264, Issue 4 C925-C931, Copyright © 1993 by American Physiological Society

ARTICLES

Regulation of CFTR Cl- conductance in secretion by cellular energy levels

C. L. Bell and P. M. Quinton
Division of Biomedical Sciences, University of California, Riverside 92521.

Recent studies suggested dual regulation of the Cl- conductance (GCl) affected in cystic fibrosis, one by protein kinase A-dependent phosphorylation and a second by low-affinity ATP binding. We proposed that ATP binding may couple the transport demands to the energy level of the cell. In the present study we examined this hypothesis further in a purely secretory function using the epithelial cell line T84. We used a depletion-permeabilization protocol on cells grown on permeable supports to deplete the cells of endogenous ATP and to provide access to the intracellular compartment for the impermeable nucleotides adenosine 3',5'-cyclic monophosphate (cAMP) and ATP. In contrast to non-depleted permeabilized cells, which responded to 0.1 mM cAMP with an increase in transepithelial potential (delta Vt = 29.8 +/- 3.0 mV, n = 4) and conductance (delta Gt = 1.23 +/- 0.54 mS/cm2, n = 4), addition of cAMP to ATP-depleted cells resulted in insignificant changes in Vt (delta Vt = 0.7 +/- 0.2 mV, n = 26; P < 0.05) and Gt (delta Gt = 0.020 +/- 0.003 mS/cm2, n = 26; P < 0.05). However, the cAMP response was restored by addition of 5 mM ATP (delta Vt = 21.7 +/- 1.5 mV, n = 26; delta Gt = 0.59 +/- 0.06 mS/cm2, n = 26). ATP dose-response experiments, taken together with the effect of cAMP with and without ATP, suggest that phosphorylation is necessary, but not sufficient, for activation. The data provide evidence for a second level of regulation of GCl, which requires high concentrations of ATP.(ABSTRACT TRUNCATED AT 250 WORDS)


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