Renal cortical basolateral Na+/HCO3- cotransporter. III. Evidence for a regulatory protein in the inhibitory effect of protein kinase A.
The activity of the Na-H antiporter is inhibited by cyclic AMP-dependent protein kinase A (cAMP-PKA). The inhibitory effect of PKA on the Na-H antiporter is mediated through a regulatory protein that can be dissociated from the antiporter by limited protein digestion. PKA also inhibits the activity of the Na+/HCO3- cotransporter. We investigated whether the activity of Na+/HCO3- cotransporter and the effect of PKA on this transporter may also be regulated by limited protein digestion. In rabbit renal cortical basolateral membranes (BLM) and in solubilized BLM reconstituted in liposomes (proteoliposomes), trypsin (100 micrograms) increased 22Na uptake in the presence of HCO3 but not in the presence of gluconate, indicating that trypsin does not alter diffusive 22Na uptake but directly stimulates the Na+/HCO3- cotransporter activity. In proteoliposomes phosphorylated with ATP, the catalytic subunit (CSU) of cAMP-PKA decreased the activity of the Na+/HCO3- cotransporter (expressed as nanomoles/mg protein/3s) from 23 +/- 10 to 14 +/- 6 (P < 0.01). In the presence of trypsin, the inhibitory effect of CSU of cAMP-PKA on the activity of Na+/HCO3- cotransporter was blunted. To identify a fraction that was responsible for the inhibitory effect of the CSU on the Na+/HCO3- cotransporter activity, solubilized proteins were separated by size exclusion chromatography. The effect of CSU of cAMP-PKA on the Na+/HCO3- cotransporter activity was assayed in proteoliposomes digested with trypsin with the addition of a fraction containing the 42 kDa protein (fraction S+) or without the 42 kDa protein (fraction S-).(ABSTRACT TRUNCATED AT 250 WORDS)
Bernardo
A A
AA
Section of Nephrology, University of Illinois at Chicago, USA.
Kear
F T
FT
Stim
J A
JA
Qiu
Y Y
YY
Ruiz
O S
OS
Weidman
H
H
Arruda
J A
JA
D000818:Animals; D002462:Cell Membrane / Q000378:metabolism; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism* / Q000494:pharmacology; D017136:Ion Transport / Q000187:drug effects; D007672:Kidney Cortex / Q000378:metabolism*; D011188:Potassium / Q000378:metabolism*; D011817:Rabbits; D017923:Sodium-Hydrogen Exchangers / Q000378:metabolism*
D017923:Sodium-Hydrogen Exchangers; D017868:Cyclic AMP-Dependent Protein Kinases; D011188:Potassium