Regulation of renal Na+/HCO3- cotransporter stimulation by CO2: role of phosphorylation, exocytosis and protein synthesis.
The sodium bicarbonate cotransporter (NBC1) mediates bicarbonate reabsorption in the renal proximal tubule. NBC1 activity is stimulated by 10% CO2, however, the mechanism is poorly understood. Here, we examined the mechanism of NBC1 regulation by 10% CO2 using an immortalized human proximal tubule cell line (HK2). In cells exposed to 10% CO2, the cotransporter activity (measured as deltapH/min) increased within minutes and this increase was maintained for 6 to 24 h. Early NBC1 stimulation was accompanied by increased NBC1 phosphorylation. Basolateral membrane NBC1 protein increased by 30 min and reached a maximum at 6 h. Increased NBC activity at 6 h was accounted for by increased NBC exocytosis to the basolateral membrane and not by decreased endocytosis. Latruncullin B (an actin cytoskeleton inhibitor) did not prevent CO2-induced stimulation, while nocodazole (a microtubule-disrupting agent) abrogated the stimulatory effect of 10% CO2. A significant increase in NBC1 mRNA expression level was observed at 6 h and maintained for 24 h. Total NBC1 protein increased at 12 to 24 h with 10% CO2 incubation and this effect was blocked by cycloheximide. In summary, the present study demonstrates that early activation of NBC1 activity by 10% CO2 was mediated by NBC1 phosphorylation. The stimulation of cotransporter activity observed at 6 h was due to exocytosis, while the late effect starting from 12 h was accounted for by increased protein synthesis.
Espiritu
D J D
DJ
Section of Nephrology, Department of Medicine, University of Illinois at Chicago, Chicago, IL 60612-7315, USA.
Yang
V L
VL
Bernardo
A A
AA
Arruda
J A L
JA
D000199:Actins / Q000187:drug effects / Q000502:physiology*; D001639:Bicarbonates / Q000378:metabolism*; D002245:Carbon Dioxide / Q000494:pharmacology*; D002478:Cells, Cultured; D003513:Cycloheximide / Q000494:pharmacology; D005089:Exocytosis / Q000187:drug effects / Q000502:physiology*; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007687:Kidney Tubules, Proximal / Q000378:metabolism*; D015739:Nocodazole / Q000494:pharmacology; D010766:Phosphorylation; D011500:Protein Synthesis Inhibitors / Q000494:pharmacology; D012333:RNA, Messenger / Q000378:metabolism; D027982:Sodium-Bicarbonate Symporters / Q000378:metabolism*
D000199:Actins; D001639:Bicarbonates; D011500:Protein Synthesis Inhibitors; D012333:RNA, Messenger; D027982:Sodium-Bicarbonate Symporters; D002245:Carbon Dioxide; D003513:Cycloheximide; D015739:Nocodazole