PH-SENSITIVE ACTIVATION OF THE INTRACELLULAR-PH REGULATION SYSTEM IN SQUID AXONS BY ATP-GAMMA-S
PH-SENSITIVE ACTIVATION OF THE INTRACELLULAR-PH REGULATION SYSTEM IN SQUID AXONS BY ATP-GAMMA-S
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DOI：
10.1038/332262a0
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发表时间：
1988-03-17
期刊：
影响因子：
64.8
通讯作者：
RUSSELL, JM
中科院分区：
文献类型：
--
作者：
BORON, WF;HOGAN, E;RUSSELL, JM
The regulation of intracellularpH (pHi) is essential for normal cell function1, and controlled changes inpHimay play a central role in cell activation2. Sodium-dependent Cl–HCO3exchange is the dominant mechanism ofpHiregulation in the invertebrate cells examined3–6, and also occurs in mammalian cells7,8. The transporter extrudes acid from the cell by exchanging extracellular Na+and HCO3−(ref. 9) (or a related species) for intracellular Cl−(refs 3, 4). It is blocked by the stilbene derivatives DIDS (4,4′-diisothiocyano-stilbene-2,2′-disulphonate, ref. 10) and SITS (4-acetamido-4′-isothiocyano-stilbene-2,2′-disulphonate, ref. 3), and has a stoichiometry of two intracellular H+neutralized for each Na+taken up and each Cl–extruded by the axon11. Because the inwardly-directed Na+concentration gradient is sufficiently large to energize both the HCO3−influx and Cl−efflux, this electroneutral exchanger could be a classic secondary active transporter, thermodynamically independent of ATP hydrolysis. However, at least in the squid axon, the exchanger has an absolute requirement for ATP (ref. 3). Thus, a major unresolved issue is whether this Na-dependent Cl–HCO3exchanger stoichiometri-cally hydrolyses ATP (the pump hypothesis), or whether ATP activates the transporter by a mechanism such as phosphorylation or simple binding (the activation hypothesis). We have now explored the role of ATP inpHiregulation by dialysing axons with the ATP analogue ATP-γ-S. In many systems, ATP–γ–S is an acceptable substrate for protein kinases12,13, whereas the resulting thiophosphorylated proteins are not as readily hydrolysed by phosphatases as are phosphorylated proteins14,15. Our results rule out the pump hypothesis, and show that the basis of the axon's ATP requirement is thepH-dependent activation (by, for instance, phosphorylation or ATP binding) of the exchanger itself, or of an essential activator.
