pH and monovalent cations regulate cytosolic free Ca2+ in E. coli

The results here show for the first time that pH and monovalent cations can regulate cytosolic free Ca2+ in E. coli through Ca2+ influx and efflux, monitored using aequorin. At pH 7.5 the resting cytosolic free Ca2+ was 0.2-0.5 µM. In the presence of external Ca2+ (1 mM) at alkaline pH this rose to 4 µM, being reduced to 0.9 µM at acid pH. Removal of external Ca2+ caused an immediate decrease in cytosolic free Ca2+ at 50-100nM s− 1. Efflux rates were the same at pH 5.5, 7.5 and 9.5. Thus, ChaA, a putative Ca2+/H+exchanger, appeared not to be a major Ca2+-efflux pathway. In the absence of added Na+, but with 1 mM external Ca2+, cytosolic free Ca2+ rose to approximately 10 µM. The addition of Na+(half maximum 60 mM) largely blocked this increase and immediately stimulated Ca2+ efflux. However, this effect was not specific, since K+ also stimulated efflux. In contrast, an increase in osmotic pressure by addition of sucrose did not significantly stimulate Ca2+ efflux. The results were consistent with H+ and monovalent cations competing with Ca2+ for a non-selective ion influx channel. Ca2+ entry and efflux in chaA and yrbG knockouts were not significantly different from wild type, confirming that neither ChaA nor YrbG appear to play a major role in regulating cytosolic Ca2+ in Escherichia coli. The number of Ca2+ ions calculated to move per cell per second ranged from < 1 to 100, depending on conditions. Yet a single eukaryote Ca2+ channel, conductance 100 pS, should conduct > 6 million ions per second. This raises fundamental questions about the nature and regulation of Ca2+ transport in bacteria, and other small living systems such as mitochondria, requiring a new mathematical approach to describe such ion movements. The results have important significance in the adaptation of E. coli to different ionic environments such as the gut, fresh water and in sea water near sewage effluents.