Free calcium transients in chemotactic and non-chemotactic strains of Escherichia coli determined by using recombinant aequorin

Abstract

Intracellular Ca2+ has been previously implicated in the chemotactic response of Escherichia coli. However, no correlative measurements of intracellular free Ca2+ have been made during bacterial chemotaxis, essential if this is to be established. In order to monitor internal free Ca2+ in E. coli during challenge with chemotactic agents, the Ca(2+)-activated photoprotein aequorin was expressed in a chemotactic strain (AB1157) and a non-chemotactic strain [BL21(DE3)] of E. coli. Repellents were found to cause an increase (50-150 nM) in intracellular free Ca2+, whereas attractants caused a small but consistent decrease in intracellular free Ca2+. These data are in agreement with the proposed model that an increase in intracellular free Ca2+ causes tumbling. The effect of increasing external Ca2+ on the regulation of intracellular free Ca2+ in both strains was monitored by using aequorin. The resting level of free Ca2+ in E. coli (AB1157) was found to be 100 nM, which agrees with previous data [Gangola and Rosen (1987) J. Biol. Chem. 262, 12570-12574]. As these results also show differences in the regulation of intracellular free Ca2+ between the two strains in the presence of high external Ca2+ concentrations, this may have implications for the effect of high-Ca2+ environments on E. coli.

A calcium signal is involved in heterocyst differentiation in the cyanobacterium Anabaena sp. PCC7120

“…Since Ca 2+ was first studied in bacteria fifty years ago (Norris & Jensen, 1957) it has been implicated in a broad range of physiological processes in prokaryotes that include chemotaxis and motility (Tisa & Adler, 1992Watkins et al, 1995;Pitta et al, 1997), pathogenesis (Rose et al, 1993;Straley et al, 1993), the cell cycle and the control of the initiation of replication (Jiménez-Sanchez et al, 1993;Yu & Margolin, 1997), quorum sensing (Wherten & Lundgren, 2001) and spore and fruiting body formation (Inouye et al, 1983;O'Hara & Hageman, 1990). In many cases, the involvement of Ca 2+ in the regulation of cellular processes has been roughly described in terms of an influx or efflux of Ca 2+ from the cytosol.…”

Use of Recombinant Aequorin to Study Calcium Homeostasis and Monitor Calcium Transients in Response to Heat and Cold Shock in Cyanobacteria

“…The fluorescent dyes show a general resistance to entry into plant cells (Bush and Jones, 1990), and in bacteria, considerable problems have been encountered with dye loading and autofluorescence (Gangola and Rosen, 1987). Fortunately, the possibility of transforming animal, plant, and bacterial cells with the Ca 2ϩ -bindingsensitive luminescent protein apoaequorin (Knight et al, 1991a(Knight et al, , 1991bBrini et al, 1995) has allowed the quantitation of intracellular Ca 2ϩ fluxes accompanying diverse stimuli (Knight et al, 1991a(Knight et al, , 1991bWatkins et al, 1995;Okazaki et al, 1996;Sedbrook et al, 1996;Takahashi et al, 1997;Chandra and Low, 1997;Gong et al, 1998;Volotovski et al, 1998).…”

Crystallization and preliminary X-ray diffraction studies of the calcium-binding protein CalD fromStreptomyces coelicolor

“…In contrast to eukaryotes, calcium-mediated regulatory roles are less evident in prokaryotes, despite accumulated evidence that Ca 2+ is involved in a myriad of physiologically important bacterial activities, including cell division (Yu & Margolin, 1997), chemotaxis (Tisa & Adler, 1995;Watkins et al, 1995) and cell differentiation (Norris et al, 1991;Michiels et al, 2002;Yonekawa et al, 2001). Calerythrin, a well studied CaBP from Saccharopolyspora erythraea, was the first prokaryotic calcium-binding protein identified to contain four canonical calcium-binding EF-hand motifs (Leadlay et al, 1984;Swan et al, 1987) and was proposed to function as a Ca 2+ buffer rather than as a Ca 2+ sensor (Cox & Bairoch, 1988).…”