Mitochondrial localization as a determinant of capacitative Ca2+ entry in HeLa cells.
Whether different subsets of mitochondria play distinct roles in shaping intracellular Ca2+ signals is presently unresolved. Here, we determine the role of mitochondria located beneath the plasma membrane in controlling (a) Ca2+ release from the endoplasmic reticulum (ER) and (b) capacitative Ca2+ entry. By over-expression of the dynactin subunit dynamitin, and consequent inhibition of the fission factor, dynamin-related protein (Drp-1), mitochondria were relocalised from the plasma membrane towards the nuclear periphery in HeLa cells. The impact of these changes on free calcium concentration in the cytosol ([Ca2+]c), mitochondria ([Ca2+]m) and ER ([Ca2+]ER) was then monitored with specifically-targeted aequorins. Whilst dynamitin over-expression increased the number of close contacts between the ER and mitochondria by >2.5-fold, assessed using organelle-targeted GFP variants, histamine-induced changes in organellar [Ca2+] were unaffected. By contrast, Ca2+ influx elicited significantly smaller increases in [Ca2+]c and [Ca2+]m in dynamitin-expressing than in control cells. These data suggest that the strategic localisation of a subset of mitochondria beneath the plasma membrane is required for normal Ca2+ influx, but that the transfer of Ca2+ ions between the ER and mitochondria is relatively insensitive to gross changes in the spatial relationship between these two organelles.
Henry Wellcome Laboratories for Integrated Cell Signalling and Department of Biochemistry, School of Medical Sciences, University of Bristol, University Walk, Bristol BS8 1TD, UK.
Aniko
A
Varadi
Vincenzo
V
Cirulli
Guy A
GA
Rutter
D002118:Calcium / Q000032:analysis / Q000378:metabolism*; D015220:Calcium Channels / Q000032:analysis / Q000378:metabolism*; D000072159:Dynactin Complex; D006367:HeLa Cells; D006801:Humans; D008869:Microtubule-Associated Proteins / Q000032:analysis / Q000096:biosynthesis; D008928:Mitochondria / Q000737:chemistry* / Q000378:metabolism*
D015220:Calcium Channels; C000606361:DCTN2 protein, human; D000072159:Dynactin Complex; D008869:Microtubule-Associated Proteins; D002118:Calcium