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. 2006 Dec 18;175(6):901–911. doi: 10.1083/jcb.200608073

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Copyright © 2006, The Rockefeller University Press

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Figure 5. — Coupling of the ER and mitochondrial Ca²⁺ channels depends on the presence of grp75. Mitochondrial Ca²⁺ uptake was measured in control siRNA–transfected HeLa cells (control); after siRNA-driven down-regulation of grp75 (siRNA-grp75); control siRNA and OMM-IP₃R-LBD_224-605–transfected cells; and siRNA-grp75 and OMM-IP₃R-LBD_224-605 cotransfected cells. Cells were also cotransfected with the mtAEQmut probe and mitochondrial Ca²⁺ response to 100 μM histamine was measured as described in the Materials and methods. Inset shows the effect of grp75 siRNA on grp75 levels after 24 h of transfection. Controls transfected only with Lipofectamine showed no difference in respect to control siRNA (not depicted). (B) Silencing of grp75 reverts the stimulatory effect of IP₃R-LBD_224-605 targeted both to the OMM and ER surface. The percent increase of [Ca²⁺]_m peaks normalized to the mean of controls are shown in cells cotransfected with mtAEQmut and control siRNA (siRNA-grp75) and OMM-IP₃R-LBD_224-605 or ER-IP₃R-LBD_224-605 after stimulation with 100 μM histamine. The stimulatory effect of both the OMM- and ER-targeted IP₃R-LBD_224-605 was inhibited after the cotransfection with siRNA-grp75 (+), whereas the control Ca²⁺ peaks remained unaffected. Data normalized to the mean ± the SEM of the control group are shown in percentages. For absolute values see Table S1. *, P < 0.05; **, P < 0.01.