Fig. 4: Sec22b affects global but not local calcium signaling. | Communications Biology

Fig. 4: Sec22b affects global but not local calcium signaling.

From: Sec22b regulates phagosome maturation by promoting ORP8-mediated lipid exchange at endoplasmic reticulum-phagosome contact sites

Fig. 4

a–c Fura-2 analysis (F340/F380 ratio) of store-operated calcium influx after 8 min of 1 µM Tg/2 mM Ca2+ re-addition in a WT MEFs transfected with YFP-STIM1 and mCh-Sec22b or RFP-KDEL controls (n = 4/5 biologically independent samples; 124/163 cells KDEL/Sec22b), in b WT MEFs transfected only with GFP-KDEL, EGFP-Sec22b, or EGFP-Sec22b-P33 (n = 13/20/19 biologically independent samples, 124/217/117 cells KDEL/Sec22b/P33) and in c WT and Stim1−/− shCTR and shSec22b stable cell lines (n = 15/13;19/20 biologically independent samples; 537/408;702/484 cells WT;Stim1−/− shCTR/shSec22b). d Quantification of STIM1 expression by Western blot in WT MEF shCTR and shSec22b (n = 8 biologically independent samples; see also Supplementary Fig. S3b). e Quantification of percent Sec22b knockdown by Western blot in Stim1−/− and Stim1−/−;Stim2−/− MEF shCTR and shSec22b (n = 5/5 biologically independent samples; see also Supplementary Fig. S3a, b). f Quantification of periphagosomal calcium hotspots (arrows) and g phagocytic index in shCTR and shSec22b phagocytic WT MEFs loaded with Fluo-8 (blue-green ratio pseudocolor, representing fluorescence over mean cytosolic fluorescence F/Fave) in cells exposed to IgG-RBC (magenta) for 30 min (n = 7/8 biologically independent samples; 589/847 phagosomes; 256/366 cells shCTR/shSec22b). White bar = 3 µm. Bar graphs show means +SEM. White bars = 3 µm. Bar graphs show means +SEM. Error bars are omitted from traces in (a–c) for clarity.

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