Figure 4. Loss of mcu-1 triggers... | Rockefeller University Press

Figure 4.

Loss of mcu-1 triggers plasma-membrane Ca2+oscillations within synapses of ADF neurons and activates intestinal UPRmt. (A) Representative fluorescence traces of average background-subtracted fluorescence intensity ΔF/F0 of plasma membrane Ca2+ indicator GCaMP6f of WT and mcu-1(ju1154) mutant animals in the 60 s. (B) Quantification of PM-GCaMP6 maximal fluorescence intensity changes in WT and mcu-1(ju1154) animals. n ≥ 8 worms. (C) Quantification of the frequency with GCaMP6f fluorescence intensity changes in WT and mcu-1(ju1154) animals. n ≥ 20 worms. (D) Representative confocal photomicrographs of TMBIM-2::GFP animals with the presence or absence of mcu-1(ju1154). The imaging used z-planes. Scale bar, 10 μm. (E) Representative photomicrographs of dve-1 reporter expression in WT and ADF neuron mcu-1 knockdown animals (srh-142p::Cas9+u6p::mcu-1 sgRNA). Scale bar, 250 μm. (F) Quantification of the number of intestinal nuclei puncta with GFP signal per worm in E. n ≥ 12 worms. (G) Deletions of mcu-1 by CRISPR/Cas9 are detected by T7E1 assay. Representative DNA gels of T7E1 assay show mcu-1 PCR products amplified from genomic DNA of WT or srh-142p::Cas9+u6p::mcu-1-sg worms. ***P < 0.001; **P < 0.01 via unpaired two-tailed Student’s t test. Error bars, SEM. Source data are available for this figure: SourceData F4.

Loss of mcu-1 triggers plasma-membrane Ca ²⁺ oscillations within synapses of ADF neurons and activates intestinal UPR ^mt . (A) Representative fluorescence traces of average background-subtracted fluorescence intensity ΔF/F0 of plasma membrane Ca²⁺ indicator GCaMP6f of WT and mcu-1(ju1154) mutant animals in the 60 s. (B) Quantification of PM-GCaMP6 maximal fluorescence intensity changes in WT and mcu-1(ju1154) animals. n ≥ 8 worms. (C) Quantification of the frequency with GCaMP6f fluorescence intensity changes in WT and mcu-1(ju1154) animals. n ≥ 20 worms. (D) Representative confocal photomicrographs of TMBIM-2::GFP animals with the presence or absence of mcu-1(ju1154). The imaging used z-planes. Scale bar, 10 μm. (E) Representative photomicrographs of dve-1 reporter expression in WT and ADF neuron mcu-1 knockdown animals (srh-142p::Cas9+u6p::mcu-1 sgRNA). Scale bar, 250 μm. (F) Quantification of the number of intestinal nuclei puncta with GFP signal per worm in E. n ≥ 12 worms. (G) Deletions of mcu-1 by CRISPR/Cas9 are detected by T7E1 assay. Representative DNA gels of T7E1 assay show mcu-1 PCR products amplified from genomic DNA of WT or srh-142p::Cas9+u6p::mcu-1-sg worms. ***P < 0.001; **P < 0.01 via unpaired two-tailed Student’s t test. Error bars, SEM. Source data are available for this figure: SourceData F4.

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