Supplementary File: Simultaneous Measurement of Changes in Mitochondrial and Endoplasmic Reticulum Free Calcium in Pancreatic Beta Cells

The following supporting information can be downloaded at: https://zenodo.org/record/7735311#.ZBExInZBxEY, Movie S1: Image series of DS-1-transduced INS1-832/13 cells treated by agents known to modulate the ER and mitochondria Ca²⁺. Green represents the ER Ca²⁺, and red represents the mitochondria Ca²⁺. The caption in the movie shows the treatment conditions. The movie shows the images taken once every 10 s; Figure S1: The figure (a–c) shows correlations between BFP-KDEL, MitoTracker, or G-CEPIA versus G-CEPIA, R-CEPIA and R-CEPIA, respectively; r values are Pearson’s correlation coefficients. Images shown at the bottom (in d) show green and red signals were nonoverlapping; Figure S2: To confirm that that the green and red fluorescent signals from G-CEPIA-er and R-CEPIA3-mt do not overlap, images acquired separately using argon (green) and neon (red) lasers are shown here. All images were acquired from cells recorded in standard imaging solution containing 11 mM glucose. DS-1 yielded signals in both the green and red channels, while G-CEPIA-er and R-CEPIA3-mt were only visible on their respective channels; Figure S3: Representative time courses of ER and Mito signals recorded from individual cells shown in Figure 2. Figure S4: Time courses of G-CEPIA-er traces where only the ER probe was transfected into cells; Figure S5: Time courses of R-CEPIA3-mt traces where only the Mito probe was transfected into cells; Figure S6: Representative ER and Mito traces shown for individual cells from Figure 3; Figure S7: (a) Plasmid map of DS-1. (b) Translated protein sequence for the DNA coding DS-1; Figure S8: Annotated sequence of DS-1 for (a) DNA and (b) amino acids. The G-CEPIA-er is shown in green and codon optimized R-CEPIA3-mt is shown in red. The T2A sequence is highlighted in yellow.