Extended Data Fig. 2: Validation of C-terminal tagged ALPK3 hPSC cell lines. | Nature Cardiovascular Research

Extended Data Fig. 2: Validation of C-terminal tagged ALPK3 hPSC cell lines.

From: Alpha kinase 3 signaling at the M-band maintains sarcomere integrity and proteostasis in striated muscle

Extended Data Fig. 2: Validation of C-terminal tagged ALPK3 hPSC cell lines.

a. Gene targeting outline of ALPK3-tdTomato hPSC cell line and Sanger sequencing confirmation of junction. b. Gene targeting outline of ALPK3-SBP3xFLAG hPSC cell line and Sanger sequencing confirmation of junction of gene fusion. c. Stem cell markers of ALPK3-SBP3xFLAG and ALPK3-tdTomato hPSC cell lines. Scale bar = 50 µm. d. Representative immunofluorescent micrograph of ALPK3-SBP3xFLAG and WT hPSC-CM’s stained for OBSCN (green), FLAG (magenta), and DAPI (blue) or MYBPC3 (green), FLAG (magenta), and DAPI (blue). Scale bar = 20 µm. e. Western blot of WT and ALPK3-SBP3xFLAG hPSC-CMs immunoprecipitated with anti-FLAG antibody. f. Flow cytometry of ALPK3-tdTomato stained for stromal cell marker CD90. g. Violin plot of ALPK3 expression across 23 cell types identified in mouse skeletal muscle. h. Representative immunofluorescent micrograph of WT and ALPK3-tdTomato hPSC-skeletal muscle stained for alpha-actinin (green), tdTomato (magenta), and DAPI. Scale bar = 15 µm. Line profile demonstrating alpha-actinin and ALPK3tdTomato organization. i. Immunofluorescence of WT and ALPK3mut hCOs staining alpha-actinin (magenta) and obscurin (green), scale bar = 10 µm.

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