Extended Data Fig. 1: Characterization of SNAP-Omp25 mouse.
From: Old mitochondria regulate niche renewal via α-ketoglutarate metabolism in stem cells
a, Schematic of lox-Stop-lox-SNAPtag-Omp25-construct. b, Left, Representative image of SNAP-Omp25 expression in Lgr5-EGFP positive crypts 5 days after a single dose of 80 mg/kg of tamoxifen in a Lgr5EGFP-IRES-creERT2/+Rosa26lox-Stop-lox-SNAPtag-omp25/+ mouse repeated independently four times. Scale bar 25 µm. Right, Representative image of SNAP-Omp25 localization to the outer membrane of mitochondria in organoids. Repeated independently three times. Scale bar 1 µm. c, Relative weight change over 18 months after a single dose of 80 mg/kg of tamoxifen. Monthly data points represent mean and colored area between dashed lines show the 50% interquartile range). (n = 6) two-tailed Student’s unpaired t-test at 18 months d, Flow cytometer frequencies of SNAP+ cells out of LiveEpcam+ cells two weeks after recombination (3 months) and 15 months after recombination (18 months) (n = 4) two-tailed Student’s unpaired t-test. Data mean ± SD. e, Flow cytometer frequencies of crypt cells: Paneth cells (CD24hiSSChi) and ISCs (Cd24medLgr5-EGFPhi) out of LiveEpcam+ cells. (n = 3) Data mean±S.D. f, Top, Schematic of in vivo labelling for mitochondrial age classes. Bottom, different labelling time-windows to establish optimal young mitochondria label (histogram showing Snap staining when young label is given at indicated time after snap-block.) g, Frequencies of crypts with indicated number of ISCsmito-O in in vivo labelled tissue. (n = 3) One-way ANOVA with two-tailed Dunnett’s post-hoc test. Data mean±S.D. h, Left, Flow cytometer frequencies of ISCsmito-O out of all ISCs (Cd24medLgr5-EGFPhi). (n = 5). Data mean ± SD. Right, Representative FACS pseudo color plot of ISCsmito-O gating out of all ISCs (Cd24medLgr5-EGFPhi). For full sort scheme see Supplementary Fig. 1.
