Extended Data Fig. 5: Identification of LONP1 interacting proteins by TurboID-mediated proximity labeling.
From: Proteolytic rewiring of mitochondria by LONP1 directs cell identity switching of adipocytes
(a) Heat map showing species of fatty acids and carnitines identified by LC-MS system in in vitro-differentiated WT and LONP1 AKO beige adipocytes. n = 5 per group. (b, c) The levels of indicated species of carnitine (b) and fatty acid (c) in in vitro-differentiated WT and LONP1 AKO beige adipocytes. n = 5 per group. (d) Schematic of LONP1-TurboID reconstitution using the proximity-dependent biotinylation catalyzed by promiscuous biotin ligases. LONP1-N-TurboID constructs (mitochondrial targeted sequence; substrate binding ___domain of LONP1; full-length TurboID; HA-tag), or the reference constructs TurboID (mitochondrial targeted sequence; full-length TurboID; HA-tag). (e) Immunoblot analysis of total biotinylated proteins in 293 T cells using Streptavidin-HRP antibody. (f) The percentage of mitochondrial proteins within all identified biotinylated proteins using MitoCarta2.0. (g) Functional protein association networks analysis of identified mitochondrial proteins. (h, i) Blue native polyacrylamide gel electrophoresis (BN-PAGE) analyses of the mitochondrial respiratory chain complexes in WT and LONP1 AKO adipocytes followed by immunoblot analysis (h) using antibodies against complex I (anti-NDUFS1), complex II (anti-SDHA), complex III (anti-UQCRC2), complex IV (anti-MTCO1) and complex V (anti-ATP5A) or in-gel activity of complex I (i). Data shown are representative examples of 3 independent experiments. All data are shown as the mean ± SEM. P value was calculated by unpaired two-sided Student’s t-test. Source numerical data and unprocessed blots are available in source data.
