Journal: Nature Communications
Article Title: Coupling of ribosome biogenesis and translation initiation in human mitochondria
doi: 10.1038/s41467-025-58827-x
Figure Lengend Snippet: a MTG3 FLAG co-isolates mtSSU MRPs and several mtSSU assembly factors. FLAG-immunoprecipitation was performed with lysed mitochondria from HEK293 wild type (WT) cells and a stable HEK293 cell line inducibly expressing MTG3 FLAG , and subsequently analyzed via western blotting with indicated antibodies (total = 3%, eluate = 100%). Similar results were obtained in n ≥ 3 biologically independent experiments. b Schematic depiction of the mature 12S rRNA secondary structure. Regions are depicted by their level of maturation in each state, with M corresponding to the mature mtSSU (PDB: 3J9M ;). c Cryo-EM structures of the MTG3-TFB1M-mtRBFA (in) -bound small mitoribosomal subunit (mtSSU) intermediate (state A), METTL15-mtRBFA (in) -bound mtSSU (state B), and METTL15-mtRBFA (out) -bound mtSSU (state C). The 12S rRNA (red) and indicated biogenesis factors are shown as cartoon and the remaining mitoribosomal proteins (MRPs) are indicated as white transparent surface. MTG3: light blue, TFB1M: lime green, mtRBFA: indigo, METTL15: turquoise, mS38: beige . Close-up views of the immature decoding center ( d ) and of the foot region ( e ) in state A. Coloring as in ( c ), with cryo-EM densities (from map A3) of immature rRNA helices shown as red surface. Close-up views of the immature decoding center with cryo-EM densities (from maps B-C3) ( f , h ) and of the foot region ( g , i ) in state B and C, respectively. ( e , g , i ) Close-up views of the foot region in each state, showing the densities for MTG3 (state A, map A3), or MTG3 NTD and h44 with altered trajectory (state B-C, 15 Å low-pass filtered maps B1 and C1). Mature h44 is depicted by red dashed lines and would clash with MTG3 and MTG3 NTD .
Article Snippet: HEK293 cell lines (Human Embryonic Kidney 293-Flp-In T-Rex, Thermo Fisher Scientific; R78007) were cultured in DMEM (Dulbecco’s Modified Eagle’s Medium) supplemented with 10% FCS (Fetal Calf Serum), 2 mM L-glutamine, 1 mM sodium pyruvate and 50 μg/ml uridine under standard cultivation conditions (37 °C under 5% CO 2 humidified atmosphere).
Techniques: Immunoprecipitation, Expressing, Western Blot, Cryo-EM Sample Prep
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