Skip to main content
. Author manuscript; available in PMC: 2012 Nov 1.
Published in final edited form as: Cell Metab. 2011 Sep 7;14(3):428–434. doi: 10.1016/j.cmet.2011.07.010

Figure 3. Patient fibroblasts have a defect in Met-tRNAMet formylation.

Figure 3

(A) In metazoan mitochondria, a single tRNAMet species acts as both initiator and elongator tRNAMet. After aminoacylation of tRNAMet by the mitochondrial methionyl-tRNA synthetase (MetRSmt), a portion of Met-tRNAMet is formylated by MTFMT to generate fMet-tRNAMet. fMet-tRNAMet is used by the mitochondrial IF2 (IF2mt) to initiate translation, whereas Met-tRNAMet is recognized by the mitochondrial EF-Tu (EF-Tumt) for the elongation of translation products. (B) Total RNA from control (lanes 5 – 7) and patient fibroblasts (P1: lanes 8 – 10; P2: lanes 11 – 13) was separated by acid-urea PAGE. Total RNA from MCH58 cells is shown as a reference (lanes 1 – 4). The mitochondrial tRNAMet (top panel) and the cytoplasmic initiator tRNAiMet (bottom panel) were detected by Northern hybridization. Total RNA was isolated under acidic conditions, which preserves both the Met-tRNAMet and fMet-tRNAMet (ac); tRNAs were treated with copper sulfate (Cu2+), which specifically deacylates Met-tRNAMet, but not fMet-tRNAMet; or with base (OH-) which deacylates both Met-tRNAMet and fMet-tRNAMet. Base treated tRNA was re-aminoacylated in vitro with Met using MetRS generating a Met-tRNAMet standard (M).