| First Author | Manjunath LE | Year | 2020 |
| Journal | J Biol Chem | Volume | 295 |
| Issue | 50 | Pages | 17009-17026 |
| PubMed ID | 33028634 | Mgi Jnum | J:299407 |
| Mgi Id | MGI:6488249 | Doi | 10.1074/jbc.RA120.014253 |
| Citation | Manjunath LE, et al. (2020) Stop codon read-through of mammalian MTCH2 leading to an unstable isoform regulates mitochondrial membrane potential. J Biol Chem 295(50):17009-17026 |
| abstractText | Stop codon read-through (SCR) is a process of continuation of translation beyond a stop codon. This phenomenon, which occurs only in certain mRNAs under specific conditions, leads to a longer isoform with properties different from that of the canonical isoform. MTCH2, which encodes a mitochondrial protein that regulates mitochondrial metabolism, was selected as a potential read-through candidate based on evolutionary conservation observed in the proximal region of its 3' UTR. Here, we demonstrate translational read-through across two evolutionarily conserved, in-frame stop codons of MTCH2 using luminescence- and fluorescence-based assays, and by analyzing ribosome-profiling and mass spectrometry (MS) data. This phenomenon generates two isoforms, MTCH2x and MTCH2xx (single- and double-SCR products, respectively), in addition to the canonical isoform MTCH2, from the same mRNA. Our experiments revealed that a cis-acting 12-nucleotide sequence in the proximal 3' UTR of MTCH2 is the necessary signal for SCR. Functional characterization showed that MTCH2 and MTCH2x were localized to mitochondria with a long t 1/2 (>36 h). However, MTCH2xx was found predominantly in the cytoplasm. This mislocalization and its unique C terminus led to increased degradation, as shown by greatly reduced t 1/2 (<1 h). MTCH2 read-through-deficient cells, generated using CRISPR-Cas9, showed increased MTCH2 expression and, consistent with this, decreased mitochondrial membrane potential. Thus, double-SCR of MTCH2 regulates its own expression levels contributing toward the maintenance of normal mitochondrial membrane potential. |
