{"title":"Selection of initiator tRNA and start codon by mammalian mitochondrial initiation factor 3 in leaderless mRNA translation","authors":"Muhoon Lee, Taisei Wakigawa, Qimin Jia, Chang Liu, Ruiyuan Huang, Shuai Huang, Asuteka Nagao, Tsutomu Suzuki, Kozo Tomita, Shintaro Iwasaki, Nono Takeuchi-Tomita","doi":"10.1093/nar/gkaf021","DOIUrl":null,"url":null,"abstract":"The mammalian mitochondrial protein synthesis system produces 13 essential subunits of oxidative phosphorylation (OXPHOS) complexes. Translation initiation in mammalian mitochondria is characterized by the use of leaderless messenger RNAs (mRNAs) and non-AUG start codons, where the proofreading function of IF-3mt still remains elusive. Here, we developed a reconstituted mammalian mitochondrial translation system using in vitro transcribed and native mitochondrial transfer RNAs (tRNAs) to investigate IF-3mt’s proofreading function. Similar to bacterial IF-3, IF-3mt permits an initiator tRNA to participate in initiation by discriminating the three G–C pairs in its anticodon stem, and by the cognate interactions of its anticodon with the AUG start codon. As a result, IF-3mt promotes the accurate initiation of leaderless mRNAs. Nevertheless, IF-3mt can also facilitate initiation from the non-AUG(AUA) start codon through its unique N- and C-terminal extensions, in concert with the 5-methylcytidine (m5C) or 5-formylcytidine (f5C) modification at the anticodon wobble position of mt-tRNAMet. This is partly because the IF-3mt-specific N- and C-terminal extensions and the KKGK-motif favor leaderless mRNA initiation and relax non-AUG start codon discrimination. Analyses of IF-3mt-depleted human cells revealed that IF-3mt indeed participates in translating the open reading frames (ORFs) of leaderless mRNAs, as well as the internal ORFs of dicistronic mRNAs.","PeriodicalId":19471,"journal":{"name":"Nucleic Acids Research","volume":"29 1","pages":""},"PeriodicalIF":16.6000,"publicationDate":"2025-01-29","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":"0","resultStr":null,"platform":"Semanticscholar","paperid":null,"PeriodicalName":"Nucleic Acids Research","FirstCategoryId":"99","ListUrlMain":"https://doi.org/10.1093/nar/gkaf021","RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q1","JCRName":"BIOCHEMISTRY & MOLECULAR BIOLOGY","Score":null,"Total":0}
引用次数: 0
Abstract
The mammalian mitochondrial protein synthesis system produces 13 essential subunits of oxidative phosphorylation (OXPHOS) complexes. Translation initiation in mammalian mitochondria is characterized by the use of leaderless messenger RNAs (mRNAs) and non-AUG start codons, where the proofreading function of IF-3mt still remains elusive. Here, we developed a reconstituted mammalian mitochondrial translation system using in vitro transcribed and native mitochondrial transfer RNAs (tRNAs) to investigate IF-3mt’s proofreading function. Similar to bacterial IF-3, IF-3mt permits an initiator tRNA to participate in initiation by discriminating the three G–C pairs in its anticodon stem, and by the cognate interactions of its anticodon with the AUG start codon. As a result, IF-3mt promotes the accurate initiation of leaderless mRNAs. Nevertheless, IF-3mt can also facilitate initiation from the non-AUG(AUA) start codon through its unique N- and C-terminal extensions, in concert with the 5-methylcytidine (m5C) or 5-formylcytidine (f5C) modification at the anticodon wobble position of mt-tRNAMet. This is partly because the IF-3mt-specific N- and C-terminal extensions and the KKGK-motif favor leaderless mRNA initiation and relax non-AUG start codon discrimination. Analyses of IF-3mt-depleted human cells revealed that IF-3mt indeed participates in translating the open reading frames (ORFs) of leaderless mRNAs, as well as the internal ORFs of dicistronic mRNAs.
期刊介绍:
Nucleic Acids Research (NAR) is a scientific journal that publishes research on various aspects of nucleic acids and proteins involved in nucleic acid metabolism and interactions. It covers areas such as chemistry and synthetic biology, computational biology, gene regulation, chromatin and epigenetics, genome integrity, repair and replication, genomics, molecular biology, nucleic acid enzymes, RNA, and structural biology. The journal also includes a Survey and Summary section for brief reviews. Additionally, each year, the first issue is dedicated to biological databases, and an issue in July focuses on web-based software resources for the biological community. Nucleic Acids Research is indexed by several services including Abstracts on Hygiene and Communicable Diseases, Animal Breeding Abstracts, Agricultural Engineering Abstracts, Agbiotech News and Information, BIOSIS Previews, CAB Abstracts, and EMBASE.