PMID- 32589964 OWN - NLM STAT- MEDLINE DCOM- 20200924 LR - 20240207 IS - 1097-4164 (Electronic) IS - 1097-2765 (Print) IS - 1097-2765 (Linking) VI - 79 IP - 4 DP - 2020 Aug 20 TI - Selective Translation Complex Profiling Reveals Staged Initiation and Co-translational Assembly of Initiation Factor Complexes. PG - 546-560.e7 LID - S1097-2765(20)30389-0 [pii] LID - 10.1016/j.molcel.2020.06.004 [doi] AB - Translational control targeting the initiation phase is central to the regulation of gene expression. Understanding all of its aspects requires substantial technological advancements. Here we modified yeast translation complex profile sequencing (TCP-seq), related to ribosome profiling, and adapted it for mammalian cells. Human TCP-seq, capable of capturing footprints of 40S subunits (40Ss) in addition to 80S ribosomes (80Ss), revealed that mammalian and yeast 40Ss distribute similarly across 5'TRs, indicating considerable evolutionary conservation. We further developed yeast and human selective TCP-seq (Sel-TCP-seq), enabling selection of 40Ss and 80Ss associated with immuno-targeted factors. Sel-TCP-seq demonstrated that eIF2 and eIF3 travel along 5' UTRs with scanning 40Ss to successively dissociate upon AUG recognition; notably, a proportion of eIF3 lingers on during the initial elongation cycles. Highlighting Sel-TCP-seq versatility, we also identified four initiating 48S conformational intermediates, provided novel insights into ATF4 and GCN4 mRNA translational control, and demonstrated co-translational assembly of initiation factor complexes. CI - Copyright (c) 2020 The Author(s). Published by Elsevier Inc. All rights reserved. FAU - Wagner, Susan AU - Wagner S AD - EMBL-Australia Collaborating Group, Department of Genome Sciences, John Curtin School of Medical Research, Australian National University, Canberra, ACT 2601, Australia; Laboratory of Regulation of Gene Expression, Institute of Microbiology of the Czech Academy of Sciences, Videnska 1083, 142 20, Prague, Czech Republic. Electronic address: susan.wagner@anu.edu.au. FAU - Herrmannova, Anna AU - Herrmannova A AD - Laboratory of Regulation of Gene Expression, Institute of Microbiology of the Czech Academy of Sciences, Videnska 1083, 142 20, Prague, Czech Republic. FAU - Hronova, Vladislava AU - Hronova V AD - Laboratory of Regulation of Gene Expression, Institute of Microbiology of the Czech Academy of Sciences, Videnska 1083, 142 20, Prague, Czech Republic. FAU - Gunisova, Stanislava AU - Gunisova S AD - Laboratory of Regulation of Gene Expression, Institute of Microbiology of the Czech Academy of Sciences, Videnska 1083, 142 20, Prague, Czech Republic. FAU - Sen, Neelam D AU - Sen ND AD - Laboratory of Gene Regulation and Development, Eunice Kennedy Shriver National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, MD 20892, USA. FAU - Hannan, Ross D AU - Hannan RD AD - Australian Cancer Research Foundation Department of Cancer Biology and Therapeutics, John Curtin School of Medical Research, Australian National University, Canberra, ACT 2601, Australia. FAU - Hinnebusch, Alan G AU - Hinnebusch AG AD - Laboratory of Gene Regulation and Development, Eunice Kennedy Shriver National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, MD 20892, USA. FAU - Shirokikh, Nikolay E AU - Shirokikh NE AD - EMBL-Australia Collaborating Group, Department of Genome Sciences, John Curtin School of Medical Research, Australian National University, Canberra, ACT 2601, Australia. FAU - Preiss, Thomas AU - Preiss T AD - EMBL-Australia Collaborating Group, Department of Genome Sciences, John Curtin School of Medical Research, Australian National University, Canberra, ACT 2601, Australia; Victor Chang Cardiac Research Institute, Darlinghurst, NSW 2010, Australia. Electronic address: thomas.preiss@anu.edu.au. FAU - Valasek, Leos Shivaya AU - Valasek LS AD - Laboratory of Regulation of Gene Expression, Institute of Microbiology of the Czech Academy of Sciences, Videnska 1083, 142 20, Prague, Czech Republic. Electronic address: valasekl@biomed.cas.cz. LA - eng GR - WT_/Wellcome Trust/United Kingdom GR - 090812/B/09/Z/WT_/Wellcome Trust/United Kingdom PT - Journal Article PT - Research Support, Non-U.S. Gov't DEP - 20200625 PL - United States TA - Mol Cell JT - Molecular cell JID - 9802571 RN - 0 (5' Untranslated Regions) RN - 0 (ATF4 protein, human) RN - 0 (Basic-Leucine Zipper Transcription Factors) RN - 0 (Codon, Initiator) RN - 0 (Eukaryotic Initiation Factor-2) RN - 0 (Eukaryotic Initiation Factor-3) RN - 0 (GCN4 protein, S cerevisiae) RN - 0 (Multiprotein Complexes) RN - 0 (Peptide Initiation Factors) RN - 0 (Saccharomyces cerevisiae Proteins) RN - 145891-90-3 (Activating Transcription Factor 4) SB - IM CIN - Mol Cell. 2020 Aug 20;79(4):539-541. PMID: 32822578 MH - 5' Untranslated Regions MH - Activating Transcription Factor 4/genetics/metabolism MH - Basic-Leucine Zipper Transcription Factors/genetics/metabolism MH - Codon, Initiator MH - Eukaryotic Initiation Factor-2/genetics/metabolism MH - Eukaryotic Initiation Factor-3/genetics/metabolism MH - HEK293 Cells MH - Humans MH - Multiprotein Complexes/genetics/*metabolism MH - Peptide Initiation Factors/genetics/*metabolism MH - *Protein Biosynthesis MH - Ribosome Subunits, Small, Eukaryotic/genetics/metabolism MH - Ribosomes/genetics/*metabolism MH - Saccharomyces cerevisiae/genetics MH - Saccharomyces cerevisiae Proteins/genetics/metabolism PMC - PMC7447980 OTO - NOTNLM OT - ATF4 OT - GCN4 OT - Ribo-seq OT - TCP-seq OT - UTR OT - co-translational assembly OT - eIF2 OT - eIF3 OT - gene expression OT - mRNA OT - ribosome OT - ribosome profiling OT - translational control COIS- Declaration of Interests The authors declare no competing interests. EDAT- 2020/06/27 06:00 MHDA- 2020/09/25 06:00 PMCR- 2020/08/20 CRDT- 2020/06/27 06:00 PHST- 2019/11/06 00:00 [received] PHST- 2020/04/10 00:00 [revised] PHST- 2020/05/18 00:00 [accepted] PHST- 2020/06/27 06:00 [pubmed] PHST- 2020/09/25 06:00 [medline] PHST- 2020/06/27 06:00 [entrez] PHST- 2020/08/20 00:00 [pmc-release] AID - S1097-2765(20)30389-0 [pii] AID - 10.1016/j.molcel.2020.06.004 [doi] PST - ppublish SO - Mol Cell. 2020 Aug 20;79(4):546-560.e7. doi: 10.1016/j.molcel.2020.06.004. Epub 2020 Jun 25.