title: Quantitative analysis of mammalian translation initiation sites by FACS-seq.
abstract: An approach combining fluorescence-activated cell sorting and high-throughput DNA sequencing (FACS-seq) was employed to determine the efficiency of start codon recognition for all possible translation initiation sites (TIS) utilizing AUG start codons. Using FACS-seq, we measured translation from a genetic reporter library representing all 65,536 possible TIS sequences spanning the -6 to +5 positions. We found that the motif RYMRMVAUGGC enhanced start codon recognition and translation efficiency. However, dinucleotide interactions, which cannot be conveyed by a single motif, were also important for modeling TIS efficiency. Our dataset combined with modeling allowed us to predict genome-wide translation initiation efficiency for all mRNA transcripts. Additionally, we screened somatic TIS mutations associated with tumorigenesis to identify candidate driver mutations consistent with known tumor expression patterns. Finally, we implemented a quantitative leaky scanning model to predict alternative initiation sites that produce truncated protein isoforms and compared predictions with ribosome footprint profiling data. The comprehensive analysis of the TIS sequence space enables quantitative predictions of translation initiation based on genome sequence.
authors.affiliation: Department of Chemical Engineering, Stanford University, Stanford, CA, USA.
authors.forename: William L
authors.initials: WL
authors.lastname: Noderer
authors.affiliation: The Program in Epithelial Biology, Stanford University School of Medicine, Stanford, CA, USA.
authors.forename: Ross J
authors.initials: RJ
authors.lastname: Flockhart
authors.affiliation: The Program in Epithelial Biology, Stanford University School of Medicine, Stanford, CA, USA The Program in Cancer Biology, Stanford University School of Medicine, Stanford, CA, USA.
authors.forename: Aparna
authors.initials: A
authors.lastname: Bhaduri
authors.affiliation: Department of Chemical Engineering, Stanford University, Stanford, CA, USA.
authors.forename: Alexander J
authors.initials: AJ
authors.lastname: Diaz de Arce
authors.affiliation: The Program in Epithelial Biology, Stanford University School of Medicine, Stanford, CA, USA.
authors.forename: Jiajing
authors.initials: J
authors.lastname: Zhang
authors.affiliation: The Program in Epithelial Biology, Stanford University School of Medicine, Stanford, CA, USA Veterans Affairs Palo Alto Healthcare System, Palo Alto, CA, USA.
authors.forename: Paul A
authors.initials: PA
authors.lastname: Khavari
authors.affiliation: Department of Chemical Engineering, Stanford University, Stanford, CA, USA cliff.wang@stanford.edu.
authors.forename: Clifford L
authors.initials: CL
authors.lastname: Wang
mesh_terms: D020816:Amino Acid Motifs; D000818:Animals; D063646:Carcinogenesis; D045744:Cell Line, Tumor; D018387:Codon, Initiator; D005434:Flow Cytometry; D056726:Genetic Association Studies; D059014:High-Throughput Nucleotide Sequencing / Q000379:methods*; D006801:Humans; D051379:Mice; D008962:Models, Theoretical; D009154:Mutation; D041681:NIH 3T3 Cells; D010442:Peptide Chain Initiation, Translational; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism
chemical_list: D018387:Codon, Initiator; D012333:RNA, Messenger
keywords: FACS‐seq; Kozak motif; proteome modeling; start codon; translation initiation