title: A non-canonical initiation site is required for efficient translation of the dendritically localized Shank1 mRNA.
abstract: Local protein synthesis in dendrites enables neurons to selectively change the protein complement of individual postsynaptic sites. Though it is generally assumed that this mechanism requires tight translational control of dendritically transported mRNAs, it is unclear how translation of dendritic mRNAs is regulated. We have analyzed here translational control elements of the dendritically localized mRNA coding for the postsynaptic scaffold protein Shank1. In its 5' region, the human Shank1 mRNA exhibits two alternative translation initiation sites (AUG⁺¹ and AUG⁺²¹⁴), three canonical upstream open reading frames (uORFs1-3) and a high GC content. In reporter assays, fragments of the 5'UTR with high GC content inhibit translation, suggesting a contribution of secondary structures. uORF3 is most relevant to translation control as it overlaps with the first in frame start codon (AUG⁺¹), directing translation initiation to the second in frame start codon (AUG⁺²¹⁴). Surprisingly, our analysis points to an additional uORF initiated at a non-canonical ACG start codon. Mutation of this start site leads to an almost complete loss of translation initiation at AUG⁺¹, demonstrating that this unconventional uORF is required for Shank1 synthesis. Our data identify a novel mechanism whereby initiation at a non-canonical site allows for translation of the main Shank1 ORF despite a highly structured 5'UTR.
authors.affiliation: Institut für Humangenetik, Universitätsklinikum Hamburg-Eppendorf, Hamburg, Germany.
authors.forename: Katrin
authors.initials: K
authors.lastname: Studtmann
authors.affiliation: Institut für Humangenetik, Universitätsklinikum Hamburg-Eppendorf, Hamburg, Germany.
authors.forename: Janin
authors.initials: J
authors.lastname: Olschläger-Schütt
authors.affiliation: Institut für klinische Chemie; Universitätsklinikum Hamburg-Eppendorf, Hamburg, Germany.
authors.forename: Friedrich
authors.initials: F
authors.lastname: Buck
authors.affiliation: Zentrum für Molekulare Neurobiologie Hamburg, Universitätsklinikum Hamburg-Eppendorf, Hamburg, Germany.
authors.forename: Dietmar
authors.initials: D
authors.lastname: Richter
authors.affiliation: Consiglio Nazionale delle Ricerche, Institute of Neuroscience, Milano, Italy.
authors.forename: Carlo
authors.initials: C
authors.lastname: Sala
authors.affiliation: Institut für Anatomie; Universität Ulm, Ulm, Germany.
authors.forename: Jürgen
authors.initials: J
authors.lastname: Bockmann
authors.affiliation: Institut für Humangenetik, Universitätsklinikum Hamburg-Eppendorf, Hamburg, Germany.
authors.forename: Stefan
authors.initials: S
authors.lastname: Kindler
authors.affiliation: Institut für Humangenetik, Universitätsklinikum Hamburg-Eppendorf, Hamburg, Germany.
authors.forename: Hans-Jürgen
authors.initials: HJ
authors.lastname: Kreienkamp
mesh_terms: D020121:5' Untranslated Regions; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D018387:Codon, Initiator / Q000378:metabolism; D004252:DNA Mutational Analysis; D003712:Dendrites / Q000378:metabolism*; D017353:Gene Deletion; D006801:Humans; D051379:Mice; D008969:Molecular Sequence Data; D009154:Mutation; D009419:Nerve Tissue Proteins / Q000235:genetics / Q000378:metabolism*; D016366:Open Reading Frames; D014176:Protein Biosynthesis; D012333:RNA, Messenger / Q000378:metabolism; D024363:Transcription Initiation Site
chemical_list: D020121:5' Untranslated Regions; D018387:Codon, Initiator; D009419:Nerve Tissue Proteins; D012333:RNA, Messenger; C574178:SHANK1 protein, human; C512412:SHANK1 protein, mouse