title: Targeted translational regulation using the PUF protein family scaffold.
abstract: Regulatory complexes formed on mRNAs control translation, stability, and localization. These complexes possess two activities: one that binds RNA and another--the effector--that elicits a biological function. The Pumilio and FBF (PUF) protein family of RNA binding proteins provides a versatile scaffold to design and select proteins with new specificities. Here, the PUF scaffold is used to target translational activation and repression of specific mRNAs, and to induce specific poly(A) addition and removal. To do so, we linked PUF scaffold proteins to a translational activator, GLD2, or a translational repressor, CAF1. The chimeric proteins activate or repress the targeted mRNAs in Xenopus oocytes, and elicit poly(A) addition or removal. The magnitude of translational control relates directly to the affinity of the RNA-protein complex over a 100-fold range of K(d). The chimeric proteins act on both reporter and endogenous mRNAs: an mRNA that normally is deadenylated during oocyte maturation instead receives poly(A) in the presence of an appropriate chimera. The PUF-effector strategy enables the design of proteins that affect translation and stability of specific mRNAs in vivo.
authors.affiliation: Cellular and Molecular Biology Program, University of Wisconsin, Madison, WI 53706, USA.
authors.forename: Amy
authors.initials: A
authors.lastname: Cooke
authors.forename: Andrew
authors.initials: A
authors.lastname: Prigge
authors.forename: Laura
authors.initials: L
authors.lastname: Opperman
authors.forename: Marvin
authors.initials: M
authors.lastname: Wickens
mesh_terms: D000818:Animals; D015153:Blotting, Western; D029742:Caenorhabditis elegans Proteins / Q000235:genetics / Q000378:metabolism*; D005260:Female; D015870:Gene Expression; D006801:Humans; D008156:Luciferases / Q000235:genetics / Q000378:metabolism; D009154:Mutation; D009865:Oocytes / Q000378:metabolism; D011061:Poly A / Q000235:genetics / Q000378:metabolism; D011062:Polynucleotide Adenylyltransferase / Q000235:genetics / Q000378:metabolism*; D011485:Protein Binding; D014176:Protein Biosynthesis; D012313:RNA / Q000235:genetics / Q000378:metabolism; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D016601:RNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D014982:Xenopus laevis
chemical_list: D029742:Caenorhabditis elegans Proteins; C510538:PUF-8 protein, C elegans; C471242:PUM1 protein, human; D012333:RNA, Messenger; D016601:RNA-Binding Proteins; D011993:Recombinant Fusion Proteins; C505139:ccf-1 protein, C elegans; C109286:fem-3-binding protein, C elegans; D011061:Poly A; D012313:RNA; D008156:Luciferases; C491056:GLD-2 protein, C elegans; D011062:Polynucleotide Adenylyltransferase