Sequential dimerization of human zipcode-binding protein IMP1 on RNA: a cooperative mechanism providing RNP stability.
Active cytoplasmic RNA localization depends on the attachment of RNA-binding proteins that dictate the destination of the RNA molecule. In this study, we used an electrophoretic mobility-shift assay in combination with equilibrium and kinetic analyses to characterize the assembly of the human zipcode-binding protein IMP1 on targets in the 3'-UTR from Igf-II mRNA and in H19 RNA. In both cases, two molecules of IMP1 bound to RNA by a sequential, cooperative mechanism, characterized by an initial fast step, followed by a slow second step. The first step created an obligatory assembly intermediate of low stability, whereas the second step was the discriminatory event that converted a putative RNA target into a 'locked' stable RNP. The ability to dimerize was also observed between members of the IMP family of zipcode-binding proteins, providing a multitude of further interaction possibilities within RNP granules and with the localization apparatus.
Nielsen
Jacob
J
Institute of Molecular Biology, University of Copenhagen, Copenhagen, Denmark.
Kristensen
Mette Ahm
MA
Willemoës
Martin
M
Nielsen
Finn Cilius
FC
Christiansen
Jan
J
D020413:3' Untranslated Regions / Q000378:metabolism; D020816:Amino Acid Motifs; D001665:Binding Sites; D019281:Dimerization; D006801:Humans; D007335:Insulin-Like Growth Factor II / Q000235:genetics; D007700:Kinetics; D011485:Protein Binding; D012313:RNA / Q000378:metabolism*; D062085:RNA, Long Noncoding; D022661:RNA, Untranslated / Q000378:metabolism; D016601:RNA-Binding Proteins / Q000737:chemistry / Q000378:metabolism*; D012261:Ribonucleoproteins / Q000378:metabolism
D020413:3' Untranslated Regions; C094335:H19 long non-coding RNA; C406144:IGF2BP1 protein, human; D062085:RNA, Long Noncoding; D022661:RNA, Untranslated; D016601:RNA-Binding Proteins; D012261:Ribonucleoproteins; D012313:RNA; D007335:Insulin-Like Growth Factor II