CPEB3 and CPEB4 in neurons: analysis of RNA-binding specificity and translational control of AMPA receptor GluR2 mRNA.
CPEB is a sequence-specific RNA-binding protein that promotes polyadenylation-induced translation in oocytes and neurons. Vertebrates contain three additional genes that encode CPEB-like proteins, all of which are expressed in the brain. Here, we use SELEX, RNA structure probing, and RNA footprinting to show that CPEB and the CPEB-like proteins interact with different RNA sequences and thus constitute different classes of RNA-binding proteins. In transfected neurons, CPEB3 represses the translation of a reporter RNA in tethered function assays; in response to NMDA receptor activation, translation is stimulated. In contrast to CPEB, CPEB3-mediated translation is unlikely to involve cytoplasmic polyadenylation, as it requires neither the cis-acting AAUAAA nor the trans-acting cleavage and polyadenylation specificity factor, both of which are necessary for CPEB-induced polyadenylation. One target of CPEB3-mediated translation is GluR2 mRNA; not only does CPEB3 bind this RNA in vitro and in vivo, but an RNAi knockdown of CPEB3 in neurons results in elevated levels of GluR2 protein. These results indicate that CPEB3 is a sequence-specific translational regulatory protein.
Huang
Yi-Shuian
YS
Program in Molecular Medicine, University of Massachusetts Medical School, Worcester, MA 01605, USA.
Kan
Ming-Chung
MC
Lin
Chien-Ling
CL
Richter
Joel D
JD
D020413:3' Untranslated Regions / Q000235:genetics / Q000378:metabolism; D000818:Animals; D002460:Cell Line; D008297:Male; D051379:Mice; D009474:Neurons / Q000166:cytology / Q000378:metabolism*; D026723:Polyadenylation / Q000235:genetics; D011485:Protein Binding / Q000235:genetics; D014176:Protein Biosynthesis / Q000235:genetics; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism*; D016601:RNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D051381:Rats; D018091:Receptors, AMPA / Q000096:biosynthesis*; D014162:Transfection; D014981:Xenopus
D020413:3' Untranslated Regions; C477603:Cpeb3 protein, mouse; C477604:Cpeb4 protein, mouse; D012333:RNA, Messenger; D016601:RNA-Binding Proteins; D018091:Receptors, AMPA; C104722:glutamate receptor ionotropic, AMPA 2