CPEB interacts with an ovary-specific eIF4E and 4E-T in early Xenopus oocytes.
CPEB (cytoplasmic polyadenylation element-binding protein) is an important regulator of translation in oocytes and neurons. Although previous studies of CPEB in late Xenopus oocytes involve the eIF4E-binding protein maskin as the key factor for the repression of maternal mRNA, a second mechanism must exist, since maskin is absent earlier in oogenesis. Using co-immunoprecipitation and gel filtration assays, we show that CPEB specifically interacts, via protein/protein interactions, with the RNA helicase Xp54, the RNA-binding proteins P100(Pat1) and RAP55, the eIF4E-binding protein 4E-T, and an eIF4E protein. Remarkably, these CPEB complex proteins have been characterized, in one or more organism, as P-body, maternal, or neuronal granule components. We do not detect interactions with eIF4E1a, the canonical cap-binding factor, eIF4G, or eIF4A or with proteins expressed late in oogenesis, including maskin, PARN, and 4E-BP1. The eIF4E protein was identified as eIF4E1b, a close homolog of eIF4E1a, whose expression is restricted to oocytes and early embryos. Although eIF4E1b possesses all residues required for cap and eIF4G binding, it binds m(7)GTP weakly, and in pull-down assays, rather than binding eIF4G, it binds 4E-T, in a manner independent of the consensus eIF4E-binding site, YSKEELL. Wild type and Y-A mutant 4E-T (which binds eIF4E1b but not eIF4E1a), when tethered to a reporter mRNA, represses its translation in a cap-dependent manner, and injection of eIF4E1b antibody accelerates meiotic maturation. Altogether, our data suggest that CPEB, partnered with several highly conserved RNA-binding partners, inhibits protein synthesis in oocytes using a novel pairing of 4E-T and eIF4E1b.
Minshall
Nicola
N
Department of Biochemistry, University of Cambridge, Cambridge, UK.
Reiter
Marie Helene
MH
Weil
Dominique
D
Standart
Nancy
N
D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D002352:Carrier Proteins / Q000737:chemistry / Q000378:metabolism*; D039561:Eukaryotic Initiation Factor-4E / Q000737:chemistry / Q000378:metabolism*; D005260:Female; D018507:Gene Expression Regulation, Developmental; D008954:Models, Biological; D008969:Molecular Sequence Data; D009474:Neurons / Q000378:metabolism; D009865:Oocytes / Q000378:metabolism*; D010053:Ovary / Q000378:metabolism*; D012333:RNA, Messenger / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D014157:Transcription Factors / Q000737:chemistry / Q000378:metabolism*; D029867:Xenopus Proteins / Q000737:chemistry / Q000378:metabolism*; D014982:Xenopus laevis / Q000378:metabolism*; D039221:mRNA Cleavage and Polyadenylation Factors / Q000737:chemistry / Q000378:metabolism*
D002352:Carrier Proteins; C490388:Cpeb1 protein, Xenopus; D039561:Eukaryotic Initiation Factor-4E; D012333:RNA, Messenger; D014157:Transcription Factors; D029867:Xenopus Proteins; D039221:mRNA Cleavage and Polyadenylation Factors