CPEB degradation during Xenopus oocyte maturation requires a PEST domain and the 26S proteasome.
Cytoplasmic poly(A) elongation is widely utilized during the early development of many organisms as a mechanism for translational activation. Targeting of mRNAs for this mechanism requires the presence of a U-rich element, the cytoplasmic polyadenylation element (CPE), and its binding protein, CPEB. Blocking cytoplasmic polyadenylation by interfering with the CPE or CPEB prevents the translational activation of mRNAs that are crucial for oocyte maturation. The CPE sequence and CPEB are also important for translational repression of mRNAs stored in the Xenopus oocyte during oogenesis. To understand the contribution of protein metabolism to these two roles for CPEB, we have examined the mechanisms influencing the expression of CPEB during oogenesis and oocyte maturation. Through a comparison of CPEB mRNA levels, protein synthesis, and accumulation, we find that CPEB is synthesized during oogenesis and stockpiled in the oocyte. Minimal synthesis of CPEB, <3.6%, occurs during oocyte maturation. In late oocyte maturation, 75% of CPEB is degraded coincident with germinal vesicle breakdown. Using proteasome and ubiquitination inhibitors, we demonstrate that CPEB degradation occurs via the proteasome pathway, most likely through ubiquitin-conjugated intermediates. In addition, we demonstrate that degradation requires a 14 amino acid PEST domain.
Biology Department, Boston College, Chestnut Hill, Massachusetts 02467, USA.
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D000595:Amino Acid Sequence; D000818:Animals; D015152:Blotting, Northern; D003546:Cysteine Endopeptidases; D003593:Cytoplasm / Q000378:metabolism; D015151:Immunoblotting; D008957:Models, Genetic; D008969:Molecular Sequence Data; D009097:Multienzyme Complexes / Q000037:antagonists & inhibitors; D009865:Oocytes / Q000378:metabolism*; D010447:Peptide Hydrolases / Q000502:physiology*; D010957:Plasmids / Q000378:metabolism; D011374:Progesterone / Q000494:pharmacology; D046988:Proteasome Endopeptidase Complex; D017434:Protein Structure, Tertiary; D012313:RNA / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D016601:RNA-Binding Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000378:metabolism*; D017386:Sequence Homology, Amino Acid; D013997:Time Factors; D014157:Transcription Factors / Q000096:biosynthesis / Q000737:chemistry / Q000378:metabolism*; D014158:Transcription, Genetic; D014452:Ubiquitins / Q000031:analogs & derivatives* / Q000037:antagonists & inhibitors / Q000378:metabolism; D014981:Xenopus; D029867:Xenopus Proteins; D039221:mRNA Cleavage and Polyadenylation Factors
C490388:Cpeb1 protein, Xenopus; D009097:Multienzyme Complexes; D012333:RNA, Messenger; D016601:RNA-Binding Proteins; D014157:Transcription Factors; D014452:Ubiquitins; D029867:Xenopus Proteins; D039221:mRNA Cleavage and Polyadenylation Factors; C051927:ubiquitin-aldehyde; D011374:Progesterone; D012313:RNA; D010447:Peptide Hydrolases; D003546:Cysteine Endopeptidases; D046988:Proteasome Endopeptidase Complex; C061553:ATP dependent 26S protease