The broad objective of this proposal is to understand posttranscriptional control of early animal development. One mechanism that regulates maternal mRNA expression is cytoplasmic polyadenylation. CPEB, a sequence-specific RNA binding protein, is at the heart of this process;it nucleates a number of factors on specific mRNAs and controls translation by modulating poly(A) tail length. Polyadenylation does not happen en masse, but instead occurs in sequential waves that are most evident during oocyte maturation in Xenopus. The first wave happens at meiosis I (MI) while the second occurs at meiosis II (MII). One determinant for first or second wave polyadenylation is the amount of CPEB in the cell. During the MI to MII transition, some of the CPEB is destroyed, which is important for second wave polyadenylation. CPEB destruction requires multiple cdk1 phosphorylations, subsequent ubiquitination, and Pin1 (peptidylprolyl cis/trans isomerase) activity. Pin1 activity increases during meiosis and binds and controls CPEB destruction. Moreover, Pin1 mediates CPEB destruction in mammalian cells as well. Pin1 also binds phospho-maskin, the CPEB- and eIF4E- binding factor that coincides with maskin phosphorylation and dissociation from eIF4E, allowing initiation complex assembly. The goals of the first specific aim are to determine how Pin1 mediates CPEB destruction and MI to MII transitioning, and whether it regulates translation by disrupting the maskin-eIF4E interaction. The second specific aim intends construct an integrated network map of polyadenylation during maturation. To do so, CLIP (crosslink IP) will be employed;it will identify not only the mRNAs to which CPEB binds, but where on the mRNA CPEB binds. This analysis will define, on a genome wide scale, what constitutes a CPEB binding element (CPE). These experiments will be complemented by the identification of all the mRNAs that undergo cytoplasmic polyadenylation, and which of 2 non-canonical poly(A) polymerases are used. Translational control by CPEB regulates not only early development, but neuronal synaptic cellular senescence as well. Thus, these experiments have important implications of human infertility, neurodegeneration, and cancer etiology.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM046779-24
Application #
8637076
Study Section
Molecular Genetics A Study Section (MGA)
Program Officer
Bender, Michael T
Project Start
1992-02-01
Project End
2016-03-31
Budget Start
2014-04-01
Budget End
2015-03-31
Support Year
24
Fiscal Year
2014
Total Cost
$386,575
Indirect Cost
$151,575
Name
University of Massachusetts Medical School Worcester
Department
Other Basic Sciences
Type
Schools of Medicine
DUNS #
603847393
City
Worcester
State
MA
Country
United States
Zip Code
01655
Ivshina, Maria; Lasko, Paul; Richter, Joel D (2014) Cytoplasmic polyadenylation element binding proteins in development, health, and disease. Annu Rev Cell Dev Biol 30:393-415
Nechama, Morris; Lin, Chien-Ling; Richter, Joel D (2013) An unusual two-step control of CPEB destruction by Pin1. Mol Cell Biol 33:48-58
Udagawa, Tsuyoshi; Farny, Natalie G; Jakovcevski, Mira et al. (2013) Genetic and acute CPEB1 depletion ameliorate fragile X pathophysiology. Nat Med 19:1473-7
Lin, Chien-Ling; Huang, Yen-Tsung; Richter, Joel D (2012) Transient CPEB dimerization and translational control. RNA 18:1050-61
Darnell, Jennifer C; Van Driesche, Sarah J; Zhang, Chaolin et al. (2011) FMRP stalls ribosomal translocation on mRNAs linked to synaptic function and autism. Cell 146:247-61
Cao, Quiping; Padmanabhan, Kiran; Richter, Joel D (2010) Pumilio 2 controls translation by competing with eIF4E for 7-methyl guanosine cap recognition. RNA 16:221-7
Lin, Chien-Ling; Evans, Veronica; Shen, Shihao et al. (2010) The nuclear experience of CPEB: implications for RNA processing and translational control. RNA 16:338-48
Kan, Ming-Chung; Oruganty-Das, Aparna; Cooper-Morgan, Amalene et al. (2010) CPEB4 is a cell survival protein retained in the nucleus upon ischemia or endoplasmic reticulum calcium depletion. Mol Cell Biol 30:5658-71
Kim, Jong Heon; Richter, Joel D (2008) Measuring CPEB-mediated cytoplasmic polyadenylation-deadenylation in Xenopus laevis oocytes and egg extracts. Methods Enzymol 448:119-38
Kim, Jong Heon; Richter, Joel D (2007) RINGO/cdk1 and CPEB mediate poly(A) tail stabilization and translational regulation by ePAB. Genes Dev 21:2571-9

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