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Dissecting the role of the conserved CPEB/Xp54 complex in translational repression in early development

Reference	BB/C514766/1
Principal Investigator / Supervisor	Dr Nancy Standart
Co-Investigators / Co-Supervisors	Dr Nicola Minshall
Institution	University of Cambridge
Department	Biochemistry
Funding type	Research
Value (£)	259,507
Status	Completed
Type	Research Grant
Start date	01/03/2005
End date	31/08/2008
Duration	42 months

Abstract

In early development, during the period from the meiotic maturation of oocytes to eggs through to onset of zygotic transcription in the fertilised embryo, the principal means of gene regulation is translational control of the masked maternal mRNA. The cytoplasmic polyadenylation element binding protein (CPEB) is a critical regulator of translation in early development, in organisms ranging from clam and flies to frog and man. CPEB binds CPE elements in the 3 prime untranslated regions of specific mRNAs, and controls their expression at the level of translational repression in oocytes, and by cytoplasmic polyadenylation and translational activation in eggs. We have shown that CPEB interacts with a DEAD-box RNA helicase p54 in Xenopus laevis and in clam oocytes. In tethered function assays in Xenopus oocytes, Xp54 represses translation. Xp54 functions as a dimmer, and co-purified with CPEB and elF4E, the cap-binding translation inflation factor. Xp54 belongs to a highly conserved helicase family, members of which are components of large cytoplasmic granules and function in translational repression and RNA decapping in other model systems; but whose detailed role in the control of gene expression is now known. In preliminary experiments, we identified two further members of the CPEB/Xp54 complex as X4E-T and P100. Human 4E-T is a large elF4E-binding protein that transports a minor fraction of elF4E into nuclei, and, when overexpressed, inhibits translation of reporter mRNA. The Drosophila homologue, Cup, represses the translation of oskar and nanos mRNAs by interacting with their specific 3 prime UTR binding proteins, Bruno and Smaug respectively, and also with elF4E, preventing elF4E interacting with elF4G. P100 is a cytoplasmic protein in oocytes, though little is known of its role in these cells. In yeast, Pat1 (P100) interacts with Dhh1 (Xp54); both function as enhancers of decapping, which requires elF4E removal. Thus, homologues of the two Xp54 partners have documented roles in elf4E-binding and translational repression, or in decapping but neither have been investigated in early vertebrate development. Our hypothesis is that in oocytes the Xp54 complex, whose components include CPEB, 4E-T and P100, controls ElF4E-elF4G interactions leading to translational repression. We propose that 4E-T, the major focus of our proposed study, is an evolutionarily conserved adaptor protein that is used by multiple translational repressors including Bruno, Smaug, and CPEB/Xp54; making it THE critical mediator of translational repression in early development. We propose to determine the role of Xp54 in translational repression of different mRNA classes in Xenopus oocytes, to characterise X4E-T and P100 in oocytes and eggs, to undertake mapping of the X4E-T/P100 interactions with Xp54 and CPEB, in order to perform functional tests of X4E-T and P100 in translational repression and to examine the importance of CPEB/Xp54-4E-T/P100 interactions for function. The data from this study should allow us to develop an experimentally based model of gene expression control by CPEB/Xp54 involving X4E-T and P100.

Summary

unavailable

Committee	Closed Committee - Biochemistry & Cell Biology (BCB)
Research Topics	X – not assigned to a current Research Topic
Research Priority	X – Research Priority information not available
Research Initiative	X - not in an Initiative
Funding Scheme	X – not Funded via a specific Funding Scheme

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