Cup Blocks the Precocious Activation of the Orb Autoregulatory Loop

Li Chin Wong,Paul Schedl,Christos Samakovlis

doi:10.1371/journal.pone.0028261

Li Chin Wong, Paul Schedl + Show 1 more

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https://doi.org/10.1371/journal.pone.0028261

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Journal: PloS one	Publication Date: Dec 2, 2011
Citations: 34	License type: CC BY 4.0

Affiliation: Princeton University

Abstract

Translational regulation of localized mRNAs is essential for patterning and axes determination in many organisms. In the Drosophila ovary, the germline-specific Orb protein mediates the translational activation of a variety of mRNAs localized in the oocyte. One of the Orb target mRNAs is orb itself, and this autoregulatory activity ensures that Orb proteins specifically accumulate in the developing oocyte. Orb is an RNA-binding protein and is a member of the cytoplasmic polyadenylation element binding (CPEB) protein family. We report here that Cup forms a complex in vivo with Orb. We also show that cup negatively regulates orb and is required to block the precocious activation of the orb positive autoregulatory loop. In cup mutant ovaries, high levels of Orb accumulate in the nurse cells, leading to what appears to be a failure in oocyte specification as a number of oocyte markers inappropriately accumulate in nurse cells. In addition, while orb mRNA is mislocalized and destabilized, a longer poly(A) tail is maintained than in wild type ovaries. Analysis of Orb phosphoisoforms reveals that loss of cup leads to the accumulation of hyperphosphorylated Orb, suggesting that an important function of cup in orb-dependent mRNA localization pathways is to impede Orb activation.

Highlights

In eukaryotic cells, cytoplasmic polyadenylation is used to activate the on-site translation of localized mRNAs
The first is the AAUAAA motif, which is bound by the cleavage and polyadenylation specificity factor (CPSF) while the second is the U-rich cytoplasmic polyadenylation element (CPE) which is bound by the cytoplasmic polyadenylation element binding protein (CPEB) [1], [2]
Orb has two key activities in this autoregulatory loop. It functions in the localization of orb mRNA within the oocyte by anchoring the message to the oocyte cortex

Summary

Introduction

Cytoplasmic polyadenylation is used to activate the on-site translation of localized mRNAs. The first is the AAUAAA motif, which is bound by the cleavage and polyadenylation specificity factor (CPSF) while the second is the U-rich cytoplasmic polyadenylation element (CPE) which is bound by the cytoplasmic polyadenylation element binding protein (CPEB) [1], [2]. The evolutionarily conserved CPEBs are RNA-recognition motif (RRM)-type RNA-binding proteins and they have been found in species ranging from nematodes to humans. One of the founding members of the CPEB family is the Drosophila germline-specific protein Orb [3]. Orb plays a critical role in the development of the female germline and is required for mRNA localization and translational regulation throughout much of oogenesis [4], [5]

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