PAP- and GLD-2-type poly(A) polymerases are required sequentially in cytoplasmic polyadenylation and oogenesis inDrosophila

Abstract

*Cytoplasmic polyadenylation has an essential role in activating maternal mRNA translation during early development. In vertebrates, the reaction requires CPEB, an RNA-binding protein and the poly(A) polymerase GLD-2. GLD-2-type poly(A) polymerases form a family clearly distinguishable from canonical poly(A) polymerases (PAPs). In Drosophila, canonical PAP is involved in cytoplasmic polyadenylation with Orb, the Drosophila CPEB, during mid-oogenesis. We show that the female germline GLD-2 is encoded by wispy. Wispy acts as a poly(A) polymerase in a tethering assay and in vivo for cytoplasmic polyadenylation of specific mRNA targets during late oogenesis and early embryogenesis. wispy function is required at the final stage of oogenesis for metaphase of meiosis I arrest and for progression beyond this stage. By contrast, canonical PAP acts with Orb for the earliest steps of oogenesis. Both Wispy and PAP interact with Orb genetically and physically in an ovarian complex. We conclude that two distinct poly(A) polymerases have a role in cytoplasmic polyadenylation in the female germline, each of them being specifically required for different steps of oogenesis.

Regulation of Cyclin A protein in meiosis and early embryogenesis

“…The poly(A) tail of cyclin A mRNA is short during prophase I arrest, and its lengthening correlates with appearance of the protein at oocyte maturation. It remains to be determined how this polyadenylation is controlled, but a likely possibility is that it may involve the GLD2 poly(A) polymerase shown to polyadenylate other mRNAs at maturation (2,33). The poly(A) tail is further increased at egg activation in a mechanism that requires active PNG kinase, and this may contribute to efficient translation of Cyclin A during early embryogenesis.…”

“…The regulation of the meiotic cell cycle at maturation is best understood in Xenopus, where control of translation by polyadenylation leads to the translation of both Cyclin B to produce MPF and the Mos kinase, which is crucial in maintaining the secondary meiotic arrest (1). In Drosophila oogenesis, some mRNAs become polyadenylated at maturation, dependent on the GLD2 cytoplasmic poly(A) polymerase (2). The cyclin B mRNA is polyadenylated initially at oocyte maturation, coinciding with appearance of the protein in meiosis (3,4).…”

“…In Drosophila oogenesis, poly(A) tail lengthening occurs both at oocyte maturation and activation (3). Lengthening of the poly(A) tail of cyclin B mRNA at each of these meiotic transitions is correlated with renewed appearance of the protein (3,4), and polyadenylation of the mRNA for the meiosis-specific APC/C activator Cortex by the GLD2 cytoplasmic poly(A) polymerase has been shown to be required for its translation at maturation (2). Given these precedents, we tested whether polyadenylation could be an additional mechanism besides loss of Bruno contributing to the increase of Cyclin A protein at maturation.…”

Translational regulation of the cell cycle: when, where, how and why?

“…Translation of Cyclin B also is promoted by Drosophila CPEB, called ORB ( figure 2c). In addition, a GLD-2 homologue is required at maturation for polyadenylation and translation of many proteins, including the Drosophila orthologue of Mos and a meiosis-specific activator of the APC/C called Cortex [42,101,102].…”

Translational activation maintains germline tissue homeostasis during adulthood

“…With the exception of a cytosolic form of canonical poly(A) polymerase in Drosophila which is important for early female germ cell stages, 47 little is known about the expression of cytoPAPs in other GSC lineages. However, as the known translational repressive networks utilizes deadenylases to silence gene expression in Drosophila GSCs, 48 it is quite conceivable that cytoplasmic poly (A) polymerases will emerge as conserved regulators of opposing cell fate decisions in GSCs.…”