GLD-2 is a cytoplasmic poly(A) polymerase present in the Caenorhabditis elegans germ line and embryo. It is a divergent member of the DNA polymerase  nucleotidyl transferase superfamily, which includes CCA-adding enzymes, DNA polymerases and eukaryotic nuclear poly(A) polymerases. The polyadenylation activity of GLD-2 is stimulated by physical interaction with an RNA binding protein, GLD-3. To test whether GLD-3 might stimulate GLD-2 by recruiting it to RNA, we tethered C. elegans GLD-2 to mRNAs in Xenopus oocytes by using MS2 coat protein. Tethered GLD-2 adds poly(A) and stimulates translation of the mRNA, demonstrating that recruitment is sufficient to stimulate polyadenylation activity. We use the same tethered assay to identify human and mouse poly(A) polymerases related to GLD-2. This may provide entrees to previously uncharacterized modes of polyadenylation in mammalian cells.
Cytoplasmic polyadenylation is important in the control of mRNA stability and translation, and for early animal development and synaptic plasticity. Here, we focus on vertebrate poly(A) polymerases that are members of the recently described GLD2 family. We identify and characterize two closely related GLD2 proteins in Xenopus oocytes, and show that they possess PAP activity in vivo and in vitro and that they bind known polyadenylation factors and mRNAs known to receive poly(A) during development. We propose that at least two distinct polyadenylation complexes exist in Xenopus oocytes, one of which contains GLD2; the other, maskin and Pumilio. GLD2 protein interacts with the polyadenylation factor, CPEB, in a conserved manner. mRNAs that encode GLD2 in mammals are expressed in many tissues. In the brain, mouse, and human GLD2 mRNAs are abundant in anatomical regions necessary for long-term cognitive and emotional learning. In the hippocampus, mouse GLD2 mRNA colocalizes with CPEB1 and Pumilio1 mRNAs, both of which are likely involved in synaptic plasticity. We suggest that mammalian GLD2 poly(A) polymerases are important in synaptic translation, and in polyadenylation throughout the soma.
SUMMARY Cytoplasmic polyadenylation is a key mechanism of gene control. In C. elegans, GLD-2 and GLD-3 provide the catalytic and RNA-binding subunits, respectively, of a major cytoplasmic poly(A) polymerase (PAP). Here we identify RNP-8 as a second GLD-2 partner. RNP-8 binds GLD-2 and stimulates GLD-2 activity to form a functional PAP, much like GLD-3. Moreover, GLD-2/RNP-8 and GLD-2/GLD-3 exist as separate complexes that form selectively during development, and RNP-8 and GLD-3 appear to have distinct RNA-binding specificities. Therefore, GLD-2 can form either of two discrete PAPs. In C.elegans hermaphrodites, gamete production begins with spermatogenesis and transitions later to oogenesis. We suggest that the combinatorial use of GLD-2 contributes to this transition, as GLD-2/GLD-3 promotes spermatogenesis, while GLD-2/RNP-8 specifies oogenesis. Indeed, RNP-8 and GLD-3 antagonize each other, as evidenced by genetic co-suppression and molecular competition for GLD-2 binding. We conclude that GLD-2 and its binding partners control gamete identity.
During the development of multicellular organisms, the processes of growth and differentiation are kept in balance to generate and maintain tissues and organs of the correct size, shape and cellular composition. We have investigated the molecular controls of growth and differentiation in the Caenorhabditis elegans germline. A single somatic cell, called the distal tip cell, promotes mitotic proliferation in the adjacent germline by GLP-1/Notch signalling. Within the germline, the decisions between mitosis and meiosis and between spermatogenesis and oogenesis are controlled by a group of conserved RNA regulators. FBF, a member of the PUF (for Pumilio and FBF) family of RNA-binding proteins, promotes mitosis by repressing gld-1 mRNA activity; the GLD-1, GLD-2, GLD-3 and NOS-3 proteins promote entry into meiosis by regulating mRNAs that remain unknown. The regulatory balance between opposing FBF and GLD activities is crucial for controlling the extent of germline proliferation. PUF proteins regulate germline stem cells in both Drosophila and C. elegans and are localized to germline stem cells of the mammalian testis. Therefore, this post-transcriptional regulatory switch may be an ancient mechanism for controlling maintenance of stem cells versus differentiation.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
hi@scite.ai
10624 S. Eastern Ave., Ste. A-614
Henderson, NV 89052, USA
Copyright © 2024 scite LLC. All rights reserved.
Made with 💙 for researchers
Part of the Research Solutions Family.