Two conserved regulatory cytoplasmic poly(A) polymerases, GLD-4 and GLD-2, regulate meiotic progression in C. elegans

Abstract

Translational regulation is heavily employed during developmental processes to control the timely accumulation of proteins independently of gene transcription. In particular, mRNA poly(A) tail metabolism in the cytoplasm is a key determinant for balancing an mRNA's translational output and its decay rate. Noncanonical poly(A) polymerases (PAPs), such as germline development defective-2 (GLD-2), can mediate poly(A) tail extension. Little is known about the regulation and functional complexity of cytoplasmic PAPs. Here we report the discovery of Caenorhabditis elegans GLD-4, a cytoplasmic PAP present in P granules that is orthologous to Trf4/5p from budding yeast. GLD-4 enzymatic activity is enhanced by its interaction with GLS-1, a protein associated with the RNA-binding protein GLD-3. GLD-4 is predominantly expressed in germ cells, and its activity is essential for early meiotic progression of male and female gametes in the absence of GLD-2. For commitment into female meiosis, both PAPs converge on at least one common target mRNA—i.e., gld-1 mRNA—and, as a consequence, counteract the repressive action of two PUF proteins and the putative deadenylase CCR-4. Together our findings suggest that two different cytoplasmic PAPs stabilize and translationally activate several meiotic mRNAs to provide a strong fail-safe mechanism for early meiotic progression.

mir-35 is involved in intestine cell G1/S transition and germ cell proliferation in C. elegans

“…In the gonad, GLP-1 (abnormal Germ Line Proliferation)/Notch signaling is essential to promote germ line divisions in distal mitotic region [19][20][21][22]. GLD (defective in Germ Line Development) family plays a key role in determining meiosis entry and indirectly regulates mitotic proliferation of the germline [23][24][25][26][27]. We found that mir-35 may down-regulate gld-1 to ensure a normal germline proliferation in distal the mir-35 mutant gk262, less intestinal nuclei were observed than the wide type, at 3-fold and L3/L4 stages, respectively (Table 1, P < 0.001, Student t-test).…”

Translational activation maintains germline tissue homeostasis during adulthood

“…18 However, a role in adult tissue homeostasis, regulating the mitosis-to-meiosis decision had remained uncharacterized. By following aging hermaphrodites, we found that GLD-4 maintains the normal size of the proliferative zone and prevents its progressive shrinkage.…”

“…GLD-4 shows poly(A) polymerase activity in a heterologous in vivo tethering system, but its endogenous activity on gene-specific Atail length extension appears rather moderate at steady state. 17,18,21 Importantly, GLD-4 depends on its co-factor GLS-1 for efficient PAP activity in tethering assays, suggesting that the GLD-4/GLS-1 complex comprises the active cytoPAP. 18 To reveal the impact of GLD-4 on global poly(A) tail metabolism, we recently measured A-tail lengths on bulk RNA and found that poly(A)-tails were only mildly reduced in the absence of GLD-4 16 or GLS-1 (unpublished results), arguing that GLD-4 cytoPAP may have an intrinsic low enzymatic activity that is similar to that of its nuclear counterparts.…”

“…16,17 GLD-4 promotes translational efficiency GLD-4 is in its enzymatic domain evolutionarily most similar to that of noncanonical TRF4-type poly(A) polymerase family members. 18,19 Yet, in many organisms TRF4 proteins are nuclear enzymes that primarily target a variety of non-coding RNA substrates by adding short adenosine stretches (»10-20 nts) for exosomemediated degradation. 20 By contrast, GLD-4 is predominantly located in the cytoplasm, 18 presumably targeting mRNAs for enhanced expression.…”

“…18,19 Yet, in many organisms TRF4 proteins are nuclear enzymes that primarily target a variety of non-coding RNA substrates by adding short adenosine stretches (»10-20 nts) for exosomemediated degradation. 20 By contrast, GLD-4 is predominantly located in the cytoplasm, 18 presumably targeting mRNAs for enhanced expression. GLD-4 shows poly(A) polymerase activity in a heterologous in vivo tethering system, but its endogenous activity on gene-specific Atail length extension appears rather moderate at steady state.…”