scholarly journals Multi-modal regulation of C. elegans hermaphrodite spermatogenesis by the GLD-1-FOG-2 complex

2018 ◽  
Author(s):  
Shuang Hu ◽  
Lauren E. Ryan ◽  
Ebru Kaymak ◽  
Lindsay Freeberg ◽  
Te-Wen Lo ◽  
...  
Keyword(s):  
Sex Determination ◽  
Germ Cell ◽  
Germ Cells ◽  
Translational Control ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Wild Type ◽  
C Elegans ◽  
Target Mrnas ◽  
Mrna Targets

AbstractProper germ cell sex determination in Caenorhabditis nematodes requires a network of RNA-binding proteins (RBPs) and their target mRNAs. In some species, changes in this network enabled limited XX spermatogenesis, and thus self-fertility. In C. elegans, one of these selfing species, the global sex-determining gene tra-2 is regulated in germ cells by a conserved RBP, GLD-1, via the 3’ untranslated region (3’UTR) of its transcript. A C. elegans-specific GLD-1 cofactor, FOG-2, is also required for hermaphrodite sperm fate, but how it modifies GLD-1 function is unknown. Germline feminization in gld-1 and fog-2 null mutants has been interpreted as due to cell-autonomous elevation of TRA-2 translation. Consistent with the proposed role of FOG-2 in translational control, the abundance of nearly all GLD-1 target mRNAs (including tra-2) is unchanged in fog-2 mutants. Epitope tagging reveals abundant TRA-2 expression in somatic tissues, but an undetectably low level in wild-type germ cells. Loss of gld-1 function elevates germline TRA-2 expression to detectable levels, but loss of fog-2 function does not. A simple quantitative model of tra-2 activity constrained by these results can successfully sort genotypes into normal or feminized groups. Surprisingly, fog-2 and gld-1 activity enable the sperm fate even when GLD-1 cannot bind to the tra-2 3’ UTR. This suggests the GLD-1-FOG-2 complex regulates uncharacterized sites within tra-2, or other mRNA targets. Finally, we quantify the RNA-binding capacities of dominant missense alleles of GLD-1 that act genetically as “hyperrepressors” of tra-2 activity. These variants bind RNA more weakly in vitro than does wild-type GLD-1. These results indicate that gld-1 and fog-2 regulate germline sex via multiple interactions, and that our understanding of the control and evolution of germ cell sex determination in the C. elegans hermaphrodite is far from complete.

scholarly journals Positive mRNA Translational Control in Germ Cells by Initiation Factor Selectivity

2015 ◽  
Vol 2015 ◽  
pp. 1-11 ◽  
Author(s):  
Andrew J. Friday ◽  
Brett D. Keiper
Keyword(s):  
Germ Cell ◽  
Germ Cells ◽  
Translation Initiation ◽  
Translational Control ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Positive Function ◽  
Initiation Factor ◽  
C Elegans ◽  
Initiation Factors

Ultimately, the production of new proteins in undetermined cells pushes them to new fates. Other proteins hold a stem cell in a mode of self-renewal. In germ cells, these decision-making proteins are produced largely from translational control of preexisting mRNAs. To date, all of the regulation has been attributed to RNA binding proteins (RBPs) that repress mRNAs in many models of germ cell development (Drosophila, mouse,C. elegans, andXenopus). In this review, we focus on the selective, positive function of translation initiation factors eIF4E and eIF4G, which recruit mRNAs to ribosomes upon derepression. Evidence now shows that the two events are not separate but rather are coordinated through composite complexes of repressors and germ cell isoforms of eIF4 factors. Strikingly, the initiation factor isoforms are themselves mRNA selective. The mRNP complexes of translation factors and RBPs are built on specific populations of mRNAs to prime them for subsequent translation initiation. Simple rearrangement of the partners causes a dormant mRNP to become synthetically active in germ cells when and where they are required to support gametogenesis.

scholarly journals CPB-3 and CGH-1 localize to motile particles within dendrites in C. elegans PVD sensory neurons

2021 ◽  
Vol 14 (1) ◽  
Author(s):  
Kathrin Spendier ◽  
Eugenia C. Olesnicky ◽  
Daniel Forand ◽  
Margaret Wolf ◽  
Darrell J. Killian
Keyword(s):  
Translational Control ◽  
Binding Proteins ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Rna Helicases ◽  
Cytoplasmic Polyadenylation ◽  
C Elegans ◽  
Mrna Targets ◽  
Dead Box ◽  
Dendrite Development

Abstract Objective RNA-binding proteins (RBPs) are important regulators of gene expression that influence mRNA splicing, stability, localization, transport, and translational control. In particular, RBPs play an important role in neurons, which have a complex morphology. Previously, we showed that there are many RBPs that play a conserved role in dendrite development in Drosophila dendritic arborization neurons and Caenorhabditis elegans (C. elegans) PVD neurons including the cytoplasmic polyadenylation element binding proteins (CPEBs), Orb in Drosophila and CPB-3 in C. elegans, and the DEAD box RNA helicases, Me31B in Drosophila and CGH-1 in C. elegans. During these studies, we observed that fluorescently-labeled CPB-3 and CGH-1 localize to cytoplasmic particles that are motile, and our research aims to further characterize these RBP-containing particles in live neurons. Results Here we extend on previous work to show that CPB-3 and CGH-1 localize to motile particles within dendrites that move at a speed consistent with microtubule-based transport. This is consistent with a model in which CPB-3 and CGH-1 influence dendrite development through the transport and localization of their mRNA targets. Moreover, CPB-3 and CGH-1 rarely localize to the same particles suggesting that these RBPs function in discrete ribonucleoprotein particles (RNPs) that may regulate distinct mRNAs.

scholarly journals Investigating the Roles of RNA Binding Protein Combinations in Neuronal Function and Organismal Behavior

2019 ◽  
Vol 4 (Spring 2019) ◽  
Author(s):  
Alexa Vandenburg
Keyword(s):  
Binding Sites ◽  
Neuronal Development ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Binding Protein ◽  
Rna Binding Protein ◽  
Wild Type ◽  
C Elegans ◽  
Neuronal Gene ◽  
Touch Response

The Norris lab recently identified two RNA binding proteins required for proper neuron-specific splicing. The lab conducted touch- response behavioral assays to assess the function of these proteins in touch-sensing neurons. After isolating C. elegans worms with specific phenotypes, the lab used automated computer tracking and video analysis to record the worms’ behavior. The behavior of mutant worms differed from that of wild-type worms. The Norris lab also discovered two possible RNA binding protein sites in SAD-1, a neuronal gene implicated in the neuronal development of C. elegans1. These two binding sites may control the splicing of SAD-1. The lab transferred mutated DNA into the genome of wild-type worms by injecting a mutated plasmid. The newly transformed worms fluoresced green, indicating that the two binding sites control SAD-1 splicing.

scholarly journals CELF RNA binding proteins promote axon regeneration in C. elegans and mammals through alternative splicing of Syntaxins

eLife ◽  
2016 ◽  
Vol 5 ◽  
Author(s):  
Lizhen Chen ◽  
Zhijie Liu ◽  
Bing Zhou ◽  
Chaoliang Wei ◽  
Yu Zhou ◽  
...  
Keyword(s):  
Gene Expression ◽  
Transcriptional Regulation ◽  
Alternative Splicing ◽  
Binding Proteins ◽  
Rna Binding ◽  
Axon Regeneration ◽  
Rna Binding Proteins ◽  
Mrna Isoforms ◽  
C Elegans ◽  
Mrna Targets

Axon injury triggers dramatic changes in gene expression. While transcriptional regulation of injury-induced gene expression is widely studied, less is known about the roles of RNA binding proteins (RBPs) in post-transcriptional regulation during axon regeneration. In C. elegans the CELF (CUGBP and Etr-3 Like Factor) family RBP UNC-75 is required for axon regeneration. Using crosslinking immunoprecipitation coupled with deep sequencing (CLIP-seq) we identify a set of genes involved in synaptic transmission as mRNA targets of UNC-75. In particular, we show that UNC-75 regulates alternative splicing of two mRNA isoforms of the SNARE Syntaxin/unc-64. In C. elegans mutants lacking unc-75 or its targets, regenerating axons form growth cones, yet are deficient in extension. Extending these findings to mammalian axon regeneration, we show that mouse Celf2 expression is upregulated after peripheral nerve injury and that Celf2 mutant mice are defective in axon regeneration. Further, mRNAs for several Syntaxins show CELF2 dependent regulation. Our data delineate a post-transcriptional regulatory pathway with a conserved role in regenerative axon extension.

scholarly journals NANOS2 suppresses the cell cycle by repressing mTORC1 activators in embryonic male germ cells

2020 ◽  
Author(s):  
Ryuki Shimada ◽  
Hiroko Koike ◽  
Takamasa Hirano ◽  
Yumiko Saga
Keyword(s):  
Cell Cycle ◽  
Germ Cell ◽  
Germ Cells ◽  
Rna Binding ◽  
Initial Step ◽  
Sexually Dimorphic ◽  
Wild Type ◽  
Male Germ Cells ◽  
Cycle Arrest ◽  
Male Specific

AbstractDuring murine germ cell development, male germ cells enter the mitotically arrested G0 stage, which is an initial step of sexually dimorphic differentiation. The male specific RNA-binding protein NANOS2 has a key role in suppressing the cell cycle in germ cells. However, the detailed mechanism of how NANOS2 regulates the cell cycle remains unclear. Using single-cell RNA sequencing (scRNA-seq), we extracted the cell cycle state of each germ cell in wild-type and Nanos2-KO testes, and revealed that Nanos2 expression starts in mitotic cells and induces mitotic arrest. We also found that NANOS2 and p38 MAPK work in parallel to regulate the cell cycle, suggesting that several different cascades are involved in the induction of cell cycle arrest. Furthermore, we identified Rheb, a regulator of mTORC1, and Ptma as possible targets of NANOS2. We propose that the repression of the cell cycle is a primary function of NANOS2 and that it is mediated via the suppression of mTORC1 activity by repressing Rheb in a post-transcriptional manner.

scholarly journals The mir-35 family links maternal germline sex to embryonic viability in C. elegans

2019 ◽  
Author(s):  
Lars Benner ◽  
Katherine Prothro ◽  
Katherine McJunkin
Keyword(s):  
Sex Determination ◽  
Germ Cells ◽  
Sex Reversal ◽  
Loss Of Function ◽  
Wild Type ◽  
Partial Loss ◽  
C Elegans ◽  
Germline Sex Determination ◽  
Embryonic Viability ◽  
Male Gene

AbstractThe germline sex determination pathway in C. elegans determines whether germ cells develop as oocytes or sperm, with no previously known effect on viability. The mir-35 family of microRNAs are expressed in the C. elegans germline and embryo and are essential for both viability and normal hermaphroditic sex determination, preventing aberrant male gene expression in XX hermaphrodite embryos. Here we show that combining feminizing mutations with partial loss of function of the mir-35 family results in enhanced penetrance embryonic lethality that preferentially kills XO animals. This lethal phenotype is due to altered signaling through the germline sex determination pathway, and maternal germline feminization is sufficient to induce enhanced lethality. These findings reveal a surprising pleiotropy of sperm-fate promoting pathways on organismal viability. Overall, our results demonstrate an unexpectedly strong link between sex determination and embryonic viability, and suggest that in wild type animals, mir-35 family members buffer against misregulation of pathways outside the sex determination program, allowing for clean sex reversal rather than deleterious effects of perturbing sex determination genes.

scholarly journals ANTAGONISTIC ROLES FOR ATAXIN-2 STRUCTURED AND DISORDERED DOMAINS IN RNP CONDENSATION

2020 ◽  
Author(s):  
Amanjot Singh ◽  
Joern Huelsmeier ◽  
Arvind Reddy Kandi ◽  
Sai Shruti Pothapragada ◽  
Jens Hillebrand ◽  
...  
Keyword(s):  
Translational Control ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Biological Significance ◽  
Spinocerebellar Ataxia Type ◽  
Granule Formation ◽  
Intrinsically Disordered ◽  
Target Mrnas ◽  
Ataxin 2 ◽  
Rnp Granules

ABSTRACTAtaxin-2 is a conserved translational control protein associated with spinocerebellar ataxia type II (SCA2) and amyotrophic lateral sclerosis (ALS) as well as an important target for ALS therapeutics under development. Despite its clinical and biological significance, Ataxin-2’s activities, mechanisms and functions are not well understood. While Drosophila Ataxin-2 (Atx2) mediates mRNP condensation via a C-terminal intrinsically disordered domain (cIDR), how Ataxin-2 IDRs work with structured (Lsm, Lsm-AD and PAM2) domains to enable positive and negative regulation of target mRNAs remains unclear. Using TRIBE (Targets of RNA-Binding Proteins Identified by Editing) technology, we identified and analysed Atx-2 target mRNAs in the Drosophila brain. We show that Atx2 preferentially interacts with AU-rich elements (AREs) in 3’UTRs and plays a broad role in stabilization of identified target mRNAs. Strikingly, Atx2 interaction with its targets is dependent on the cIDR domain required for neuronal-granule formation. In contrast, Atx2 lacking its Lsm domain not only interacts more efficiently with the target mRNA identified, but also forms larger RNP granules. Providing an extensive dataset of Atx2-interacting brain mRNAs, our results demonstrate that Atx2: (a) interacts with target mRNAs within RNP granules; (b) modulates the turnover of these target mRNAs; (c) has an additional essential role outside of mRNP granules; and (d) contains distinct protein domains that drive or oppose RNP-granule assembly. These findings increase understanding of neuronal translational control mechanisms and inform Ataxin-2-based interventions in development for SCA2 and ALS.

scholarly journals Functional reconstruction of NANOS3 expression in the germ cell lineage by a novel transgenic reporter reveals distinct subcellular localizations of NANOS3

Reproduction ◽  
2010 ◽  
Vol 139 (2) ◽  
pp. 381-393 ◽  
Author(s):  
Masashi Yamaji ◽  
Takashi Tanaka ◽  
Mayo Shigeta ◽  
Shinichiro Chuma ◽  
Yumiko Saga ◽  
...  
Keyword(s):  
Germ Cell ◽  
Germ Cells ◽  
Binding Proteins ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Primordial Germ Cells ◽  
Cell Lineage ◽  
Regulatory Elements ◽  
Initiation Factor ◽  
Processing Bodies

Mutations of RNA-binding proteins such as NANOS3, TIAL1, and DND1 in mice have been known to result in the failure of survival and/or proliferation of primordial germ cells (PGCs) soon after their fate is specified (around embryonic day (E) 8.0), leading to the infertility of these animals. However, the mechanisms of actions of these RNA-binding proteins remain largely unresolved. As a foundation to explore the role of these RNA-binding proteins in germ cells, we established a novel transgenic reporter strain that expresses NANOS3 fused with EGFP under the control of Nanos3 regulatory elements. NANOS3–EGFP exhibited exclusive expression in PGCs as early as E7.25, and continued to be expressed in female germ cells until around E14.5 and in male germ cells throughout the fetal period with declining expression levels after E16.5. NANOS3–EGFP resumed strong expression in postnatal spermatogonia and continued to be expressed in undifferentiated spermatogonial cells in adults. Importantly, the Nanos3–EGFP transgene rescued the sterile phenotype of Nanos3 homozygous mutants, demonstrating the functional equivalency of NANOS3–EGFP with endogenous NANOS3. We found that throughout germ cell development, a predominant amount of  NANOS3–EGFP co-localized with TIAL1 (also known as TIAR) and phosphorylated eukaryotic initiation factor 2α, markers for the stress granules, whereas a fraction of it showed co-localization with DCP1A, a marker for the processing bodies. On the other hand, NANOS3–EGFP did not co-localize with Tudor domain-containing protein 1, a marker for the intermitochondrial cements, in spermatogenic cells. These findings unveil the presence of distinct posttranscriptional regulations in PGCs soon after their specification, for which RNA-binding proteins such as NANOS3 and TIAL1 would play critical functions.

nos-1 and nos-2, two genes related to Drosophila nanos, regulate primordial germ cell development and survival in Caenorhabditis elegans

Development ◽  
1999 ◽  
Vol 126 (21) ◽  
pp. 4861-4871 ◽  
Author(s):  
K. Subramaniam ◽  
G. Seydoux
Keyword(s):  
Caenorhabditis Elegans ◽  
Germ Cell ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Primordial Germ Cell ◽  
Gene Families ◽  
Cell Development ◽  
Embryonic Patterning ◽  
Germ Cell Development ◽  
C Elegans

In Drosophila, the posterior determinant nanos is required for embryonic patterning and for primordial germ cell (PGC) development. We have identified three genes in Caenorhabditis elegans that contain a putative zinc-binding domain similar to the one found in nanos, and show that two of these genes function during PGC development. Like Drosophila nanos, C. elegans nos-1 and nos-2 are not generally required for PGC fate specification, but instead regulate specific aspects of PGC development. nos-2 is expressed in PGCs around the time of gastrulation from a maternal RNA associated with P granules, and is required for the efficient incorporation of PGCs into the somatic gonad. nos-1 is expressed in PGCs after gastrulation, and is required redundantly with nos-2 to prevent PGCs from dividing in starved animals and to maintain germ cell viability during larval development. In the absence of nos-1 and nos-2, germ cells cease proliferation at the end of the second larval stage, and die in a manner that is partially dependent on the apoptosis gene ced-4. Our results also indicate that putative RNA-binding proteins related to Drosophila Pumilio are required for the same PGC processes as nos-1 and nos-2. These studies demonstrate that evolutionarily distant organisms utilize conserved factors to regulate early germ cell development and survival, and that these factors include members of the nanos and pumilio gene families.

scholarly journals Culture medium, gas atmosphere and MAPK inhibition affect regulation of RNA-binding protein targets during mouse preimplantation development

Reproduction ◽  
2011 ◽  
Vol 142 (5) ◽  
pp. 689-698 ◽  
Author(s):  
Michele D Calder ◽  
Patricia H Watson ◽  
Andrew J Watson
Keyword(s):  
Mrna Expression ◽  
Culture Medium ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Mapk Pathway ◽  
Preimplantation Development ◽  
Cell Stage ◽  
Target Mrnas ◽  
Mrna Targets ◽  
Gas Atmosphere

During oogenesis, mammalian oocytes accumulate maternal mRNAs that support the embryo until embryonic genome activation. RNA-binding proteins (RBP) may regulate the stability and turnover of maternal and embryonic mRNAs. We hypothesised that varying embryo culture conditions, such as culture medium, oxygen tension and MAPK inhibition, affects regulation of RBPs and their targets during preimplantation development. STAU1, ELAVL1, KHSRP and ZFP36 proteins and mRNAs were detected throughout mouse preimplantation development, whereasElavl2mRNA decreased after the two-cell stage. Potential target mRNAs of RBP regulation,Gclc,Slc2a1andSlc7a1were detected during mouse preimplantation development.GclcmRNA was significantly elevated in embryos cultured in Whitten's medium compared with embryos cultured in KSOMaa, andGclcmRNA was elevated under high-oxygen conditions. Inhibition of the p38 MAPK pathway reducedSlc7a1mRNA expression while inhibition of ERK increasedSlc2a1mRNA expression. The half-lives of the potential RBP mRNA targets are not regulated in parallel;Slc2a1mRNA displayed the longest half-life. Our results indicate that mRNAs and proteins encoding five RBPs are present during preimplantation development and more importantly, demonstrate that expression of RBP target mRNAs are regulated by culture medium, gas atmosphere and MAPK pathways.

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