scholarly journals Rap2 and TNIK control Plexin-dependent synaptic tiling in C. elegans

2017 ◽  
Author(s):  
Xi Chen ◽  
Akihiro C.E. Shibata ◽  
Ardalan Hendi ◽  
Mizuki Kurashina ◽  
Ethan Fortes ◽  
...  
Keyword(s):  
Synapse Formation ◽  
Small Gtpase ◽  
Negative Regulator ◽  
Gtpase Activity ◽  
C Elegans ◽  
Receptor Interactions ◽  
Presynaptic Assembly ◽  
Intracellular Mechanisms

AbstractDuring development, neurons form synapses with their fate-determined targets. While we begin to elucidate the mechanisms by which extracellular ligand-receptor interactions enhance synapse specificity by inhibiting synaptogenesis, our knowledge about their intracellular mechanisms remains limited. Here we show that Rap2 GTPase (rap-2) and its effector, TNIK (mig-15), act downstream of Plexin (plx-1) to restrict presynaptic assembly and to form tiled synaptic innervation in C. elegans. Both constitutively GTP- and GDP-forms of rap-2 mutants exhibit synaptic tiling defects as plx-1 mutants, suggesting that cycling of the RAP-2 nucleotide state is critical for synapse inhibition. Consistently, RAP-2 activity is locally suppressed by PLX-1. Excessive ectopic synapse formation in mig-15 mutants causes a severe synaptic tiling defect. Conversely, overexpression of mig-15 strongly inhibited synapse formation, suggesting that mig-15 is a negative regulator of synapse formation. These results reveal that subcellular regulation of small GTPase activity by Plexin shapes proper synapse patterning in vivo.

scholarly journals Rap2 and TNIK control Plexin-dependent tiled synaptic innervation in C. elegans

eLife ◽  
2018 ◽  
Vol 7 ◽  
Author(s):  
Xi Chen ◽  
Akihiro CE Shibata ◽  
Ardalan Hendi ◽  
Mizuki Kurashina ◽  
Ethan Fortes ◽  
...  
Keyword(s):  
Synapse Formation ◽  
Small Gtpase ◽  
Negative Regulator ◽  
Gtpase Activity ◽  
C Elegans ◽  
Receptor Interactions ◽  
Presynaptic Assembly ◽  
Intracellular Mechanisms

During development, neurons form synapses with their fate-determined targets. While we begin to elucidate the mechanisms by which extracellular ligand-receptor interactions enhance synapse specificity by inhibiting synaptogenesis, our knowledge about their intracellular mechanisms remains limited. Here we show that Rap2 GTPase (rap-2) and its effector, TNIK (mig-15), act genetically downstream of Plexin (plx-1) to restrict presynaptic assembly and to form tiled synaptic innervation in C. elegans. Both constitutively GTP- and GDP-forms of rap-2 mutants exhibit synaptic tiling defects as plx-1 mutants, suggesting that cycling of the RAP-2 nucleotide state is critical for synapse inhibition. Consistently, PLX-1 suppresses local RAP-2 activity. Excessive ectopic synapse formation in mig-15 mutants causes a severe synaptic tiling defect. Conversely, overexpression of mig-15 strongly inhibited synapse formation, suggesting that mig-15 is a negative regulator of synapse formation. These results reveal that subcellular regulation of small GTPase activity by Plexin shapes proper synapse patterning in vivo.

scholarly journals The RhoGAP SPV-1 regulates calcium signaling to control the contractility of theC. elegansspermatheca during embryo transits

2018 ◽  
Author(s):  
Jeff Bouffard ◽  
Alyssa D. Cecchetelli ◽  
Coleman Clifford ◽  
Kriti Sethi ◽  
Ronen Zaidel-Bar ◽  
...  
Keyword(s):  
Calcium Signaling ◽  
Signaling Pathways ◽  
Calcium Release ◽  
Small Gtpase ◽  
Negative Regulator ◽  
Calcium Signal ◽  
C Elegans ◽  
Spatial Regulation ◽  
Calcium Indicator

AbstractContractility of the non-muscle and smooth muscle cells that comprise biological tubing is regulated by the Rho-ROCK and calcium signaling pathways. Although many molecular details about these signaling pathways are known, less is known about how they are coordinated spatiotemporally in biological tubes. The spermatheca of theC. elegansreproductive system enables study of the signaling pathways regulating actomyosin contractility in live adult animals. The RhoGAP SPV-1 was previously identified as a negative regulator of RHO-1/Rho and spermathecal contractility. Here, we uncover a role for SPV-1 as a key regulator of calcium signaling.spv-1mutants expressing the calcium indicator GCaMP in the spermatheca exhibit premature calcium release, elevated calcium levels, and disrupted spatial regulation of calcium signaling during spermathecal contraction. Although RHO-1 is required for spermathecal contractility, RHO-1 does not play a significant role in regulating calcium. In contrast, activation of CDC-42 recapitulates many aspects ofspv-1mutant calcium signaling. Depletion ofcdc-42by RNAi does not suppress the premature or elevated calcium signal seen inspv-1mutants, suggesting other targets remain to be identified. Our results suggest SPV-1 works through both the Rho-ROCK and calcium signaling pathways to coordinate cellular contractility.Highlight SummaryThroughin vivoimaging of the calcium sensor GCaMP, we show that the RhoGAP SPV-1 is a key regulator of calcium signaling in theC. elegansspermatheca. Our data suggests SPV-1 acts at least partially through the small GTPase CDC-42 to modulate calcium signaling, while also acting on RHO-1 to modulate Rho-ROCK signaling. This places SPV-1 as a central regulator of cellular contractility.

scholarly journals RHO-1 and the Rho GEF RHGF-1 interact with UNC-6/Netrin signaling to regulate growth cone protrusion and microtubule organization in C. elegans

2019 ◽  
Author(s):  
Mahekta R. Gujar ◽  
Aubrie M. Stricker ◽  
Erik A. Lundquist
Keyword(s):  
Axon Guidance ◽  
Growth Cone ◽  
Neural Circuit ◽  
Growth Cones ◽  
Negative Regulator ◽  
Microtubule Organization ◽  
C Elegans ◽  
Guidance Cue ◽  
Rho Gef

AbstractUNC-6/Netrin is a conserved axon guidance cue that directs growth cone migrations in the dorsal-ventral axis of C. elegans and in the vertebrate spinal cord. UNC-6/Netrin is expressed in ventral cells, and growth cones migrate ventrally toward or dorsally away from UNC-6/Netrin. Recent studies of growth cone behavior during outgrowth in vivo in C. elegans have led to a polarity/protrusion model in directed growth cone migration away from UNC-6/Netrin. In this model, UNC-6/Netrin first polarizes the growth cone via the UNC-5 receptor, leading to dorsally biased protrusion and F-actin accumulation. UNC-6/Netrin then regulates protrusion based on this polarity. The receptor UNC-40/DCC drives protrusion dorsally, away from the UNC-6/Netrin source, and the UNC-5 receptor inhibits protrusion ventrally, near the UNC-6/Netrin source, resulting in dorsal migration. UNC-5 inhibits protrusion in part by excluding microtubules from the growth cone, which are pro-protrusive. Here we report that the RHO-1/RhoA GTPase and its activator GEF RHGF-1 inhibit growth cone protrusion and MT accumulation in growth cones, similar to UNC-5. However, growth cone polarity of protrusion and F-actin were unaffected by RHO-1 and RHGF-1. Thus, RHO-1 signaling acts specifically as a negative regulator of protrusion and MT accumulation, and not polarity. Genetic interactions suggest that RHO-1 and RHGF-1 act with UNC-5, as well as with a parallel pathway, to regulate protrusion. The cytoskeletal interacting molecule UNC-33/CRMP was required for RHO-1 activity to inhibit MT accumulation, suggesting that UNC-33/CRMP might act downstream of RHO-1. In sum, these studies describe a new role of RHO-1 and RHGF-1 in regulation of growth cone protrusion by UNC-6/Netrin.Author SummaryNeural circuits are formed by precise connections between axons. During axon formation, the growth cone leads the axon to its proper target in a process called axon guidance. Growth cone outgrowth involves asymmetric protrusion driven by extracellular cues that stimulate and inhibit protrusion. How guidance cues regulate growth cone protrusion in neural circuit formation is incompletely understood. This work shows that the signaling molecule RHO-1 acts downstream of the UNC-6/Netrin guidance cue to inhibit growth cone protrusion in part by excluding microtubules from the growth cone, which are structural elements that drive protrusion.

scholarly journals The Amyloid Precursor-like Protein APL-1 Regulates Axon Regeneration

2018 ◽  
Author(s):  
Lewie Zeng ◽  
Rachid El Bejjani ◽  
Marc Hammarlund
Keyword(s):  
Motor Neurons ◽  
Neuronal Development ◽  
Axon Regeneration ◽  
Neuronal Response ◽  
Small Gtpase ◽  
C Elegans ◽  
Protein Levels ◽  
Mature Neurons ◽  
And Function

AbstractMembers of the Amyloid Precursor Protein (APP) family have important functions during neuronal development. However, their physiological functions in the mature nervous system are not fully understood. Here we use the C. elegans GABAergic motor neurons to study the post-developmental function of the APP-like protein APL-1 in vivo. We find that apl-1 has minimum roles in the maintenance of gross neuron morphology and function. However, we show that apl-1 is an inhibitor of axon regeneration, acting on mature neurons to limit regrowth in response to injury. The small GTPase Rab6/RAB-6.2 also inhibits regeneration, and does so in part by maintaining protein levels of APL-1. To inhibit regeneration, APL-1 functions via the E2 domain of its ectodomain; the cytoplasmic tail, transmembrane anchoring, and the E1 domain are not required for this function. Our data defines a novel role for APL-1 in modulating the neuronal response to injury.

scholarly journals Kinesin-3 mediated delivery of presynaptic neurexin stabilizes growing dendritic spines and postsynaptic components in vivo

2021 ◽  
Author(s):  
Devyn Oliver ◽  
Shankar Ramachandran ◽  
Alison Philbrook ◽  
Christopher M Lambert ◽  
Ken C. Q. Nguyen ◽  
...  
Keyword(s):  
Motor Neurons ◽  
Dendritic Spines ◽  
Synapse Formation ◽  
Synaptic Activity ◽  
Presynaptic Terminals ◽  
Adhesion Protein ◽  
C Elegans ◽  
Specific Regulation ◽  
Receptor Clusters

A high degree of cell and circuit-specific regulation has complicated efforts to precisely define roles for synaptic adhesion proteins in establishing circuit connectivity. Here, we take advantage of the strengths of C. elegans for cell-specific analyses to investigate molecular coordination of pre- and postsynaptic development. We show that developing dendritic spines emerge from the dendrites of wild type GABAergic motor neurons following the localization of active zone proteins and the formation of immature synaptic vesicle assemblies in presynaptic terminals. Similarly, clusters of postsynaptic receptors and F-actin are visible in GABAergic dendrites prior to spine outgrowth. Surprisingly, these developmental processes occur without a requirement for synaptic activity. Likewise, the initial stages of spine outgrowth and receptor clustering are not altered by deletion of the C. elegans ortholog of the transsynaptic adhesion protein, neurexin/NRX-1. Over time, however, dendritic spines and postsynaptic receptor clusters are destabilized in the absence of presynaptic NRX-1/neurexin and collapse prior to adulthood. The kinesin-3 family member, UNC-104, delivers NRX-1 to presynaptic terminals and ongoing UNC-104 delivery is required into adulthood for the maintenance of postsynaptic structure. Our findings provide novel insights into the temporal order of synapse formation events in vivo and demonstrate a requirement for transsynaptic adhesion in stabilizing mature circuit connectivity.

scholarly journals Conserved and divergent aspects of Robo receptor signaling and regulation between Drosophila Robo1 and C. elegans SAX-3

Genetics ◽  
2021 ◽  
Vol 217 (3) ◽  
Author(s):  
Trent Daiber ◽  
Christine J VanderZwan-Butler ◽  
Greg J Bashaw ◽  
Timothy A Evans
Keyword(s):  
Fruit Fly ◽  
Negative Regulator ◽  
Genetic Rescue ◽  
Signaling Mechanism ◽  
Endosomal Sorting ◽  
C Elegans ◽  
Midline Crossing ◽  
Robo Receptors ◽  
Guidance Receptors

Abstract The evolutionarily conserved Roundabout (Robo) family of axon guidance receptors control midline crossing of axons in response to the midline repellant ligand Slit in bilaterian animals including insects, nematodes, and vertebrates. Despite this strong evolutionary conservation, it is unclear whether the signaling mechanism(s) downstream of Robo receptors are similarly conserved. To directly compare midline repulsive signaling in Robo family members from different species, here we use a transgenic approach to express the Robo family receptor SAX-3 from the nematode Caenorhabditis elegans in neurons of the fruit fly, Drosophila melanogaster. We examine SAX-3’s ability to repel Drosophila axons from the Slit-expressing midline in gain of function assays, and test SAX-3’s ability to substitute for Drosophila Robo1 during fly embryonic development in genetic rescue experiments. We show that C. elegans SAX-3 is properly translated and localized to neuronal axons when expressed in the Drosophila embryonic CNS, and that SAX-3 can signal midline repulsion in Drosophila embryonic neurons, although not as efficiently as Drosophila Robo1. Using a series of Robo1/SAX-3 chimeras, we show that the SAX-3 cytoplasmic domain can signal midline repulsion to the same extent as Robo1 when combined with the Robo1 ectodomain. We show that SAX-3 is not subject to endosomal sorting by the negative regulator Commissureless (Comm) in Drosophila neurons in vivo, and that peri-membrane and ectodomain sequences are both required for Comm sorting of Drosophila Robo1.

scholarly journals The RabGAP TBC-11 controls Argonaute localization for proper microRNA function in C. elegans

PLoS Genetics ◽  
2021 ◽  
Vol 17 (4) ◽  
pp. e1009511
Author(s):  
Pascale Michaud ◽  
Vivek Nilesh Shah ◽  
Pauline Adjibade ◽  
Francois Houle ◽  
Miguel Quévillon Huberdeau ◽  
...  
Keyword(s):  
Intracellular Trafficking ◽  
Small Gtpase ◽  
Gene Repression ◽  
High Molecular Weight Complex ◽  
Forward Genetic Screen ◽  
C Elegans ◽  
Argonaute Proteins ◽  
Microrna Pathway ◽  
Microrna Function

Once loaded onto Argonaute proteins, microRNAs form a silencing complex called miRISC that targets mostly the 3’UTR of mRNAs to silence their translation. How microRNAs are transported to and from their target mRNA remains poorly characterized. While some reports linked intracellular trafficking to microRNA activity, it is still unclear how these pathways coordinate for proper microRNA-mediated gene silencing and turnover. Through a forward genetic screen using Caenorhabditis elegans, we identified the RabGAP tbc-11 as an important factor for the microRNA pathway. We show that TBC-11 acts mainly through the small GTPase RAB-6 and that its regulation is required for microRNA function. The absence of functional TBC-11 increases the pool of microRNA-unloaded Argonaute ALG-1 that is likely associated to endomembranes. Furthermore, in this condition, this pool of Argonaute accumulates in a perinuclear region and forms a high molecular weight complex. Altogether, our data suggest that the alteration of TBC-11 generates a fraction of ALG-1 that cannot bind to target mRNAs, leading to defective gene repression. Our results establish the importance of intracellular trafficking for microRNA function and demonstrate the involvement of a small GTPase and its GAP in proper Argonaute localization in vivo.

scholarly journals Molecular Evolution of a Sex Determination Protein: FEM-2 (PP2C) in Caenorhabditis

Genetics ◽  
1998 ◽  
Vol 149 (3) ◽  
pp. 1353-1362 ◽  
Author(s):  
Dave Hansen ◽  
Dave Pilgrim
Keyword(s):  
Sex Determination ◽  
Phosphatase Activity ◽  
Catalytic Domain ◽  
Sequence Similarity ◽  
Signal Transduction Pathway ◽  
Negative Regulator ◽  
Amino Terminus ◽  
C Elegans

Abstract Somatic sex determination in Caenorhabditis elegans involves a signal transduction pathway linking a membrane receptor to a transcription factor. The fem-2 gene is central to this pathway, producing a protein phosphatase (FEM-2) of the type 2C (PP2C). FEM-2 contains a long amino terminus that is absent in canonical PP2C enzymes. The function of this domain is difficult to predict, since it shows no sequence similarity to any other known proteins or motifs. Here we report the cloning of the fem-2 homologue from Caenorhabditis briggsae (Cb-fem-2). The sequence identity is much higher than that observed for other C. briggsae homologues of C. elegans sex determination proteins. However, this level is not uniform across the entire lengths of the proteins; it is much lower in the amino termini. Thus, the two domains of the same protein are evolving at different rates, suggesting that they have different functional constraints. Consistent with this, Cb-FEM-2 is able to replace some, but not all, of the Ce-FEM-2 in vivo function. We show that removal of the amino terminus from Ce-FEM-2 has no effect on its in vitro phosphatase activity, or its ability to replace the in vivo function of a yeast PP2C enzyme, but that it is necessary for proper FEM-2 function in worms. This demonstrates that the amino terminus is not an extended catalytic domain or a direct negative regulator of phosphatase activity.

Abstract 114: Downregulation of Small GTPase Rnd3 in Cardiomyocytes Leads to Hyperinflammatory Response in Post-myocardial Infarction Heart

2016 ◽  
Vol 119 (suppl_1) ◽  
Author(s):  
Yuan Dai ◽  
Xiangsheng Yang ◽  
Xiaojing Yue ◽  
Tingli Yang ◽  
Xin Yi ◽  
...  
Keyword(s):  
Myocardial Infarction ◽  
Inflammatory Response ◽  
Cardiac Function ◽  
Inflammatory Cells ◽  
Small Gtpase ◽  
Negative Regulator ◽  
Fine Tuning ◽  
Acute Mi

Background: Hyperinflammatory response leads to maladaptive healing and remodeling after acute myocardial infarction (MI), while a balanced inflammatory response is essential for post-MI repair and remodeling. Most of the studies have focused on inflammatory cells, extracellular matrices and myofibroblasts in the regulation of acute MI-induced inflammatory response, the investigation of crucial role of cardiomyocytes in the initiation and regulation of the responsive inflammation in heart has been overlooked. Subjects and Methods: To demonstrate that small G protein Rnd3 functions as a negative regulator to refine the inflammatory response by preventing excessive inflammation in cardiomyocytes after acute MI. Genetic mouse models with cardiomyocyte-specific knockout and overexpression of Rnd3 were studied along with multiple cell culture approaches. An acute MI was introduced by ligation of the left anterior descending artery. Results: Mice with cardiomyocyte-specific deletion of Rnd3 (heterozygous, Rnd3 +/- ) showed a significantly higher mortality, worse cardiac function, and increased infarct size compared to wild-type control mice after MI. Massive inflammatory cell infiltration along with a profound increase in the expression levels of cytokines, MMP2 and MMP9 were observed in the infarcted and remote areas in Rnd3 +/- heart. Mechanistic studies revealed that Rnd3 bound to NFκB complex and attenuated NFκB activity by 1) physically interacting with NFκB component p65; 2) competing with NFκB component p50 for p65 binding; 3) blocking p65 nuclear import. The mechanisms were demonstrated in vitro and in vivo . Cardiomyocyte-specific overexpression of Rnd3 in mice inhibited hyperactivation of NFκB signaling, attenuated excessive inflammatory response, reduced infarct areas and ameliorated post-MI cardiac function. Conclusions: 1) This study demonstrates cardiomyocytes as an intrinsic and crucial component in the initiation of the inflammatory cascade in acute MI; 2) discover Rnd3 as a new “fine-tuning” factor situated at the nexus of the inflammatory response; 3) reveal a novel molecular mechanism through Rnd3-mediated NFκB signaling axes responsible for a balanced inflammatory response after acute MI.

scholarly journals Redox signaling modulates Rho activity and tissue contractility in the C. elegans spermatheca

2019 ◽  
Author(s):  
Charlotte A. Kelley ◽  
Sasha De Henau ◽  
Liam Bell ◽  
Tobias B. Dansen ◽  
Erin J. Cram
Keyword(s):  
Hydrogen Peroxide ◽  
Active Site ◽  
Myoepithelial Cells ◽  
Redox Signaling ◽  
Small Gtpase ◽  
C Elegans ◽  
Hydrogen Peroxide Treatment ◽  
Fine Tune ◽  
Contractile Tissue

AbstractActomyosin based contractility in smooth muscle and non-muscle cells is regulated by signaling through the small GTPase Rho and by calcium-activated pathways. We use the myoepithelial cells of the Caenorhabditis elegans spermatheca to study the mechanisms of coordinated myosin activation in vivo. Here, we demonstrate that redox signaling regulates RHO-1/Rho activity in this contractile tissue. Exogenous hydrogen peroxide treatment decreases spermathecal contractility by inhibiting RHO-1, which is mediated through a conserved cysteine in its active site (C20). Further, we identify a gradient of oxidation across the spermathecal tissue, which is regulated by the cytosolic superoxide dismutase, SOD-1. SOD-1 functions in the Rho pathway to inhibit RHO-1 through oxidation of C20. Our results suggest that SOD-1 functions to regulate the redox environment and to fine-tune Rho activity across the spermatheca.

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