Watteyne, Jan et al. (2021) International Worm Meeting "Untying the Gordian Knot: unravelling spatiotemporal activity in the C. elegans neuropeptide-receptor network"

Watteyne, Jan, Ripoll-Sanchez, Lidia, Vandewyer, Elke, Schafer, William R., Vertes, Petra E., Beets, Isabel, Cho, Sumin

[International Worm Meeting, 2021]

Neuropeptides are important modulators of organismal physiology and behavior. Yet, the rules and constraints by which these modulatory substances achieve both local and widespread physiological effects remain poorly understood. Currently lacking is a comprehensive analysis of the neuropeptide signaling network at an organismal systems level. The C. elegans genome shows a broad diversity of neuropeptide pathways, harboring around 150 genes encoding neuropeptides and a similar number of peptide GPCRs. Using reverse pharmacology, we have systematically mapped the biochemical network of neuropeptide-receptor interactions in the C. elegans nervous system. By screening for neuropeptide-GPCR couples, we identified receptors for all C. elegans RFamide-like peptides (FLPs) and many neuropeptide-like proteins (NLPs). These peptidergic pathways are organized into a dense signaling network including promiscuous neuropeptides and receptors. To further understand the functional organization of peptidergic circuits, we have adopted genetically-encoded sensors for neuropeptide-receptor activation that allow characterizing the spatiotemporal activity patterns of neuromodulatory signaling axes in the network. Using optogenetics, we found that conditional signaling of CAPA-1 neuropeptides, through activation of the neuromedin U receptor NMUR-1, underpins experience-dependent plasticity of salt chemotaxis behavior in C. elegans. CAPA-1 signaling from ASG neurons is specifically required for the retrieval, but not the acquisition, of learned salt avoidance. This highlights temporal aspects of neuropeptide signaling as important organizational motifs within the neuropeptide network, which we are further addressing with activity readouts of neuropeptide-receptor signaling. These findings and tools act as a scaffold to investigate how flexible behaviors emerge from neuromodulatory networks.

Ripoll-Sanchez, Lidia et al. (2021) International Worm Meeting "Mapping the neuropeptidergic connectome of Caenorhabditis elegans"

Vertes, Petra E., Miller III, David M., Schafer, William R., Watteyne, Jan, Ripoll-Sanchez, Lidia, Hobert, Oliver, Beets, Isabel, Weinreb, Alexis, Taylor, Seth R., Hammarlund, Marc

[International Worm Meeting, 2021]

The synaptically-wired neuronal circuitry is modulated by monoamines and neuropeptides, which act mostly through extrasynaptic volume transmission. This modulation is critical to nervous system function, yet little is known about the structure and function of extrasynaptic signaling networks at a whole-organism level. To this end, we used the recently published single neuron gene expression from the CeNGEN database along with deorphanization data for neuropeptide-activated G-protein coupled receptors (GPCRs) (see Jan Watteyne et al. abstract for this meeting) to generate a draft connectome of neuropeptide signaling networks in C. elegans. We based our network on single-cell neuronal expression patterns of 93 neuropeptide-receptor couples. In our baseline network edges were formed when the sending neuron expressed a given neuropeptide, the receiving neuron expressed the cognate receptor, and both neurons extended overlapping processes in the same neuropil. We also generated an unrestricted network with no spatial restriction on edge formation which allowed for potential long-range signaling. Since all 302 neurons of the adult hermaphrodite express at least one neuropeptide precursor gene and nearly all express at least one neuropeptide GPCR, both the baseline (with 31866 edges) and the unrestricted (with 54267 edges) neuropeptide networks were extremely dense compared to the synaptic connectome (with 2284 edges). In addition to its high density, the neuropeptide connectome differs in significant ways from the synaptic and monoamine signaling networks. For example, whereas the synaptic network consists of a small (11 neurons) core of high-degree hubs and a low-degree periphery, the neuropeptides network is more decentralised with a great number (103 neurons) of very high-degree nodes that form an interconnected rich club. Moreover, in contrast to the monoamine network, which shows very low reciprocity, the neuropeptide connectome shows higher than expected reciprocity, even though the networks formed by individual ligand-receptor couples are not. Finally, although the premotor neurons of the synaptic rich club have high neuropeptide degree, several of the most important nodes in the neuropeptide network are little-studied neurons that may be specialised for peptidergic neuromodulation. In the future, functional studies of these neurons and their role in behaviour may provide new insight into the control of behavioral states.

De Fruyt, Nathan et al. (2021) International Worm Meeting "Neuropeptidergic modulation of C. elegans learning behavior"

Fadda, Melissa, De Fruyt, Nathan, Schoofs, Liliane, Beets, Isabel, Watteyne, Jan, Van Damme, Sara

[International Worm Meeting, 2021]

Neuropeptides are an evolutionarily conserved group of neuromodulators that regulate a wide range of adaptive behaviors, such as learning. Yet, unravelling the molecular and circuit mechanisms underlying this neuropeptidergic modulation is challenging due to the diversity of neuropeptide signaling pathways, and their 'wireless' extrasynaptic mode of action. Using reverse pharmacology, we have constructed a molecular map of the C. elegans neuropeptide-receptor network. Phylogenetic reconstruction of the evolutionary history of nematode neuropeptide systems across bilaterian animals revealed several nematode-specific diversifications of neuropeptide signaling in addition to evolutionarily ancient neuropeptide pathways. One of these ancient, conserved pathways is a neuropeptide Y/F (NPY/F)-like signaling system that is an important regulator of learning behavior both in Proto- and Deuterostomia. We found that NPY/F-like FLP-34 neuropeptides are required in serotonergic neurons for aversive olfactory associative learning, which is functionally similar to the role of NPY in vertebrate learning as well as to the role of NPF in invertebrate learning. NPY/F-like neuropeptides are released from serotonergic neurons and signal through the G protein-coupled receptor NPR-11 in the excitatory AIA interneurons to facilitate olfactory aversive learning. In addition, signaling through NPY/F-like receptor NPR-11 also affects learning in salt gustatory plasticity, a gustatory associative learning paradigm. NPY/F-like signaling is not the only neuropeptidergic signaling system affecting learning behavior; we discovered additional neuropeptides that appear to be important to the learning process as well, including peptides that are expressed in non-neuronal cells. Our current research focuses on unravelling the functions of such non-neuronal neuropeptide messengers in learning and other types of behavioral plasticity.

Beets, Isabel et al. (2015) International Worm Meeting "Large-scale deorphanization of C. elegans neuropeptide receptors."

Caers, Jelle, Watteyne, Jan, Schoofs, Liliane, Peymen, Katleen, Van Sinay, Elien, Baytemur, Esra, Beets, Isabel

[International Worm Meeting, 2015]

Neuropeptides are key modulators of adaptive behaviors and represent one of the largest groups of neural messengers, with over 250 bioactive peptides predicted in C. elegans. Most of them are thought to act by binding of G protein-coupled receptors (GPCRs). Despite the broad diversity of neuropeptides and their physiological effects, a handful of neuropeptide-specific receptors have been characterized and the majority of peptide GPCRs remain orphan receptors, i.e. have no known ligand.We are therefore undertaking a large-scale deorphanization initiative - the Peptide-GPCR project - that aims to match all predicted peptide GPCRs of C. elegans to their cognate neuropeptide ligand(s). Using an in vitro reverse pharmacology approach, more than 150 peptide GPCR candidates are expressed in a heterologous cellular system and screened with a library of all known and predicted C. elegans peptides of the established NLP and FLP families. Activation of each receptor is measured by a calcium reporter read out. We have found several novel and evolutionary conserved neuropeptidergic systems including signaling pathways related to mammalian neuromedin, neuropeptide FF, and neuropeptide Y systems. Our results provide insight into neuropeptide signaling networks, and present a scaffold for further unravelling how neuropeptidergic states modulate adaptive behaviors and physiology. Peptide GPCRs are screened randomly, but community members are invited to help us prioritize candidates and steer the project's progression via http://worm.peptide-gpcr.org.

Wibisono, Phillip et al. (2021) International Worm Meeting "The neuropeptide receptor NMUR-1 regulates the specificity of C. elegans innate immunity against pathogen infection"

Chen, Chia-Hui, Watteyne, Jan, Sun, Jingru, Wibisono, Shawndra, Liu, Yiyong, Beets, Isabel, Sellegounder, Durai, Wibisono, Phillip

[International Worm Meeting, 2021]

Increasing evidence indicates that the innate immune system can generate high levels of specificity. However, the underlying molecular basis for such specificity is not well understood. Like other invertebrates, Caenorhabditis elegans does not have the adaptive immune system and only has innate immunity, and yet it can differentiate different pathogen attacks and launch proportionate innate immune responses. We found that functional loss of NMUR-1, a neuronal GPCR homologous to the mammalian receptors for the neuropeptide neuromedin U, had diverse effects on C. elegans survival against various bacterial pathogens. Further investigation revealed that NMUR-1 modulates C. elegans transcription activity by regulating the expression of transcription factors, which, in turn, controls the expression of distinct immune genes in response to different pathogens. Our study uncovered a molecular basis for the specificity of C. elegans innate immunity and could provide mechanistic insights into understanding the specificity of vertebrate innate immunity.

Armand I et al. (1996) Worm Breeder's Gazette "The Control of Body Size"

Emmons SW, Leroi AM, Armand I

[Worm Breeder's Gazette, 1996]

Exactly 100 years ago (Jan 10, 1896) Edwin Conklin published a paper entitled "Cell Size and Body Size." In it he posed the following question: do species that vary in adult body size differ in cell number or in cell size? His answer, for a genus of intertidal snails, was that species that vary in body size vary in cell number, but that cell size is more or less constant over evolutionary time.

Nelson GA et al. "Nematode radiobiology and development in space. Results from IML-1."

Richards DG, Benton ER, Kazarians GA, Schubert WW, Benton EV, Henke R, Nelson GA

The International Microgravity Laboratory #1 Spacelab mission was launched on 22-Jan-1992 for an 8-day mission. The Radiat experiment was one of 17 investigations which used the ESA Biorack on IML-1 and it had two objectives. The first objective was to isolate and characterize mutations induced by cosmic rays; the second was to assess the fidelity of development in 0-gravity over two consecutive generations. Two strategies were used to isolate mutations in a set of essential genes or a specific gene and to correlate the genetic events with the passage of charged particles. The results were isolation of 60 lethal mutations whose phenotypes are related to the local pattern of energy deposition. 12 mutations in the unc-22 gene include large deletions as characterized by DNA hybridization studies. Development of nematodes proceeded through two consecutive generations with no obvious defects. Cytoplasmic determinants in embryos, nuclear location and symmetry of cellular anatomy were normal as were Mendelian segregation and recombination of

Ellis HM et al. (1983) Worm Breeder's Gazette "monoclonal antibodies directed against the Drosophila nervous system cross-react with C elegans."

Yuan JY, Horvitz HR, Ellis HM

[Worm Breeder's Gazette, 1983]

We have begun to screen a collection of monoclonal antibodies that were generated against Drosophila embryos and stain a variety of Drosophila nervous tissues. These antibodies were kindly provided by Lily and Yuh Nung Jan of the University of California at San Francisco. Worms were fixed overnight at 4 C in 2% Formaldehyde, squashed, frozen and fractured as described by Albertson et al. (W.B.G. Vol.7 No.1) and transferred through a series of alchohols (95,70,50% EtOH). Three out of nine monoclonal antibodies that we have tested so far stain C. elegans. 20B6-B6* stains many nerve processes including dorsal and ventral cords, sub-lateral cords, cord commissures, amphid commissures and processes in the head. It is not yet clear whether this antibody stains all or only some processes. 18H12-C3* stains processes in a manner similar to 20B6-B6* as well as sperm and muscle ( or a structure with a muscle-like staining pattern). 21A4-E4 stains sperm and early (pre-morphogenetic) eggs. This staining is non- nuclear in both sperm and eggs and appears to be symmetrically distributed within eggs.

Schroeder, Nathan E et al. (2011) International Worm Meeting "Rapid and reversible arborization of IL2 neurons during dauer development."

Schroeder, Nathan E, Androwski, Rebecca J, Barr, Maureen M

[International Worm Meeting, 2011]

Dendrite morphology and plasticity profoundly affect neuronal signaling and behavioral outputs (1). However, little is known regarding the molecular signals governing morphogenesis of dendrite structure. During non-dauer stages of C. elegans development the inner labial (IL2) neurons, a set of six ciliated putative chemosensory neurons, display a bipolar morphology with an unbranched dendrite and axon. Under adverse environmental conditions, C. elegans can develop into a dauer larva, an alternative juvenile stage with altered morphology and behavior. We found that during the dauer stage, the IL2 neurons exhibit hierarchical dendritic branching and a switch from a bipolar to multipolar morphology. During dauer formation the ventral and dorsal IL2 primary dendrites establish branching and extend de novo processes from the cell bodies which undergo additional branching. The lateral IL2 neurons branch exclusively at the distal dendrites, forming a circular "crown" extending around the circumference of the head. Using time-lapse imaging, we found that plasticity in the IL2s begins with the onset of the dauer molt. Following the cessation of pharyngeal pumping puncta begin forming and resorbing in a dynamic fashion along the primary dendrite for several hours. Rapid and dynamic branch formation with periodic pruning events occur during a 3-4 hour period preceding radial shrinkage. Following recovery from dauer, the branches are incompletely resorbed, leaving behind occasional remnant secondary branches. Using a forward genetics screen, we isolated 28 candidate mutants with branching defects. Variations in defects include ectopic branches, disorganized branching or an incomplete crown. Several mutants were backcrossed and are being identified using traditional mapping and whole genome sequencing. We are currently using laser ablation to examine tiling and possible roles of surrounding tissue on IL2 branching. Additionally, we are testing candidate genes that may play a role in the IL2 dauer branching phenotype. Various developmental disorders are associated with defects in dendrite structure (1). IL2 branching in dauers may serve as a new and rapid model to understand the molecular basis of arborization and dendritic pruning that underlie these disorders. 1. Jan and Jan. 2010. Nat. Rev. Neurosci. 11:316-328.

Paulo Bettencourt et al. (2003) International Worm Meeting "Comprehensive genome wide RNAi screen for cell division genes in C. elegans"

Martin Jones, Joanne Stamford, Pierre Gonczy, Paulo Bettencourt, Anthony Hyman, Alan Coulson, Chris Echeverri, Steven Jones, Mark Hewitson, Karen Oegema

[International Worm Meeting, 2003]

To identify all genes required for cell division in a basic animal system we performed a genomic RNAi screen in C. elegans covering more than 99% of the genome as currently annotated. Aiming to identify genes in early development, we injected in vitro synthesized dsRNAs, and followed the first two cell divisions in the embryo at high resolution by DIC video microscopy, and the overall development of the progeny at a dissection microscope level. In addition, roughly 200 cell division genes were analysed by fluorescence video microscopy in GFP marker strains, giving further insight into the processes controlled by the genes addressed. The injection approach proved to be highly successful. Overall we scored RNAi phenotypes on 8% of all genes, 77% of which were defects in embryonic development. Compared to the Ahringer screen, published earlier this year (Kamath et al., Nature 421, 16 Jan 2003), we discovered about 30% more early genes, the silencing of which causes embryonic lethality. Judged by the nearly 100% hit rate on known genetic loci for the first cell division, we are confident we have identified and categorized a comprehensive set of cell division genes in C. elegans.