Rajan, Malini et al. (2019) International Worm Meeting "NHR-14 loss of function couples intestinal iron uptake with innate immunity in C. elegans through PQM-1 signaling."

Romney, Steven, Leibold, Elizabeth, Jackson, Laurie, Anderson, Cole, Rindler, Paul, Gertz, Jason, Ferreira dos Santos, Maria, Rajan, Malini

[International Worm Meeting, 2019]

Iron is essential for survival of most organisms, but is toxic in excess. All organisms have thus developed mechanisms to sense, acquire and sequester iron. In C. elegans, iron uptake and sequestration are regulated by HIF-1. We previously showed that hif-1 loss of function mutants are developmentally delayed when grown under iron limitation. Here we identify nhr-14, encoding a nuclear receptor homologous to vertebrate HNF4, in a genetic screen conducted for mutations that rescue the developmental delay of hif-1 mutants under iron limitation. NHR-14 is highly expressed in intestinal cell nuclei and in head cells. We found NHR-14 subcellular localization and expression are not regulated by iron. Loss of nhr-14 leads to the upregulation of the intestinal metal transporter SMF-3 that increases iron uptake in hif-1 mutants, rescuing the low iron-dependent developmental delay. Loss of nhr-14 also promotes the nuclear localization of the zinc-finger transcription factor PQM-1, which activates smf-3 through the interaction with GATA-like DAF-16-associated elements (DAEs) in the smf-3 promoter. In addition to smf-3, RNA-seq analysis revealed upregulation of innate immune response genes as well as DAF-16/FoxO-suppressed Class 2 genes, which are known to be regulated by PQM-1. Consistent with the upregulation of innate immune response genes, nhr-14 mutants showed enhanced resistance to the human pathogen Pseudomonas aeruginosa that depends in part on the upregulation of smf-3 dependent iron uptake as well as the upregulation of innate immune response genes. P. aeruginosa reduces expression of nhr-14 in wild-type N2 worms, which is associated with PQM-1 nuclear localization. We propose that increased iron uptake by SMF-3 is a strategy to limit the acquisition of intestinal iron by pathogens, and may serve as a critical component of the host innate immune response. In summary, the identification of NHR-14 adds a new layer of regulation that links iron metabolism with the innate immune response and provides new insights by which C. elegans responds to pathogen infections.

Sorkac, Altar et al. (2011) International Worm Meeting "Notch signaling regulates synaptic transmission at the neuromuscular junction."

Singh, Komudi, Hart, Anne, Sorkac, Altar, DiIorio, Michael

[International Worm Meeting, 2011]

The role of Notch signaling during development has been characterized in many organisms, including the nematode C. elegans. In the canonical pathway, Notch receptors are cleaved by the g-secretase/presenilin complex upon ligand binding, releasing the intracellular domain that enters the nucleus to act as a transcription factor. In C. elegans, Notch receptors LIN-12 and GLP-1 are activated by DSL ligands working with DOS-motif proteins that act as co-activators. Here, we show that Notch signaling impacts synaptic transmission. Loss of two DOS co-ligands, OSM-7 and OSM-11, resulted in hypersensitivity to inhibitors of cholinesterase (Hic) on aldicarb. glp-1(gf) or glp-1(lf) had no impact on aldicarb response; whereas lin-12 null animals were Hic and lin-12(gf) mutants were Ric (Resistance to inhibitors of cholinesterase). These findings suggest a model in which OSM-7 and OSM-11 act on LIN-12 Notch receptors to regulate synaptic transmission at the neuromuscular junction.

Sorkac, Altar et al. (2013) International Worm Meeting "Notch signaling regulates synaptic transmission at the C. elegans neuromuscular junction."

Hart, Anne, Sorkac, Altar, Graham, Hannah, Singh, Komudi, DiIorio, Michael

[International Worm Meeting, 2013]

Notch is a signaling pathway that has been conserved across the animal kingdom. The role of Notch signaling during nervous system development has been extensively studied whereas its effects outside cell fate specification are less well described. Notch signaling is active in adult mice, Drosophila and C. elegans, regulating synaptic activity and other neurobiological processes. Notch receptors LIN-12 and GLP-1 are activated by DSL proteins with DOS-motif proteins acting as co-ligands. In C. elegans, Notch signaling affects avoidance of aversive chemicals and quiescence. Intriguingly, Notch DOS co-ligands OSM-7 and OSM-11 are regulated by environmental osmolarity; animals lacking either one of these co-ligands are preadapted to osmotic stress. Here, we show that Notch signaling impacts synaptic transmission at the neuromuscular junction of C. elegans, possibly in this neuroethological context. Complete loss of the Notch receptor LIN-12 results in hypersensitivity to aldicarb (Hic), an inhibitor of acetylcholinesterase. Loss the DSL ligand DSL-3 or either DOS co-ligand, OSM-7 or OSM-11, also results in hypersensitivity; these ligands likely activate LIN-12 in this paradigm. Notch signaling in adults is sufficient as overexpression of OSM-11 exclusively in adult animals causes resistance to aldicarb (Ric). Moreover, knock-down by RNAi of osm-11 in early larval stages does not cause adult aldicarb hypersensitivity. Also, osm-11 knock-down after the L2 larval stage results in hypersensitivity. Loss of the transcription factor LAG-1 resulted in hypersensitivity to aldicarb suggesting that LIN-12 signals via the canonical pathway. Next, the site of action of LIN-12 will be elucidated and candidate downstream targets will be examined in order to further our understanding of how Notch signaling regulates synaptic transmission.

Van Buskirk, Cheryl (2021) International Worm Meeting "Tips for early career scientists on NSF-CAREER award proposals"

Van Buskirk, Cheryl

[International Worm Meeting, 2021]

In keeping with the focus of this year's meeting on early career scientists, this poster aims to provide guidance and encouragement to new worm faculty in writing winning NSF Faculty Early Career Development (CAREER) proposals. The NSF-CAREER program is an exceptional funding opportunity for new investigators who are equally committed to outstanding research and education. The award is NSF-wide and thus is available to most areas of basic scientific research. If you are not yet tenured and looking to authentically integrate your research with education and/or outreach, this award is for you. Having served as a reviewer for these proposals, and currently in my last year of a CAREER award, I'd like to share some DOs and DONTs of proposal preparation and discuss ways to effectively integrate research and education/outreach components.

Lisa Girard et al. (2001) International C. elegans Meeting "Role of egl-5 in EGF/Wnt signal integration in the specification of P12 fate"

Scott Emmons, Paul Sternberg, Henrique B Ferreira, Lisa Girard

[International C. elegans Meeting, 2001]

The C. elegans Hox gene egl-5 is most similar, based on sequence analysis, to Abdominal-B . Consistent with its assignment into this paralog group, egl-5 is expressed in the posterior region of the worm. Immuno-staining results have shown that in the hermaphrodite, egl-5 is expressed in the hermaphrodite specific neuron, body wall muscle, posterior lateral microtubule neuron, PVC interneuron, the rectal epithelial cells K, F, B, U and the P12 neuroectoblast cell. 1 The two most posterior P cells are P11 and P12. The anterior products of their first division are both neuroblasts. P11.p fuses with the epidermal syncytium and P12.p divides again during late L1. The anterior division products, the epidermal cells P12.pa and P11.p are distinguishable by their distinct nuclear morphologies and positions. Previous genetic analysis indicates that P12 fate specification requires the synergistic action of the EGF and the Wnt signaling pathways. Reduction - or loss of function mutations in components of the EGF or the Wnt pathway result in partially penetrant P12 to P11 or P11 to P12 transformations. Double mutants of EGF and Wnt pathway components significantly enhance the frequency of transformation. P12 is not specified in an egl-5(lf) mutant and overexpression of egl-5 can rescue the loss of P12 specification phenotype of let-23 mutants. 2 In order to understand how information from the EGF and Wnt pathway are integrated at a cis -regulatory level, we have undertaken an analysis of the egl-5 promoter in collaboration with Scott Emmons to determine elements required for egl-5 expression in P12. 3 Do the two pathways converge on the egl-5 promoter or upstream of it? Beginning with a large promoter construct provided by Ferreira and Emmons, we have identified an approximately 1.3 kb. fragment of egl-5 promoter sufficient to drive expression of a heterologous promoter in P12. This 1.3 kb fragment contains six sites of approximately 20-40 bp each which are conserved between C. elegans and C. briggsae . Specific deletion of at least two of these sites in certain combinations eliminates expression in P12, suggesting that these sites might represent P12 enhancers. Complementary to our cis regulatory analysis, we have been using EMS screens to identify, and are currently characterizing, potential trans-acting factors as well as other genes which may participate in signal integration. Ferreira et al .(1999) Dev. Biol. 207:215-228 Jiang and Sternberg (1998) Development 125:2337-2347

Gautam Kao et al. (2006) European Worm Meeting "The ATPase ASNA-1 Positively Regulates Insulin Secretion in C. elegans and Mammals and Promotes Growth Non-autonomously"

Simon Tuck, Gautam Kao, Maria Still, Peter Naredia, Cecilia Nordensson, Agneta Rannlund

[European Worm Meeting, 2006]

Gautam Kao1, Cecilia Nordensson2, Maria Still2, Agneta Rnnlund1, Peter Naredi2 & Simon Tuck1. C. elegans embryos that hatch in the absence of food arrest growth during the first larval stage (L1). While much has been learned about the later diapause, called dauer that worms enter on encountering adverse conditions little is known presently about the mechanisms governing L1 arrest. We have identified and characterized a gene, asna-1, which when mutated, causes a complete but reversible arrest at the L1 stage even in the presence of food. We find that asna-1 encodes an ATPase that functions non-autonomously to regulate larval growth. asna-1 is expressed in a restricted set of sensory neurons and in the intestine; cells that produce insulin. asna-1 mutants have reduced insulin signaling while overexpression of asna-1 mimics the effects of overexpressing insulin. The human homologue ASNA1, which rescues the C. elegans mutant phenotype, is expressed at high levels in human pancreatic ?eta cells but not in other pancreatic endocrine or exocrine cell types. Increase or decrease of ASNA1 function in mouse insulinoma cells causes an increase or decrease in insulin secretion respectively. Similarly, reducing asna-1 gene function in C. elegans causes a decrease in levels of secreted insulin. We propose that ASNA1 is a novel evolutionarily conserved modulator of insulin signaling.

Marta Maria Gaglia et al. (2006) European Worm Meeting "Transcriptional Changes Underlying Chemosensory Regulation of Lifespan"

Cynthia Kenyon, Marta Maria Gaglia

[European Worm Meeting, 2006]

Marta Maria Gaglia and Cynthia Kenyon. Mutations in the insulin/IGF-1-like receptor daf-2 alone can double the lifespan of C.elegans, suggesting that aging is not just a default state, but instead a genetically regulated process. However, it is not clear what physiological processes regulate the daf-2 pathway. Interestingly, mutants with altered chemosensation also have an extended lifespan (Apfeld and Kenyon, 1999). Both daf-2 and chemosensory mutant longevity are dependent on the activity of the FOXO transcription factor DAF-16. This suggests that external inputs through the chemosensory system may regulate the insulin pathway and in turn affect lifespan. Further studies have identified some of the cellular components of the sensory pathway of lifespan control (Alcedo and Kenyon, 2004). However, the molecular mechanism by which the chemosensory system modulates lifespan remains still largely unknown. To begin to elucidate the mechanisms of chemosensory regulation of lifespan, we have performed genome-wide expression analysis with microarrays on the long-lived chemosensory mutant daf-10. We are in the process of characterizing the genes whose expression was altered with mutants and RNA interference in sensitized backgrounds. In particular, we are interested in those genes that affect lifespan without causing a chemosensory defect, as they may act downstream of the sensory neurons. In addition we are comparing the expression changes to those occurring in daf-2 mutants, to determine how the two pathways interact to modulate lifespan.

Frederick A. Partridge et al. (2006) European Worm Meeting "The Glycosyltransferase BUS-8 is Essential for Epidermal Morphogenesis"

Jonathan Hodgkin, Frederick A. Partridge, Adam W. Tearle, Maria J. Gravato-Nobre, William R. Schafer

[European Worm Meeting, 2006]

Frederick A. Partridge1, Adam W. Tearle2, Maria J. Gravato-Nobre1, William R. Schafer2 and Jonathan Hodgkin1. In C. elegans the epidermis is a simple epithelium. It is derived from a sheet of cells on the dorsal side of the embryo that spread to enclose the ventral side prior to elongation. The epidermis is then covered by the cuticle, an extracellular structure. We independently isolated alleles of bus-8 in screens for mutants resistant to infection by Microbacterium nematophilum and for nicotine hypersensitivity. Further alleles were obtained from a non-complementation screen, and a deletion (putative null) allele was provided by Shohei Mitani. bus-8(null) animals arrest during embryogenesis. We have shown that this is due to a defect in ventral enclosure, leading to extrusion of the body contents through the anterior of the ventral side during early elongation. This is reminiscent of mutations in the cadherin hmr-1. Weaker alleles of bus-8 show a variety of larval defects, including hypersensitivity to a variety of drugs. We have examined the structure of the cuticle and epidermis by electron microscopy. The epidermis is strikingly disordered. We also see abnormalities in the surface coat and cuticle. We propose that, due to their abnormalities in epidermal morphogenesis, bus-8 mutants are more permeable to small molecules. We have cloned bus-8. It encodes a putative glycosyltransferase. This suggests that glycosylation may have a role in mediating cadherin-based adhesion. We will discuss our progress at relating the molecular identity of bus-8 to its roles during development.

Singh K et al. (2010) Neuronal Development, Synaptic Function and Behavior, Madison, WI "Notch Signaling Regulates Lethargus Quiescence and Behavioral Adaptation to Stress"

Singh K, Hart A C, Corkins M E

[Neuronal Development, Synaptic Function and Behavior, Madison, WI, 2010]

A role for Notch signaling in adult nervous system function has been confirmed by several studies. Yet, how this pathway regulates behavior remains unclear. Originally, osm-11 was identified in classical screen for genes required for avoidance of octanol in our laboratory and avoidance of high osmolarity in the Horvitz and Thomas laboratories. osm-11 encodes one of five DOS family proteins that act with C. elegans DSL family ligands to activate Notch receptors in vulval development. We have delineated roles for osm-11 and/or Notch pathway genes in adult C. elegans for octanol avoidance and dispersal that are described elsewhere. In this presentation, we focus on the role of osm-11 and the Notch pathway in quiescence and behavioral adaptation to environmental stress. We noted that over-expression of osm-11 in adult C. elegans induced dramatic and anachronistic quiescence that was similar to the sleep-like quiescence of molting lethargus. A role for osm-11 and the Notch pathway in lethargus quiescence was defined during the L4/adult molt using a new microfluidic chip assay system (developed with the help of Shawn Lockery). Simultaneous loss of two DOS genes, osm-7 and osm-11, decreased quiescence during the L4 molt suggesting Notch signaling regulates this endogenous behavior. Examination of additional Notch pathway genes and epistasis studies with genes that regulate molting/quiescence reveals complex and interesting regulation of lethargus quiescence, which will be described at the meeting. Exposure to increased osmotic stress or loss of osm-11 causes internal glycerol accumulation and consequent increased physiological osmotic stress resistance (Osr, Strange, Lamitina & Thomas laboratories). Combining this with behavioral studies, we suggest that osm-11 and Notch signaling may play a pivotal role regulating behavioral and physiological adaptation to osmotic stress. Osmotic-adapted animals (those adapted to high osmotic stress) are behaviorally distinct from animals raised under our standard conditions and the known behavioral changes of osm-11(lf) animals are reminiscent of those observed in osmotic-adapted animals. For example, osm-11 and osmotic-adapted animals have low spontaneous reversal rates, which results in increased dispersal from high osmolarity microenvironments. Secreted osm-11 in the pseudocoelom may act as a hormone,- globally regulating Notch receptors in various tissues including neurons. Osmotic-adaptation can serve as a model to identify the mechanisms underlying nervous system adaptation to environmental and physiological stress. osm-11 and Notch signaling play a significant role in this adaptation, a role which may be conserved across species.

Mutlu, Beste et al. (2015) International Worm Meeting "Increasing histone H3K9 methylation and nucleosome stability promote higher-order nuclear re-organization during embryogenesis."

Mango, Susan, Mutlu, Beste, Wang, Jie, Chen, Huei-Mei, Hall, David, Liu, Tao

[International Worm Meeting, 2015]

Open chromatin is a conserved feature of pluripotent cells and is lost upon differentiation. As cells differentiate, the nucleus shrinks and chromatin undergoes global re-organization to generate domains of euchromatin and higher-order heterochromatin1,2. While much is known about chromatin organization at the local level, much less is known about the factors that govern large-scale nuclear organization and their regulation during embryogenesis. We have used ultrastructural and genomic approaches to track nuclear organization in C. elegans embryos as they transition from a developmentally plastic state towards differentiation. As embryos mature, higher-order heterochromatin emerges, visible as electron dense regions (EDRs) by electron microscopy. Concomitant with the appearance of EDRs, methylated histone H3 accumulates, and nucleosomes become more stable, as monitored by FAIRE (formaldehyde-assisted identification of regulatory elements)3. We find that the H3K9 methyltransferase MET-2 is required to build EDRs and promote loss of plasticity, but not to stabilize nucleosomes. These results highlight two apparently independent processes that generate a differentiated nucleus: i) generation of higher-order heterochromatin by MET-2, and ii) increased nucleosome stability. These changes may help focus the transcriptional machinery to discrete regions of open chromatin within differentiated cells.References1. Yuzyuk, T., Fakhouri, T.H.I., Kiefer, J., and Mango, S.E. (2009). The polycomb complex protein mes-2/E(z) promotes the transition from developmental plasticity to differentiation in C. elegans embryos. Dev. Cell 16, 699-710.2. Gaspar-Maia, A., Alajem, A., Meshorer, E. & Ramalho-Santos, M. Open chromatin in pluripotency and reprogramming. Nat. Rev. Mol. Cell Biol. 12, 36-47 (2011).3. Giresi, P.G., Kim, J., McDaniell, R.M., Iyer, V.R., and Lieb, J.D. (2007). FAIRE (Formaldehyde-Assisted Isolation of Regulatory Elements) isolates active regulatory elements from human chromatin. Genome Res. 17, 877-885.