Michael Shapira et al. (2005) International Worm Meeting "Large scale transcriptional and functional analyses reveal central roles for a GATA transcription factor in Caenorhabditis elegans innate immune responses toward Pseudomonas aeruginosa infection."

Michal Ronen, Min Jiang, Stuart Kim, Jiming Rong, Brigham Hamlin, Michael Shapira, Man-Wah Tan, Marianne Campbell

[International Worm Meeting, 2005]

The opportunistic human pathogen Pseudomonas aeruginosa infects the worm Caenorhabditis elegans, when ingested, and causes its death within 3 days. We performed genome-wide gene expression analyses at early time points during P. aeruginosa infection, followed by a large scale RNAi-based functional analysis, to identify genes that respond to and protect from this infection. Transcriptional responses were distinct from general stress responses as previously described (e.g. Huffman et al., 2004), although some overlap exists, suggesting that the bulk of this response is a bona fide immune response and not a secondary effect of cellular damage or impaired metabolism. Induced genes included known as well as yet unknown, or putative, members of all levels of the innate immune system. Among the latter, genes encoding proteins with the yet uncharacterized DUF141 domain were noticeable in their importance for protecting worms from infection. We further unraveled the participation of the Sma/TGFbeta pathway and the intestinal GATA transcription factor, elt-2, in the regulation of the immune response. RT-PCR and functional analyses in RNAi treated worms showed that elt-2 is crucial for gene induction during infection, slowing pathogen accumulation in the intestine and improving survival. These results provide direct evidence for elt-2 being a pivotal nuclear regulator of immune responses in C. elegans. Additional evidence suggests that elt-2 may function downstream to the Sma/TGFbeta pathway.

Michael Meyer et al. (2008) Development & Evolution Meeting "NUD-1 Functions in Germline Development and Localizes Asymmetrically During Spermatogenesis"

Jennifer Miskowski, Kim Caldwell, Michael Large, Michael Meyer, Guy Caldwell

[Development & Evolution Meeting, 2008]

The C. elegans NUD-1 protein is the structural and functional ortholog of the fungal NudC protein (1). NUD-1/NudC proteins are evolutionarily conserved and have been implicated in diverse processes such as asymmetrical cell division in yeast, nuclear migration in fungi, and neuronal migration in humans; however, their exact function is unknown. When NUD-1 levels are depleted in N2 hermaphrodites by RNAi, a variety of phenotypes are found, including sterility (1), indicating a role for NUD-1 in C. elegans gonad development. Observation of nud-1(RNAi) mutants revealed possible defects in both the germ line and somatic gonad. To distinguish between these, NUD-1 RNAi was performed in rrf-1 mutants, which are resistant to RNAi in the soma. The treated animals were sterile and exhibited a wide variety of defects, suggesting that NUD-1 functions in the germ line. NUD-1 knockdown in males also results in gonad defects; sperm are generated, but their viability cannot be tested by mating due to severe tail malformations. An antibody generated against a phosphorylated version of mammalian NUDC cross-reacts specifically in C. elegans and is being used in immunofluorescence studies. Robust staining was observed in the germ line of both hermaphrodites and males. Specifically, NUD-1 was found in perinuclear puncta in primary spermatocytes with more diffuse staining in secondary spermatocytes and spermatids. NUD-1 was found to colocalize with fibrous body membrane organelles (FBMOs) which are sperm-specific organelles derived from the Golgi. Like FBMOs, NUD-1 is asymmetrically segregated to developing spermatids and away from residual bodies. Recent data suggests that the asymmetrical localization of FBMOs and NUD-1 is generated by the same molecular mechanism.

Hoy, Michael J. et al. (2015) International Worm Meeting "Worm farming: growing and harvesting large quantities of C. elegans."

Watson, Emma, Walhout, Albertha J.M., Hoy, Michael J.

[International Worm Meeting, 2015]

Mass spectrometry methods using C. elegans, such as metabolomics and protein identification, require the growth and collection of very large quantities of worms. In many cases, researchers are limited in their ability to analyze or collect low-abundance metabolites and proteins by the shear volume of worm biomass required for detection. We have optimized the procedure for growing millions of synchronized worms in liquid culture. The parent generation is grown on solid NGM media. Arrested L1 worms are grown to adulthood by the millions in liquid NGM supplemented with concentrated E. coli OP50 as a food source. These wild-type worms grow at the same rate as worms on solid media. We employ the use of Imhoff settling cones for the process of quickly washing and collecting the worms from liquid culture. These samples can then be frozen and stored at -80oC for future protein and metabolite extraction and analysis by mass spectrometry. This method provides researchers with a new, streamlined approach to the arduous task of performing experiments that require millions of worms and milliliter quantities of worm biomass.

Scheel JK et al. (1995) International C. elegans Meeting "SYNAPSE SPECIFICATION IN C. ELEGANS"

Hedgecock EM, Scheel JK

[International C. elegans Meeting, 1995]

We are interested in the molecular basis of synaptic specificity. Neurexins, presynaptic membrane proteins with large extracellular domains containing regions homologous to laminin G domains, are believed to be involved in cell-cell interactions at mammalian synapses. In mammals, there is a large variety of neurexin proteins created by a combination of alternative splicing and multiple genes.

Michael Hoffmann et al. (2006) European Worm Meeting "The Planar Cell Polarity Protein VANG-1 interferes with Intercalation Events during Epithelial Morphogenesis In C. elegans"

Michael Hoffmann, Christoph Segbert, Olaf Bossinger, Gisela Helbig

[European Worm Meeting, 2006]

Michael Hoffmann, Christoph Segbert, Gisela Helbig and Olaf Bossinger. C. elegans VANG-1 is the only homolog of the Drosophila planar cell polarity (PCP) protein Strabismus/Van Gogh. We originally identified VANG-1 as a putative binding partner of DLG-1 (Discs large), a MAGUK protein that acts as a molecular scaffold for the establishment of the apical junction in all epithelia of the C. elegans embryo. The phenotype observed in vang-1 mutant embryos show intercalation defects during epithelial morphogenesis of the intestine and the hypodermis. Similar phenotypes arise in lin-17 (frizzled) mutants or after RNAi against dsh-2 (dishevelled), indicating core components of the PCP pathway in other systems to be involved in cell intercalation in the C. elegans embryo. In addition, VANG-1 and DSH-2 do colocalize and the asymmetric distribution of VANG-1 in epithelia depends on DSH-2. Taken together these results suggests that planar cell polarity might exist in C. elegans.

Zhao-Wen Wang et al. (2003) International Worm Meeting "Presynaptic Ryanodine Receptors Mediate Spontaneous Neurotransmitter Release at the C. elegans Neuromuscular Junction"

Lawrence Salkoff, Ding-Xia Shen, Zhao-Wen Wang, Michael L Nonet

[International Worm Meeting, 2003]

Spontaneous miniature postsynaptic potentials or currents (minis) are generally considered fusion events of individual synaptic vesicles. Consistent with this notion, minis at vertebrate neuromuscular junctions (NMJs) often appear unitary, and show a normal distribution in amplitude histograms. However, at many other synapses, such as synapses in the vertebrate central nervous system, amplitude histograms of minis are positively skewed due to the presence of numerous large-amplitude events. Little is known about the origin of large-amplitude spontaneous events. At the C. elegans NMJ, amplitudes of minis vary greatly. Although their average amplitude is about 25 pA, many are over 50 pA. We investigated the mechanism of large-amplitude minis at the C. elegans NMJ. Minis were recorded from body-wall muscle cells voltage-clamped at -60 mV. Large-amplitude minis persisted in the absence of external Ca2+, suggesting that they are not endogenous evoked responses. Experiments with nicotinic and GABAergic receptor blockers, and with mutants of GABAergic receptors (unc-49) suggest that large-amplitude minis are of both cholinergic and GABAergic origins. Large-amplitude minis were absent in unc-68(r1162), a null mutant of the ryanodine receptor (RyR). Inclusion of a RyR blocker in the recording pipette to selectively block postsynaptic RyRs had no effect on minis, whereas application of a membrane permeant RyR blocker to the NMJ preparation, which blocked both presynaptic and postsynaptic RyRs, produced effects similar to that of the unc-68 mutant, suggesting that large-amplitude minis are mediated by presynaptic RyRs. The frequency of minis was also severely reduced in the RyR mutant and in wild-type animals treated with a RyR blocker. The effect of RyRs appeared specific because large-amplitude minis remained in other mutants affecting intracellular Ca2+ mobilization (itr-1 and crt-1), or release of large dense core vesicles (unc-31). In the presence of extracellular Ca2+ (1-5 mM), mini frequency was high (~40 Hz). Elimination of external Ca2+ reduced mini frequency by ~80%. Preliminary results suggest that the effect of external Ca2+ on minis is mediated at least in part by presynaptic RyRs, perhaps through a process known as Ca2+-induced Ca2+ release.

Vecseri G et al. (1991) International C. elegans Meeting "AN UNUSUAL MUTANT STRAIN OF CAENORAHBDITIS BRIGGSAE (G16)."

Buza J, Fodor A, Vecseri G

[International C. elegans Meeting, 1991]

The founder animal of strain vg-182 was isolated as a Large Roller progeny of an EMS treated C. briggsae (G16) hermaphrodite. It has segregated only a few offsprings including Lons and animals of variable phenotypes. Lon hermaphrodites were singled and self-fertilized through subsequent generations and then crossed out several times. During this process 7 of 8 fertile lines segregated Lon males, Large Dpy hermaphrodites, Small Dumpies and Large Rollers. At the end, loci represented by one X-linked dominant lon, one autosomal recessive dpy and one independent, autosomal squat allele, respectively, could be identified and named temporarily as lon-2, dpy- 2 and sqt-l. lon-2(d) proved a dominant epistatic gene over dpy-2 and a semidominant epistatic one over sqt-l. (Worms of sqt-l; +/+; lon-2 genotype exhibited Large Roller phenotype while those of sqt-l/+; lon- 2/0 genotype were large, Non-Roller males.) The male--segregating (Him) phenotype could not be separated from lon-2(d). The lon-2 hermaphrodites kept in liquid nitrogene still keep their 'him' phenotype, those which were kept in laboratory conditions lost their male-segregating ability. The possibilities of getting a mutator strain of C. briggsae are discussed.

Michael Hoffmann et al. (2005) International Worm Meeting "Analysis of the DLG-1 (Discs large) binding partners Vang-1 (Strabismus/Van Gogh) and EGL-15 during organogenesis of the intestine"

Michael Hoffmann, Olaf Bossinger

[International Worm Meeting, 2005]

The C. elegans homolog of the Drosophila tumor suppressor protein Discs large, DLG-1, acts as a molecular scaffold for the establishment of the apical junction. We identified the homolog of the planar cell polarity protein Strabismus/Van Gogh, Vang-1, and a homolog of the FGF receptor tyrosine kinase Breathless, EGL-15 as interacting partners of DLG-1. EGL-15 is able to phosphorylate Vang-1 in vitro and the subcellular distribution of Vang-1 is altered in an egl-15 background. The phenotypes observed in both, vang-1 and egl-15 mutants suggest defects in spindle positioning and/or intercalation of specific E-cells during morphogenesis of the gut epithelium. In addition, disruption of the non-canonical wingless signal transduction pathway in mutants or RNAi against beta-catenin (wrm-1), dishevelled (dsh-2) and frizzled (lin-17) exhibit similar phenotypes and are likely candidates to be involved in gut organogenesis. Furthermore, preliminary data indicate that src-1(RNAi) embryos show a vang-1-like phenotype, suggesting the involvement of the integrin signalling pathway during the process of gut morphogenesis.

Alok, G. et al. (2019) International Worm Meeting "Developing a large-scale method for bulk segregant analysis of genetic variation underlying differential requirements for SKN-1 in endoderm specification."

Torres Cleuren, Y., Alok, G.

[International Worm Meeting, 2019]

The maternally supplied SKN-1 transcription factor initiates specification of the C. elegans endoderm. We have found that the requirement for SKN-1 in endoderm specification varies widely between genetically distinct isolates of the species; while some wild isolates show a nearly absolute requirement for this factor, the majority of embryos in others generate endoderm in its absence. GWAS and analysis of RILs allowed us to identify several regions of the genome responsible for this large variation in the requirement for SKN-1. To narrow in on the specific causal sequence differences responsible for this variation, we are developing a method for large-scale bulk segregant analysis by evaluating large numbers of embryos derived from many-generation bulk intercrosses. This approach relies on our ability to sort large numbers of individual embryos with a fluorescent-activated cell sorter that effectively separates embryos that undergo endoderm differentiation from those that do not. With this approach we can analyze thousands of recombinant embryos and hope to achieve fine-scale resolution mapping of causal loci responsible for evolutionary variation in the endoderm gene regulatory network.

Spooner, Patrick et al. (2011) International Worm Meeting "Immunoglobulin domain containing isoforms of UNC-89 (obscurin) are required for myofilament organization and calcium signaling in Caenorhabditis elegans."

Duran, M. Berenice, Spooner, Patrick, Benian, Guy, Bonner, Jennifer, Norman, Kenneth

[International Worm Meeting, 2011]

The force generating machinery in muscle is regulated by the influx of calcium ions into the muscle cytoplasm. Initially, depolarization of the sarcolemma leads to calcium entry via voltage gated calcium channels, which rapidly triggers the release of calcium from the sarcoplasmic reticulum via ryanodine receptors by calcium induced calcium release. Thus, calcium needs to be rapidly, accurately and reliably regulated and the calcium channels and contractile machinery must be maintained in a highly ordered arrangement for efficient and effective muscle contraction to occur. The mechanism underlying this highly organized configuration is not fully understood. Using a combination of genetic and cell biological techniques, we have found that the C. elegans ortholog of the giant muscle protein obscurin, UNC-89, is required for muscle cell organization. The unc-89 locus encodes at least 6 isoforms as large as 900 kDa, 4 large Immunoglobin (Ig) domain rich isoforms and two smaller tandem kinase containing isoforms (Small et al. 2004; Ferrara et al. 2005). From our analyses, we have found that large Ig domain rich isoforms of UNC-89 are critical for sarcomere and SR organization. Furthermore, we have found evidence that unc-89 mutants lacking the large Ig rich isoforms have defects in excitation-contraction coupling in the pharyngeal and body wall muscle, through the coordination of calcium influx. Thus, our data implicates the large Ig domain rich isoforms of UNC-89 in maintaining muscle cell architecture that is critical for efficient and effective muscle contraction.