Min DS et al. (2000) J Neurochem "Phospholipase C, protein kinase C, Ca(2+)/calmodulin-dependent protein kinase II, and tyrosine phosphorylation are involved in carbachol-induced phospholipase D activation in Chinese hamster ovary cells expressing muscarinic acetylcholine receptor of Caenorhabditis elegans."

Jo YH, Yoon SH, Hahn SJ, Lee YH, Kim MS, Cho NJ, Lee KH, Min DS

[J Neurochem, 2000]

Recently, we have isolated a cDNA encoding a muscarinic acetylcholine receptor (mAChR) from Caenorhabditis elegans. To investigate the regulation of phospholipase D (PLD) signaling via a muscarinic receptor, we generated stable transfected Chinese hamster ovary (CHO) cells that overexpress the mAChR of C. elegans (CHO-GAR-3). Carbachol (CCh) induced inositol phosphate formation and a significantly higher Ca(2+) elevation and stimulated PLD activity through the mAChR; this was insensitive to pertussis toxin, but its activity was abolished by the phospholipase C (PLC) inhibitor U73122. Western blot analysis revealed several apparent tyrosine-phosphorylated protein bands after CCh treatment. The CCh-induced PLD activation and tyrosine phosphorylation were significantly reduced by the protein kinase C (PKC) inhibitor calphostin C and down-regulation of PKC and the tyrosine kinase inhibitor genistein. Moreover, the Ca(2+)-calmodulin-dependent protein kinase II (CaM kinase II) inhibitor KN62, in addition to chelation of extracellular or intracellular Ca(2+) by EGTA and BAPTA/AM, abolished CCh-induced PLD activation and protein tyrosine phosphorylation. Taken together, these results suggest that the PLC/PKC-PLD pathway and the CaM kinase II/tyrosine kinase-PLD pathway are involved in the activation of PLD through mAChRs of C. elegans.

Chew, Yee Lian et al. (2015) International Worm Meeting "Neuronal protein with tau-like repeats (PTL-1) regulates intestinal SKN-1 nuclear accumulation in response to oxidative stress."

Gotz, Jurgen, Chew, Yee Lian, Nicholas, Hannah

[International Worm Meeting, 2015]

Oxidative stress is a central pathomechanism in Alzheimer's disease and other diseases with tau pathology. In response to oxidative stress, the Nrf2 transcription factor induces the expression of detoxification enzymes, and recent data indicate that Nrf2 activity improves tau pathology and tau-mediated neurotoxicity (Jo et al., 2014, Nature Communications; Stack et al., 2014, Human Molecular Genetics). The homolog of Nrf2 in the model organism Caenorhabditis elegans, called SKN-1, regulates a conserved stress response by mediating the expression of detoxification enzymes in stress conditions. In response to stressors, intestinal SKN-1 localises to the nucleus to modulate gene expression. We previously showed that the sole homolog of tau/MAP2 in C. elegans, Protein with Tau-like repeats (PTL-1), regulates neuronal ageing and lifespan. Here, we tested for the involvement of PTL-1 in stress tolerance. Our findings indicate that ptl-1 mutant animals are hypersensitive to oxidative stress and are defective in nuclear accumulation of SKN-1 in stress conditions. This defect can be rescued by PTL-1 re-expression under the control of the endogenous promoter. Given the close relationship between ageing and stress tolerance, we next tested lifespan in ptl-1 and skn-1 mutant animals and found that ptl-1;skn-1 double mutants are not significantly shorter- or longer-lived than single mutants, suggesting that that both genes regulate longevity via similar processes. We also tested for intestinal SKN-1 nuclear accumulation in a transgenic line re-expressing PTL-1 in the nervous system. Data from these experiments indicate that SKN-1 in the intestine is modulated by PTL-1 in neurons. Together, this demonstrates that PTL-1 functions via neurons to modulate intestinal SKN-1, clarifying the role of PTL-1 in the stress response and longevity.

Rachael A Nimmo et al. (2003) International Worm Meeting "Worm Tales! Molecular Genetic Analysis of Male Tail Development in C. elegans."

Rachael A Nimmo, Alison Woollard, Adam Antebi

[International Worm Meeting, 2003]

The male tail of C. elegans is a specialised structure consisting of a cuticular fan in which are embedded 18 symmetrically placed mechanosensory sensilla called rays, which each have distinct characteristics due to their morphology, position and neurotransmitter usage. The mab-2 mutation causes loss of both the V and T cell derived rays1. We have found this mutation to be allelic with a rnt-1 knockout and to be rescued by the cosmid B0414 which contains rnt-1. We will further analyse the ray phenotype by lineage analysis using suitable markers. In addition, mutations appear to be temperature sensitive for sterility and an intestinal mitotic defect. It is interesting that a rnt-1::gfp reporter construct has been shown previously to be expressed in the intestine as well as seam cells2 so lending support to the hypothesis that both the ray loss and intestinal phenotypes may be caused by mutation of rnt-1.Initial attempts at cloning mab-2 involved screening genes mapping to the same region by feeding RNAi (Chromosome I feeding library was a gift from J. Ahringer). As a result of the screen two genes which phenocopied mab-2 mutants were identified. In light of our subsequent identification of mab-2, it is interesting to find that 3 genes giving a virtually identical ray loss phenotype are located within a 0.21 cM region of LGI.The two novel Mab genes are situated in a putative operon and so are presumably co-regulated. Do worm operons represent functional cassettes of genes as in bacteria? The common RNAi phenotypes of these two co-regulated genes imply interesting co-functionality. Further support for their co-functionality is provided by the fact that homologues of these two genes plus another in the operon are involved in rRNA processing in yeast. In addition, orthologues of one of these Mab genes and another gene in the operon are thought to modulate the Notch signalling pathway in mammals. We are in the process of investigating the possibility that these functions are conserved in C. elegans.References: 1) Hodgkin, J. Genetics (1983) 103: 43-64; 2) Nam S, Jin YH, Li QL, Lee KY, Jeong GB, Ito Y, Lee J, Bae SC. Mol Cell Biol. (2002) 22(2):547-54.

Chien, Shih-Chieh et al. (2009) International Worm Meeting "Do orthologs of the yeast RAM pathway mediate Wnt signaling in neuronal polarity?"

Garriga, Gian, Chien, Shih-Chieh, Oppermann, Julie, Gurling, Mark

[International Worm Meeting, 2009]

Wnts regulate cell migration and polarity along the C. elegans A/P axis. For example, Wnts control the polarity of the mechanosensory neuron ALM1,2. Although single cwn-1, cwn-2 or egl-20 Wnt mutants display normal ALM polarity, the polarity of the ALMs is often reversed in cwn-1; cwn-2 or cwn-1; egl-20 double mutants. We find that ALM polarity also requires the Frizzled receptor MOM-5 and the Dishevelleds DSH-1 and MIG-5. While Frizzled and Dishevelled proteins mediate the effects of Wnts in many developmental contexts, how these molecules signal to control neuronal polarity is unclear. We find that the MIG-15 kinase and potential components of a MIG-15 signaling pathway might be novel Wnt effectors in neuronal polarity. MIG-15, the C. elegans ortholog of Nck-interacting kinase (NIK) in mice and Misshapen in Drosophila, was shown to function in cell migrations that also require Wnt function3. We found that mig-15 mutants exhibit a low frequency of ALM polarity defects that was enhanced by a mutation in cwn-1, suggesting that MIG-15 could mediate the effects of Wnts in ALM polarity. In S. cerevisiae, Kic1p, a distant relative of MIG-15, acts in the RAM signaling pathway to regulate polarized cell growth4. We asked whether C. elegans orthologs of RAM signaling molecules also regulate ALM polarity. Through RNAi and mutant analysis in a cwn-1 sensitized background, we identified mop-25.2 and sax-2 as regulators of ALM polarity. The gene mop-25.2 is the ortholog of yeast Hym1p, which physically interacts with Kic1p, and SAX-2 is the ortholog of the RAM scaffold protein Tao3p. SAX-2 acts in the ALM to establish its polarity. We were surprised to find that cwn-1; sax-1 mutants did not have an ALM polarity phenotype. SAX-1 is the ortholog of the NDR kinase Cbk1p and a target of the Kic1p kinase. We are currently testing whether the other C. elegans NDR kinase WTS-1 provides an overlapping function in ALM polarity in the absence of SAX-1. We are now testing whether the RAM pathway homologs act downstream of the Wnts in ALM polarity. 1Hilliard MA, Bargmann CI. (2006) Dev. Cell 10: 379-390. 2Prasad BC, Clark SG. (2006) Development 133: 1757-1766. 3Chapman JO, Li, H & Lundquist EA. (2008) Dev. Biol. 324: 245-257. 4Nelson B et al. (2003) Mol. Biol. Cell 14: 3782-3803.