Barry, James et al. (2011) International Worm Meeting "EMS Mutagenesis of Early Embryos: Does checkpoint status affect the mutational spectrum?"

Barry, James, Hartman, Phil, Williams, Dean, Finstad, Whitney, Khan, Numan

[International Worm Meeting, 2011]

Mutagenesis protocols typically call for exposure of late-stage larvae or adults to a mutagen with the hope of inducing mutations in a robust germ line. If recessive, these mutations manifest themselves in the F2 generation. In contrast, we have handpicked and EMS-mutagenized early (1-4 cell) C. elegans embryos. Because all meiotic products derive from a single progenitor cell at this stage, homozygotes should be obtained in the F1 rather than the F2 generation. Using CB665 [unc-58(e665)], a strain in which secondary mutations can relieve a strong Unc phenotype, we hand-selected ca. 16,000 early embryos and exposed them to EMS for 50 minutes. We recovered 21 mutants out of ca. 6,400 viable embryos, which calculates to a mutation frequency of 3x10-3. This is considerably higher than the 10-4 to 5x10-4 typical of classical EMS mutagenesis (Anderson, P. 1995. In: Methods in Cell Biology, Vol. 48, Epstein and Shakes, eds, p. 31). This is not surprising given that cell-cycle checkpoints are muted in C. elegans embryos (Holway et al., 2006. J. Cell Biol. 172:999). Of the 21 mutants, 17 are intragenic. We have also isolated 26 EMS-induced e665 revertants using standard EMS mutagenesis, of which 25 are intragenic. We are currently sequencing the unc-58 gene in both sets of mutants to see if the mutational spectra differ from one another.

Takahashi, Kodai et al. (2015) International Worm Meeting "Oxidative stress response of a transcription factor MXL-3 in the nematode C. elegans."

Ishii, Naoaki, Yasuda, Kayo, Sasagawa, Noboru, Hartman, Phil, Takahashi, Kodai, Ishii, Takamasa

[International Worm Meeting, 2015]

It has been reported that a basic helix-loop-helix transcription factor MXL-3 (Max-like 3) represses the expression of the lysosomal lipases in the presence of nutrients. On the other hand, under starvation conditions, MXL-3 is down-regulated and the expression of the lysosomal lipases are induced. Consequently, fats are broken down by inducing lipophagy. Moreover, it has been reported that the transcription factor SKN-1, a homolog of mammalian Nrf proteins, binds to MXL-3 which is involved in oxidative stress response. These studies suggest that MXL-3 may be involved oxidative stress response. We have previously shown that X-ray irradiation extends lifespan and up-regulates the expression of mxl-3 at a dauer larval stage of the nematode Caenorhabditis elegans. It is well known that the radiation generates reactive oxygen species in response to water in vivo. To investigate the role of MXL-3 in oxidative stress response, we examined, 1) mxl-3 expression levels in wild-type animals exposed to paraquat, 2) the lifespan of a mxl-3 (ok1947) mutant under high oxygen levels, 3) paraquat sensitivity of the mxl-3 mutant. Our results showed that mxl-3 mRNA levels was up-regulated in wild type worms exposed to 30mM paraquat for 4 hours and longer. The mxl-3 mutant was hypersensitive to paraquat. Moreover, under 90% oxygen, the mean lifespan of the mxl-3 mutant was relatively more decreased in comparison to wild type. It has been reported that mxl-3 mutant is long lived in abundant food conditions. On the other hand, we observed mxl-3 mutant is short lived under high oxidative stress. In addition, mxl-3 was up-regulated in wild type animals. Our result raise the possibility that MXL-3 is not involved in response to low oxidative stress such as normoxia s but in response to hyperoxia.

Nguyen, Anh Quynh et al. (2009) International Worm Meeting "THE GENETICS OF DEET RESISTANCE IN Caenorhabditis elegans."

Limes, Julia, Freedman, Matt, Douglas, Alfred, Nguyen, Anh Quynh, Hartman, Phil, Copeland, Heather

[International Worm Meeting, 2009]

The molecular mechanism(s) by which DEET (N,N-diethyl-m-toluamide) acts as an insect "repellent" has not been resolved. Using forward genetic screens in Caenorhabditis elegans, we have isolated five DEET-resistant mutants (der-1) after EMS mutagenesis of DEET-sensitive N2. Two are allelic and have been mapped between 2.4 mu and 2.9 mu on linkage group (LG) IV. Specifically, 3F (factor) cross data between two markers unc-5 (1.78 mu) and egl-19 (3.34 mu) on LGIV showed that 11/22 (50%) Egl-19 recombinants were DEET-resistant and 11/22 (50%) were DEET sensitive for allele hf175. In case of allele hf176, the data were 19/32 (59%) and 13/32 (41%), respectively. In addition, all Lin-33 recombinants from the 3F cross between two references unc-5 (1.78 mu) and lin-33 (2.55 mu) showed DEET sensitivity in both hf175 and hf176. Several additional 3F crosses are in various stages of completion. Interestingly, wild-type strains CB4856, AB1, and AB2 have been shown to be DEET resistant relative to the wild-type strains N2 and CB4852, which are both DEET sensitive. These findings suggest that DEET-resistance may be more common than we originally expected. Successful cloning of the der-1 gene is an important step not only to elucidate the mechanism of DEET action but may prove useful in designing the next generation of chemicals that helps reducing insect-borne-diseases.

Morgan PG et al. (2000) Worm Breeder's Gazette "Mutations in two C. elegans respiratory enzymes confer variable hypersensitivity to free radicals and volatile anestetics"

Kayser EB, Sedensky MM, Ishii N, Hartman PS, Morgan PG

[Worm Breeder's Gazette, 2000]

As with mev-1, a mutation in gas-1 resulted in hypersensitivity to the superoxide anion-generating chemical paraquat. gas-1 was more hypersenstive at 20C than 15C, which was not surprising given that gas-1 is difficult to even propagate at 25oC. As with mev-1, the gas-1 mutation also rendered animals hypersensitive to hyperoxia. In addition, the life spans of both gas-1 and mev-1 mutants were significantly shortened by elevated oxygen concentrations. It was determined previously that mev-1 animals were hypermutable to oxygen (Ponder, Hartman and Ishii, unpublished data). Conversely, gas-1 was only weakly hypermutable if at all. Finally, kn1 mutation in mev-1 did not confer hypersensitivity to the volatile anaestetic halothane.

Ishii N et al. (1998) Nature "A mutation in succinate dehydrogenase cytochrome b causes oxidative stress and ageing in nematodes."

Yanase S, Yasuda K, Hartman PS, Tsuda M, Ishii N, Fujii M, Suzuki K, Senoo-Matsuda N, Ayusawa D

[Nature, 1998]

Much attention has focused on the aetiology of oxidative damage in cellular and organismal ageing. Especially toxic are the reactive oxygen byproducts of respiration and other biological processes. A mev-1(kn1) mutant of Caenorhabditis elegans has been found to be hypersensitive to raised oxygen concentrations. Unlike the wild type, its lifespan decreases dramatically as oxygen concentrations are increased from 1 to 60%. Strains bearing this mutation accumulate markers of ageing (such as fluorescent materials and protein carbonyls) faster than the wild type. We show here that mev-1 encodes a subunit of the enzyme succinate dehydrogenase cytochrome b, which is a component of complex II of the mitochondrial electron transport chain. We found that the ability of complex II to catalyse electron transport from succinate to ubiquinone is compromised in mev-1 animals. This may cause an indirect increase in superoxide levels, which in turn leads to oxygen hypersensitivity and premature ageing. Our results indicate that mev-1 governs the rate of ageing by modulating the cellular response to oxidative stress.AD - Department of Molecular Life Science, Tokai University School of Medicine, Isehara, Kanagawa, Japan. nishii@is.icc.u-tokai.ac.jpFAU - Ishii, NAU - Ishii NFAU - Fujii, MAU - Fujii MFAU - Hartman, P SAU - Hartman PSFAU - Tsuda, MAU - Tsuda MFAU - Yasuda, KAU - Yasuda KFAU - Senoo-Matsuda, NAU - Senoo-Matsuda NFAU - Yanase, SAU - Yanase SFAU - Ayusawa, DAU - Ayusawa DFAU - Suzuki, KAU - Suzuki KLA - engSI - GENBANK/H83619SI - GENBANK/L26545SI - GENBANK/N34060SI - GENBANK/R87065SI - GENBANK/S74803PT - Journal ArticleCY - ENGLANDTA - NatureJID - 0410462RN - 1339-63-5 (Ubiquinone)RN - 37337-51-2 (cytochrome b560)RN - 9035-37-4 (Cytochrome b)RN - EC 1.- (Succinate Cytochrome c Oxidoreductase)SB - IM

Larson SD et al. (2018) Philos Trans R Soc Lond B Biol Sci "Connectome to behaviour: modelling Caenorhabditis elegans at cellular resolution."

Gleeson P, Larson SD, Brown AEX

[Philos Trans R Soc Lond B Biol Sci, 2018]

It has been 30 years since the 'mind of the worm' was published in <i>Philosophical Transactions B</i> (White <i>et al</i> 1986 <i>Phil. Trans. R. Soc. Lond. B</i><b>314</b>, 1-340). Predicting <i>Caenorhabditis elegans</i>' behaviour from its wiring diagram has been an enduring challenge since then. This special theme issue of <i>Philosophical Transactions B</i> combines research from neuroscientists, physicists, mathematicians and engineers to discuss advances in neural activity imaging, behaviour quantification and multiscale simulations, and how they are bringing the goal of whole-animal modelling at cellular resolution within reach.This article is part of a discussion meeting issue 'Connectome to behaviour: modelling <i>C. elegans</i> at cellular resolution'.

Oge Arum et al. (2004) West Coast Worm Meeting "Reduction in cep-1 expression increases lifespan"

Oge Arum, Tom Johnson

[West Coast Worm Meeting, 2004]

We found that reduced expression of cep-1 (a homolog of p53) confers extension of maximal lifespan in screens of RNAi libraries. Subsequent survival assays confirmed a modest mean lifespan increase and mild resistance to heat, but no effect on resistance to oxidative, stress. cep-1 feeding-RNAi failed to increase dauer formation of N2 worms at 27 degC or of daf-2 worms at 25.5 degC. cep-1 feeding-RNAi also failed to induce nuclear-localization of DAF-16::GFP; all suggesting that CEP-1 does not participate in the daf-2 pathway. However, the daf-16 -dependence lifespan assays showed daf-16 worms on cep-1 feeding-RNAi bacteria as having lifespans similar to those of daf-16 worms on GFP feeding-RNAi bacteria. As we have often pointed out, shortening of life span by supposed 'suppression' by daf-16 mutations is not a reliable indication of direct interaction between the genes under study because of the known life-shortening effects of mutations in daf-16 and the oxidative toxicity in daf-16 hypomorphs. A cep-1 knockout has a lifespan increase slightly longer than that observed on cep-1 RNAi. A survey of all C. elegans mutants defective for response to DNA damage in the germline revealed that only cep-1 reduction causes an increased life expectancy. Thus, there does not appear to be a universal correlate of DNA damage-response mutants and increased lifespan, consistent with studies published fifteen years ago by Phil Hartman and by us. Since mammalian Sir2alpha decreases p53 expression and SIR2 overexpression in Saccharomyces cerevisiae increases replicative lifespan, we are currently studying possible cep-1 interactions with sir-2.1 in double mutants with the sir-2.1 overexpression and knockout strains. Thus, consistent with murine findings that an increase in p53 activity decreases lifespan and engenders segmental progeria, decreased expression of a C. elegans p53-like gene leads to a daf-16 -independent increase in lifespan.

Hodgkin JA (1988) Worm Breeder's Gazette "informational suppression by male-abnormal mutations: new names, new data."

Hodgkin JA

[Worm Breeder's Gazette, 1988]

As previously reported (WBG 10-1: 112, 10-2: 30, etc.) mutations at six different loci act as allele-specific suppressors of mutations in a variety of genes: tra-2, unc-54, etc. These mutations also have characteristic morphological effects on the anatomy of the adult male tail and, to a lesser extent, of the hermaphrodite vulva. For this reason they have hitherto been assigned to the mab (male abnormal) gene category: mab-1, ed) and mab-12 through mab-16 (unpublished). However, they exhibit such a distinctive set of common properties that it has been decided (with the agreement of Andy Papp, Victor Ambros, Rock Pulak and Phil Anderson, who have been responsible for isolating most of these suppressor mutations) to assign them to a new gene

K Kawamura et al. (1999) International C. elegans Meeting "Structure of Neural Network of C. elegans Observed by a Random Walker"

Y Osana, K Kawamura, K Oka, K Oshio, Y Funabashi, S Morita

[International C. elegans Meeting, 1999]

A database of synaptic connectivity of 302 neurons of the C. elegans has been constructed[1] from the observations of Albertson and Thomson[2] and White et al.[3] by some of the present authors. A network formed by 302 neurons of the C. elegans is represented on a computer by a network which consists of 302 dots combined by (arrowed) bonds. To analyse the structure of the neural network, behavior of a random walker on it is studied. The walker is displaced among dots which represent neurons over bonds which model synaptic connection. In terms of walking distance defined by minimum time steps which is necessary for the random walker to be displaced between neurons, distances among all neurons, whose synaptic connectivity are described by the above authors, have been determined. Almost all neurons are located within the walking distance of three time steps but walking distance among phalingeal neurons and somatic neurons are more than four time steps. The network is extended in a (more than) nine dimensional space around three nanohedra which are mutually combined by manifolds of less dimension. Each nanohedron consists of nine dots representing interneurons mutually connected by synapses and these nanohedra are located near the center of the network. The lattice is biased by the rectification of the chemical synapse in the sence that a random walker prefers to be displaced from sensory neurons to motor neurons. [1] K. Oshio, S. Morita, Y. Osana and K. Oka: C. elegans connectivity data, Technical report of CCEP, Keio Future No.1 (1998) [2] D. G. Albertson and J. N. Thomson: Phil. Trans R. Soc. Lond. B. 275 (1976) 299 [3] J. G. White, E. Southgate, J. N. Thomson and S. Brenner: Phil. Trans. R. Soc. Lond. B 314 (1986) 1.

Hartman PS et al. (1992) Photochem Photobiol "Inactivation of wild-type and rad mutant Caenorhabditis elegans by 8-methoxypsoralen and near ultraviolet radiation."

Marshall A, Hartman PS

[Photochem Photobiol, 1992]

Survival of wild-type and four radiation-sensitive (rad) mutants of the nematode Caenorhabditis elegans was determined after near-UV irradiation in the presence of 8-methoxypsoralen (8-MOP). Three sets of inactivation profiles were generated for each strain by irradiating synchronous populations of either early embryos, late embryos or first-stage larvae (L1s). Late embryos were consistently the most sensitive. Curiously, none of the four rad mutants were even moderately hypersensitive. Split-dose experiments indicated that DNA-DNA crosslinks were primarily responsible for lethality. Crosslink induction and repair were determined using two different assays. In both cases, little if any repair was observed in wild-type. This lack of repair thus explains why the rad mutants were not hypersensitive to 8-MOP photoinactivation. Since early embryos undergo extensive cell cycling, their resistance to 8-MOP photoinactivation suggests that replication is highly refractory to both monoadducts and crosslinks, as has been demonstrated previously for UV radiation-induced photoproducts (Hartman et al., 1991, Mutat. Res., 255, pp. 163-173).