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[
International Worm Meeting,
2021]
Animals adjust their phenotype to their environment, which is thought to provide a selective benefit under changing conditions. A famous example of such phenotypic plasticity is the slow down of growth and the delay of aging under dietary restriction (DR). Interestingly, parental DR also affects phenotypes of the F1 generation, for example, DR increases the starvation resistance of progeny (Jordan et al., 2019). Molecularly, DR is associated with a global change in proteome expression, including a reduction in ribosome levels and a proteome-wide slow-down of protein turnover (Dhondt et al., 2016; Visscher et al., 2016). Here, we ask if and how parental diet impacts the proteome of the F1 progeny. We will seek causal relationships between intergenerationally transmitted phenotypes and the inherited proteome. Finally, we aim to determine if the parental control of the proteome provides a selective benefit to their progeny. To address these questions, we have established a tool to precisely quantify ribosomal levels by measuring the luminescence from lysis of HiBit tagged to ribosomal proteins. We plan to globally analyze parental effects on the F1 proteome by proteomics and measure the dynamics of proteome changes, growth rates, and reproduction rates in different nutritional conditions by fluorescence live imaging in microchambers. The project will likely provide important insights about the molecular mechanisms and physiological significance of inter-generational inheritance induced by parental diet.
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[
Radiat Prot Dosimetry,
1994]
For the first time track structure theory has been applied to radiobiological effects in a living organism. Data for lethal mutagenesis in Caenorhabditis elegans, obtained after irradiation with nine different types of ions of atomic number 1-57 and gamma rays have yielded radiosensitivity parameters (E0, sigma 0, kappa, m = 68 Gy, 2.5 x 10(-9)
cm2, 750, 2) comparable with those found for the transformation of C3HT10 1/2 cells (180 Gy, 1.15 x 10(-10)
cm2, 750, 2) but remote from those (E0 and sigma 0 = approximately 2 Gy, approximately 5 x 10(-7)
cm2) for mammalian cell survival.
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[
Nematology,
2000]
A technique is described that refines the standard sugar flotation procedure used to isolate nematodes from their surroundings. By centrifuging nematodes in a number of increasing specific gravity solutions and plotting the fraction floating, the cumulative probability distribution of the population's specific gravity is generated. By assuming normality, the population mean, mu, and standard deviation, sigma, are found by a nonlinear least squares procedure. These density parameters along with their error covariance matrix may be used as a taxonomic physical character. A chi-squared test is derived for comparing populations. Mean and standard deviation pairs (mu, sigma) were found for the specific gravities of the adult stage of the plant parasites Pratylenchus agilis (1.068, 0.017), P. scribneri (1.073, 0.028), P. penetrans (1.058, 0.008) and the bacterial-feeder Caenorhabditis elegans (1.091, 0.016).
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[
European Worm Meeting,
1998]
The Glutathione S-Transferases (GST) are a superfamily of enzymes that catalyse the nucleophilic addition of the tripeptide glutathione (GSH) to endogenous and xenobiotic electrophilic substrates. GSTs are principally detoxification enzymes, but have been implicated in the development of resistance to a range of pesticides, herbicides and a number of drugs. These enzymes also serve as non-catalytic carrier proteins (ligandins). Recently, a sigma class related GST isolated from Ascaridia galli, displayed a high level of specific activity in the GSH dependent isomerisation of prostaglandin H to prostaglandin E (1). This suggests that certain sigma GSTs may be involved in eicosanoid metabolism. We are investigating the possible functions of members of the sigma class GST of C.elegans. To date, seven sigma class GST-like genes have been identified in C.elegans. We have amplified the corresponding cDNA for each gene by PCR from a cDNA library, and successfully expressed six of the seven cDNA sequences in Escherichia Coli. The recombinant protein product of the R03D7.6 gene has GSH binding properties, allowing purification by affinity chromatography. This protein also shows high levels of activity towards the universal GST substrate 1-chloro-2,4-dinitrobenzene (CDNB). We intend to utilise each recombinant protein firstly to raise monoclonal antibodies to localise the native enzymes in vivo. Secondly, we will conduct a series of assays to analyse the substrate specificities of each recombinant protein. In addition we would like to inject combinations of dsRNAs prepared from the GST cDNAs, to study the effects of interfering with the expression of this GST family. 1 Meyer D.J., Muimo R., Thomas M., Coates D. and Isaac R.E. (1996) Biochem. J. 313, 223-227
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[
Worm Breeder's Gazette,
1994]
In my abstract for the '93 C. elegans meeting, I provided the recipe for Flo Youngren's EZ worm mix. Since then, I have come up with a better formulation, which has less salt (for superior lawn consistency) and does not include yeast extract. These plates have three main advantages over conventional NGM plates: 1 ) Annoying salt crystals do not readily form 2) No stock solutions need to be added after autoclaving, and 3) There is minimal fuss and/or room for error when undergraduate assistants make up the plates. Here is the recipe for MYOB (Modified Youngren's, Only Bacto-peptone). For 100 liters of plates: Combine 55 g Tris-CI (Sigma reagent grade), 24 g Tris-OH (Sigma reagent grade), 310 g Bacto Peptone (Difco), 800 mg Cholesterol and 200 g NaCI. Mix thoroughly and be sure to avoid chunks; this powdered mixture can be stored for many months prior to use. Dissolve 5.9 g in 1 liter of water, add 20 g of agar and autoclave.
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Ferragud A, Scott LL, Satarasinghe PN, Shen A, Hodges TR, Friese KM, Prakash BA, Sabino V, Martin SF, Pierce JT, Wood MD, Sahn JJ, Yen RC, Shi T
[
Neuropsychopharmacology,
2018]
Repeated cycles of intoxication and withdrawal enhance the negative reinforcing properties of alcohol and lead to neuroadaptations that underlie withdrawal symptoms driving alcohol dependence. Pharmacotherapies that target these neuroadaptations may help break the cycle of dependence. The sigma-1 receptor (1R) subtype has attracted interest as a possible modulator of the rewarding and reinforcing effects of alcohol. However, whether the sigma-2 receptor, recently cloned and identified as transmembrane protein 97 (2R/TMEM97), plays a role in alcohol-related behaviors is currently unknown. Using a Caenorhabditis elegans model, we identified two novel, selective 2R/Tmem97 modulators that reduce alcohol withdrawal behavior via an ortholog of 2R/TMEM97. We then show that one of these compounds blunted withdrawal-induced excessive alcohol drinking in a well-established rodent model of alcohol dependence. These discoveries provide the first evidence that 2R/TMEM97 is involved in alcohol withdrawal behaviors and that this receptor is a potential new target for treating alcohol use disorder.
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[
J Comput Biol,
2010]
The current pairwise RNA (secondary) structural alignment algorithms are based on Sankoff's dynamic programming algorithm from 1985. Sankoff's algorithm requires O(N(6)) time and O(N(4)) space, where N denotes the length of the compared sequences, and thus its applicability is very limited. The current literature offers many heuristics for speeding up Sankoff's alignment process, some making restrictive assumptions on the length or the shape of the RNA substructures. We show how to speed up Sankoff's algorithm in practice via non-heuristic methods, without compromising optimality. Our analysis shows that the expected time complexity of the new algorithm is O(N(4)sigma(N)), where sigma(N) converges to O(N), assuming a standard polymer folding model which was supported by experimental analysis. Hence, our algorithm speeds up Sankoff's algorithm by a linear factor on average. In simulations, our algorithm speeds up computation by a factor of 3-12 for sequences of length 25-250. Code and data sets are available, upon request.
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[
International C. elegans Meeting,
1999]
The Glutathione S-Transferases (GST) are a superfamily of enzymes that catalyse the nucleophilic addition of the tripeptide glutathione (GSH) to endogenous and xenobiotic electrophilic substrates. GSTs are principally detoxification enzymes, but have been implicated in the development of resistance to a range of pesticides, herbicides and a number of drugs. These enzymes also serve as non-catalytic carrier proteins (ligandins). Recently, a sigma class related GST isolated from Ascaridia galli , displayed a high level of specific activity in the GSH dependent isomerisation of prostaglandin H to prostaglandin E 1 . This suggests that certain sigma GSTs may be involved in eicosanoid metabolism. We are investigating the possible functions of members of the sigma ( s ) class GST of C.elegans . 20 s class GST-like genes have been identified in C.elegans . We have amplified the corresponding cDNA for 7 of these genes by PCR from a cDNA library, and successfully expressed six of the seven cDNA sequences in Escherichia coli . The recombinant protein product of the R03D7.6 gene has GSH binding properties, allowing purification by affinity chromatography. This protein also shows high levels of activity towards the universal GST substrate 1-chloro-2,4-dinitrobenzene (CDNB). Antibodies raised to recombinant R03D7.6 have been used to immunoblot GST protein in crude extracts of mixed stage C. elegans and in column purified native GST, demonstrating that R03D7.6 is expressed at detectable levels. The antibody is being used to validate the expression pattern obtained with nematodes transformed with a R03D7.6:: lacZ promoter construct. 1 Meyer D.J., Muimo R., Thomas M., Coates D. and Isaac R.E. (1996) Biochem. J. 313 , 223-227
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[
Infect Immun,
2003]
Staphylococcus aureus, an important pathogen of humans and other warm-blooded animals, is also capable of killing the nematode Caenorhabditis elegans. Here, we show that C. elegans organisms that are fed S. aureus die over the course of several days in a process that is correlated with the accumulation of bacteria within the nematode digestive tract. Several S. aureus virulence determinants known or speculated to be important in rnammalian pathogenesis, including the quorum-sensing global virulence regulatory system agr and the global virulence regulator sarA, the alternative sigma factor sigma(B), alpha-hemolysin, and V8 serine protease, are required for full pathogenicity in nematodes. In addition, several defined C. elegans mutants were examined for susceptibility to S. aureus infection. Enhanced susceptibility to S. aureus killing was observed with loss-of-function mutations in the C. elegans genes
esp-2/sek-1 and
esp-8/nsy-1, which encode components of a conserved
p38 MAP kinase signaling pathway involved in nematode defense against multiple pathogens. These results suggest that key aspects of S. aureus pathogenesis have been conserved, irrespective of the host, and that specific C. elegans host factors can alter susceptibility to this gram-positive human pathogen.
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[
J Bacteriol,
2001]
We cloned the rpoN (ntrA, glnF) gene encoding the alternate sigma factor sigma(54) from the opportunistic multihost pathogen Pseudomonas aeruginosa strain PA14. A marker exchange protocol was used to construct the PA14 rpoN insertional mutation rpoN::Gen(r). PA14 rpoN::Gen(r) synthesized reduced levels of pyocyanin and displayed a variety of phenotypes typical of rpoN mutants, including a lack of motility and the failure to grow on nitrate, glutamate, or histidine as the sole nitrogen source. Compared to wild-type PA14, rpoN::Gen(r) was ca. 100-fold less virulent in a mouse thermal injury model and was significantly impaired in its ability to kill the nematode Caenorhabditis elegans. In an Arabidopsis thaliana leaf infectivity assay, although rpoN::Gen(r) exhibited significantly reduced attachment to trichomes, stomata, and the epidermal cell surface, did not attach perpendicularly to or perforate mesophyll cell walls, and proliferated less rapidly in Arabidopsis leaves, it nevertheless elicited similar disease symptoms to wild-type P. aeruginosa PA14 at later stages of infection. rpoN::Gen(r) was not impaired in virulence in a Galleria mellonella (greater wax moth) pathogenicity model. These data indicate that rpoN does not regulate the expression of any genes that encode virulence factors universally required for P. aeruginosa pathogenicity in diverse hosts.