Bergstrom AR et al. (1991) International C. elegans Meeting "IDENTIFICATION OF A Gj-LlKE PROTEIN IN C. ELEGANS MEMBRANES: POSSIBLE ROLE IN POLYAMINE METABOLISM."

Schaeffer JM, Bergstrom AR

[International C. elegans Meeting, 1991]

Guanine nucleotide-binding regulatory proteins (G-proteins) represent a family of trimeric proteins which mediate communication between a variety of cell surface receptors and effector molecules such as adenylate cyclase, phosphoinositol-specific phospholipase C and ion channels. When an appropriate agonist binds to a G protein- linked receptor, the alpha subunit of the G protein is activated and binds GTP resulting in a dissociation of the three G protein subunits. The intrinsic GTPase activity of the alpha subunit hydrolyzes GTP to GDP and the subsequent GDP-alpha subunit recombines with the other two G protein subunits ending the activation cycle. There are both stimulatory- and inhibitory-G proteins which are specifically ribosylated by bacterial toxins. We have identified and characterized a 41,000 dalton protein from C. elegans membranes with properties similar to previously described Gj-like proteins. Our laboratory previously presented evidence that specific dibenzoguanidines (and structural analogs including robenidine) are potent inhibitors of C. elegans ornithine decarboxylase, possibly acting via a G-protein. Consequently, the effect of robenidine on pertussis toxin stimulated ADP-ribosylation and GTPase activity in C. elegans has been evaluated. As shown in this poster, robenidine is a potent inhibitor of pertussis toxin-mediated ADP-ribosylation and stimulator of GTPase activity. Further work is required to determine whether there is a direct link between this Gj-like protein and ODC activity.

Furuhashi T et al. (2016) Biochem Biophys Res Commun "L-arginine, an active component of salmon milt nucleoprotein, promotes thermotolerance via Sirtuin in Caenorhabditis elegans."

Asahara Y, Matsunaga M, Furuhashi T, Sakamoto K

[Biochem Biophys Res Commun, 2016]

We previously showed that salmon milt nucleoprotein (NP) promotes thermotolerance in Caenorhabditis elegans; however, the active component and physiological mechanism of this effect has remained unclear. l-arginine (AR) is a major component of protamine and thus it has been proposed as the possible active component of NP. In this study, the viability of C. elegans treated with AR under heat stress was assessed and AR was shown to extend the survival term of the heat-stressed organisms. Additionally, AR was shown to restore the thrashing movement of the worms that is suppressed by heat stress. Treatment with AR was furthermore shown to promote thermotolerance in a DAF-16- and SIR-2.1-dependent manner, where DAF-16 and SIR-2.1 are homologs of FoxO and SirT1, respectively. Taken together, these data suggest that AR is one of the active components of NP and promotes thermotolerance via the activation of DAF-16 and SIR-2.1.

Schaeffer JM et al. (1988) Life Sci "Identification of gamma-aminobutyric acid and its binding sites in Caenorhabditis elegans."

Bergstrom AR, Schaeffer JM

[Life Sci, 1988]

Gamma-aminobutyric acid (GABA), glutamate decarboxylase and GABA-transaminase were identified in the nematode Caenorhabditis elegans. The concentration of GABA in C. elegans (0.14 ug/mg protein) is approximately 10-fold lower than the concentration of GABA in rat brain. Glutamate decarboxylase and GABA-transaminase, the GABA anabolic and catabolic enzymes, are also present in C. elegans. Crude membrane fractions were prepared from C. elegans and used to study specific [3H] GABA binding sites. GABA binds to C. elegans membranes with high affinity (37 nM) and low capacity (Bmax= 2.25 pmol/mg protein). Muscimol is a competitive inhibitor of specific GABA binding with a KI value 120 nM. None of the other GABA agonists or antagonists inhibited greater than 40% of the specific GABA binding at concentrations up to 10 -4M. Thirteen spider venoms were examined as possible GABA agonists or antagonists, the venom from Calilena agelenidae inhibits specific GABA binding with a KI value of 6 nl/ml. These results suggest that GABA has a physiological role as a neurotransmitter in C. elegans.

Toshiaki Suzuki et al. (2003) International Worm Meeting "Analysis of Ce-PARKIN in C. elegans"

Hideki Yashiroda, Yoshikuni Mizuno, Toshiaki Kitami, Keiji Tanaka, Shohei MItani, Keiko Gengyo-Ando, Toshiaki Suzuki, Noriyuki Matsuda, Nobutaka Hattori

[International Worm Meeting, 2003]

Autosomal recessive juvenile parkinsonism (AR-JP) is one of the most common forms of familial parkinsons disease characterized by selective loss of dopaminergic neurons in substantia nigra and the locus coeruleus. parkin is the causative gene of AR-JP. The human parkin gene encodes 465 amino acids with a ubiquitin-like domain in the amino-terminus and two RING finger motifs in the carboxy terminus. So far, various deletion mutations and point mutations have been discovered in patients of AR-JP, suggesting that the loss of function of Parkin is the cause of AR-JP. Recently we and others showed that Parkin has a ubiquitin-protein ligase activity and therefore suggested that the defect of protein degradation in the neurons of AR-JP patients (Shimura H. et al. Nat. Genet. 25, 302-5, 2000). To investigate the function of Parkin in vivo, we began to analyze the Ce-PARKIN of C. elegans. Two deletion mutations in parkin genes show no defect in their viabilities. The expression of Ce-PARKIN seems to be specific to subset of neurons. Therefore, we focused on the function of Ce-PARKIN in the neurons and the analysis is underway.

Hart EH et al. (2015) Vet Parasitol "A new enabling proteomics methodology to investigate membrane associated proteins from parasitic nematodes: case study using ivermectin resistant and ivermectin susceptible isolates of Caenorhabditis elegans and Haemonchus contortus."

Brophy PM, Bartley DJ, Prescott M, Hamilton JV, Wolf BT, Hart EH

[Vet Parasitol, 2015]

The mechanisms involved in anthelmintic resistance (AR) are complex but a greater understanding of AR management is essential for effective and sustainable control of parasitic helminth worms in livestock. Current tests to measure AR are time consuming and can be technically problematic, gold standard diagnostics are therefore urgently required to assist in combatting the threat from drug resistant parasites. For anthelmintics such as ivermectin (IVM), target proteins may be present in the cellular membrane. As proteins usually act in complexes and not in isolation, AR may develop and be measurable in the target associated proteins present in the parasite membrane. The model nematode Caenorhabditis elegans was used to develop a sub-proteomic assay to measure protein expression differences, between IVM resistant and IVM susceptible isolates in the presence and absence of drug challenge. Evaluation of detergents including CHAPS, ASB-14, C7BzO, Triton x100 and TBP (tributyl phosphine) determined optimal conditions for the resolution of membrane proteins in Two Dimensional Gel Electrophoresis (2DE). These sub-proteomic methodologies were then translated and evaluated using IVM-susceptible and IVM-resistant Haemonchus contortus; a pathogenic blood feeding parasitic nematode which is of global importance in livestock health, welfare and productivity. We have demonstrated the successful resolution of membrane associated proteins from both C. elegans and H. contortus isolates, using a combination of CHAPS and the zwitterionic amphiphilic surfactant ASB-14 to further support the detection of markers for AR.

Schaeffer JM et al. (1989) Biochem J "MK-801 is a potent nematocidal agent. Characterization of MK-801 binding sites in Caenorhabditis elegans."

Schaeffer JM, Turner MJ, Bergstrom AR

[Biochem J, 1989]

MK-801, an N-methyl-D-aspartate antagonist in mammalian brain tissue, is a potent nematocidal agent. Specific MK-801 binding sites have been identified and characterized in a membrane fraction prepared from the free-living nematode Caenorhabditis elegans. The high-affinity MK-801 binding site has an apparent dissociation constant, Kd, of 225 nM. Unlike the MK-801 binding site in mammalian tissues, the C. elegans binding site is not effected by glutamate or glycine, and polyamines are potent inhibitors of specific MK-801 binding.

Rohrer SP et al. (1990) Comp Biochem Physiol C "A membrane associated glutamate binding protein from Caenorhabditis elegans and Haemonchus contortus."

Rohrer SP, Evans DV, Bergstrom AR

[Comp Biochem Physiol C, 1990]

1. A glutamate binding protein has been identified in membrane preparations from the free living nematode, Caenorhabditis elegans, and from the parasitic nematode, Haemonchus contortus. 2. This putative glutamate receptor was solubilized with 30 mM octyl-B-glucoside and partially purified by anion exchange and gel filtration chromatography. 3. An 80-fold purification with recovery of 75% of the glutamate binding activity was achieved. 4. The soluble C. elegans binding protein displayed a Kd for glutamate of 0.1 microM, in close agreement with the findings for the membrane associated binding protein. 5. Quisqualate was capable of displacing glutamate from the soluble C. elegans receptor, again in agreement with previous findings for the membrane bound receptor. 6. The fact that a parasitic nematode, Haemonchus contortus, also possesses this putative glutamate receptor, strengthens the case for using C. elegans as a model system for the study of parasitic nematode neuromuscular physiology.

Schaeffer JM et al. (1994) Biochem Pharmacol "Nematocidal activity of MK-801 analogs and related drugs. Structure-activity relationships."

Bergstrom AR, Frazier EG, Schaeffer JM, Underwood D

[Biochem Pharmacol, 1994]

A series of dibenzo[a,d]cycloalkenimines were evaluated for their affinity to the (+)-5-methyl-10,11-dihydro-5H-dibenzo[a,d]cyclohepten-5,10-imine (MK-801) binding site in Caenorhabditis elegans membranes and their nematocidal activity. The (+)-MK-801 enantiomer (1) had a higher affinity (Kd = 240 nM) for its specific binding site and was a more potent nematocidal agent than the (-)-MK-801 enantiomer (-1). Ring expansion to form the dibenzo[a,d]cyclooctenimine analogs generally resulted in more potent compounds. The most potent of this series (23) was approximately 7-fold more potent than (+)-MK-801. A good correlation was established between binding affinities and nematocidal activity for all of the analogs that were tested. However, there was no correlation between binding to C. elegans membranes and affinity for mammalian MK-801 binding sites. Other noncompetitive inhibitors of the mammalian N-methyl-D-aspartate site were examined, and a series of diphenylguanidines were identified as potent competitive inhibitors of MK-801 binding to C. elegans membranes, in addition to displaying potent nematocidal activity. The most potent diphenylguanidine analog (24) was approximately 80-fold more potent than (+)-MK-801 in both its affinity for the MK-801 binding site and as a nematocidal agent. Molecular modeling studies support the hypothesis that the diphenylguanidines and MK-801 are binding to the same site and suggest that more potent compounds may be developed by effective modeling of the existing compounds.

Shaver AO et al. (2024) Trends Parasitol "Integrating metabolomics into the diagnosis and investigation of anthelmintic resistance."

Andersen EC, Shaver AO

[Trends Parasitol, 2024]

Anthelmintic resistance (AR) in parasitic nematodes poses a global health problem in livestock and domestic animals and is an emerging problem in humans. Consequently, we must understand the mechanisms of AR, including target-site resistance (TSR), in which mutations affect drug binding, and non-target site resistance (NTSR), which involves alterations in drug metabolism and detoxification processes. Because much of the focus has been on TSR, NTSR has received less attention. Here, we describe how metabolomics approaches using Caenorhabditis elegans offer the ability to disentangle nematode drug metabolism, identify metabolic changes associated with resistance, uncover novel biomarkers, and enhance diagnostic methods.

Aliota MT et al. (2010) PLoS Negl Trop Dis "Mosquito transcriptome profiles and filarial worm susceptibility in Armigeres subalbatus."

Fuchs JF, Rocheleau TA, Aliota MT, Clark AK, Christensen BM, Chen CC, Hillyer JF

[PLoS Negl Trop Dis, 2010]

BACKGROUND: Armigeres subalbatus is a natural vector of the filarial worm Brugia pahangi, but it kills Brugia malayi microfilariae by melanotic encapsulation. Because B. malayi and B. pahangi are morphologically and biologically similar, comparing Ar. subalbatus-B. pahangi susceptibility and Ar. subalbatus-B. malayi refractoriness could provide significant insight into recognition mechanisms required to mount an effective anti-filarial worm immune response in the mosquito, as well as provide considerable detail into the molecular components involved in vector competence. Previously, we assessed the transcriptional response of Ar. subalbatus to B. malayi, and now we report transcriptome profiling studies of Ar. subalbatus in relation to filarial worm infection to provide information on the molecular components involved in B. pahangi susceptibility. METHODOLOGY/PRINCIPAL FINDINGS: Utilizing microarrays, comparisons were made between mosquitoes exposed to B. pahangi, B. malayi, and uninfected bloodmeals. The time course chosen facilitated an examination of key events in the development of the parasite, beginning with the very start of filarial worm infection and spanning to well after parasites had developed to the infective stage in the mosquito. At 1, 3, 6, 12, 24 h post infection and 2-3, 5-6, 8-9, and 13-14 days post challenge there were 31, 75, 113, 76, 54, 5, 3, 13, and 2 detectable transcripts, respectively, with significant differences in transcript abundance (increase or decrease) as a result of parasite development. CONCLUSIONS/SIGNIFICANCE: Herein, we demonstrate that filarial worm susceptibility in a laboratory strain of the natural vector Ar. subalbatus involves many factors of both known and unknown function that most likely are associated with filarial worm penetration through the midgut, invasion into thoracic muscle cells, and maintenance of homeostasis in the hemolymph environment. The data show that there are distinct and separate transcriptional patterns associated with filarial worm susceptibility as compared to refractoriness, and that an infection response in Ar. subalbatus can differ significantly from that observed in Ae. aegypti, a common laboratory model.