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(Protein Kinase D1) mutants have age-dependent defects in locomotion and neuromuscular transmission.MicroPublication Biology 2023Changes in neuronal function that occur with age are an area of increasing importance. A potential significant contributor to age-dependent decline may be alterations to...
Changes in neuronal function that occur with age are an area of increasing importance. A potential significant contributor to age-dependent decline may be alterations to neurotransmitter release. Protein kinases, such as Protein Kinase C and Protein Kinase A, are well characterised modulators of neuronal function and neurotransmission. Protein Kinase D (PRKD) is a serine/threonine kinase whose role in neurons is less well characterised. Here we report that mutations in the PRKD homolog, , show an acceleration in age-dependent decline of locomotion rate and an alteration to age-dependent changes in aldicarb sensitivity. These effects could be explained by a pre- or post-synaptic function of the protein kinase as the animal ages.
PubMed: 37090152
DOI: 10.17912/micropub.biology.000800 -
Human Molecular Genetics Mar 2023The principal component of the protein homeostasis network is the ubiquitin-proteasome system. Ubiquitination is mediated by an enzymatic cascade involving, i.e. E3...
The principal component of the protein homeostasis network is the ubiquitin-proteasome system. Ubiquitination is mediated by an enzymatic cascade involving, i.e. E3 ubiquitin ligases, many of which belong to the cullin-RING ligases family. Genetic defects in the ubiquitin-proteasome system components, including cullin-RING ligases, are known causes of neurodevelopmental disorders. Using exome sequencing to diagnose a pediatric patient with developmental delay, pyramidal signs and limb ataxia, we identified a de novo missense variant c.376G>C; p.(Asp126His) in the FEM1C gene encoding a cullin-RING ligase substrate receptor. This variant alters a conserved amino acid located within a highly constrained coding region and is predicted as pathogenic by most in silico tools. In addition, a de novo FEM1C mutation of the same residue p.(Asp126Val) was associated with an undiagnosed developmental disorder, and the relevant variant (FEM1CAsp126Ala) was found to be functionally compromised in vitro. Our computational analysis showed that FEM1CAsp126His hampers protein substrate binding. To further assess its pathogenicity, we used the nematode Caenorhabditis elegans. We found that the FEM-1Asp133His animals (expressing variant homologous to the FEM1C p.(Asp126Val)) had normal muscle architecture yet impaired mobility. Mutant worms were sensitive to the acetylcholinesterase inhibitor aldicarb but not levamisole (acetylcholine receptor agonist), showing that their disabled locomotion is caused by synaptic abnormalities and not muscle dysfunction. In conclusion, we provide the first evidence from an animal model suggesting that a mutation in the evolutionarily conserved FEM1C Asp126 position causes a neurodevelopmental disorder in humans.
Topics: Animals; Humans; Child; Proteasome Endopeptidase Complex; Cullin Proteins; Acetylcholinesterase; Speech; Ubiquitin-Protein Ligases; Neurodevelopmental Disorders; Ubiquitin; Caenorhabditis elegans; Ataxia; Ubiquitin-Protein Ligase Complexes
PubMed: 36336956
DOI: 10.1093/hmg/ddac276 -
G3 (Bethesda, Md.) Dec 2022Animals exhibit phenotypic plasticity through the interaction of genes with the environment, and little is known about the genetic factors that change synaptic function...
Animals exhibit phenotypic plasticity through the interaction of genes with the environment, and little is known about the genetic factors that change synaptic function at different developmental stages. Here, we investigated the genetic determinants of how animal's sensitivity to drugs that alter synaptic activity is regulated at a specific developmental stage using the free-living nematode Caenorhabditis elegans. C. elegans enters the stress-resistant dauer larval stage under harsh conditions. Although dauer is known to have reduced permeability and increased resistance to most known exogenous chemicals, we discovered that dauer is hypersensitive to a cholinesterase inhibitor, aldicarb. To investigate genes regulating dauer-specific acetylcholine transduction, we first screened for aldicarb-resistant mutations in dauer and then performed a secondary screen to rule out aldicarb-resistant mutations that also affect adults. We isolated 2 different mutations of a single gene called cyp-34A4 or dach-1 encoding a cytochrome P450. In the nondauer stages, dach-1 is mainly expressed in the intestine, but its expression is robustly increased in the epidermis of dauers. By tissue-specific rescue experiments, we found that dach-1 modulates aldicarb sensitivity in a cell nonautonomous manner. In addition, dach-1 plays pleiotropic functions in dauers by regulating quiescence and surviving heat shock and hyperosmolar stress. Our study reveals novel functions of the cytochrome P450 in synaptic and physiological changes during the developmental plasticity.
Topics: Animals; Caenorhabditis elegans; Caenorhabditis elegans Proteins; Larva; Aldicarb; Alleles; Cytochrome P-450 Enzyme System
PubMed: 36194018
DOI: 10.1093/g3journal/jkac266 -
Frontiers in Nutrition 2022The common carbamate insecticide aldicarb is considered one of the most acutely toxic pesticides. Herein, rational design was used to synthesize two haptens with spacers...
The common carbamate insecticide aldicarb is considered one of the most acutely toxic pesticides. Herein, rational design was used to synthesize two haptens with spacers of different carbon chain lengths. The haptens were then used to immunize mice. The antibodies obtained were evaluated systematically, and a colloidal gold immunochromatographic strip was developed based on an anti-aldicarb monoclonal antibody. The 50% inhibition concentration and linear range of anti-aldicarb monoclonal antibody immunized with Hapten 1 were 0.432 ng/mL and 0.106-1.757 ng/mL, respectively. The cross-reactivities for analogs of aldicarb were all <1%. The limit of detection of the colloidal gold immunochromatographic strip was 30 μg/kg, and the average recoveries of aldicarb ranged from 80.4 to 110.5% in spiked samples. In the analysis of spiked samples, the test strip could accurately identify positive samples detected by the instrumental method in the GB 23200.112-2018 standard but produced some false positives for negative samples. This assay provides a rapid and accurate preliminary screening method for the determination of aldicarb in agricultural products and environments.
PubMed: 36082035
DOI: 10.3389/fnut.2022.976284 -
MicroPublication Biology 2022Atropine has been used as an established anticonvulsant treatment for nerve agent intoxication. Atropine reduces electroshock recovery time among aldicarb-exposed...
Atropine has been used as an established anticonvulsant treatment for nerve agent intoxication. Atropine reduces electroshock recovery time among aldicarb-exposed wild-type .
PubMed: 36017190
DOI: 10.17912/micropub.biology.000621 -
American Journal of Human Genetics Aug 2022PPFIBP1 encodes for the liprin-β1 protein, which has been shown to play a role in neuronal outgrowth and synapse formation in Drosophila melanogaster. By exome and...
PPFIBP1 encodes for the liprin-β1 protein, which has been shown to play a role in neuronal outgrowth and synapse formation in Drosophila melanogaster. By exome and genome sequencing, we detected nine ultra-rare homozygous loss-of-function variants in 16 individuals from 12 unrelated families. The individuals presented with moderate to profound developmental delay, often refractory early-onset epilepsy, and progressive microcephaly. Further common clinical findings included muscular hyper- and hypotonia, spasticity, failure to thrive and short stature, feeding difficulties, impaired vision, and congenital heart defects. Neuroimaging revealed abnormalities of brain morphology with leukoencephalopathy, ventriculomegaly, cortical abnormalities, and intracranial periventricular calcifications as major features. In a fetus with intracranial calcifications, we identified a rare homozygous missense variant that by structural analysis was predicted to disturb the topology of the SAM domain region that is essential for protein-protein interaction. For further insight into the effects of PPFIBP1 loss of function, we performed automated behavioral phenotyping of a Caenorhabditis elegans PPFIBP1/hlb-1 knockout model, which revealed defects in spontaneous and light-induced behavior and confirmed resistance to the acetylcholinesterase inhibitor aldicarb, suggesting a defect in the neuronal presynaptic zone. In conclusion, we establish bi-allelic loss-of-function variants in PPFIBP1 as a cause of an autosomal recessive severe neurodevelopmental disorder with early-onset epilepsy, microcephaly, and periventricular calcifications.
Topics: Acetylcholinesterase; Animals; Drosophila melanogaster; Epilepsy; Loss of Heterozygosity; Microcephaly; Nervous System Malformations; Neurodevelopmental Disorders; Pedigree
PubMed: 35830857
DOI: 10.1016/j.ajhg.2022.06.008 -
Toxicology Research Jun 2022Aldicarb, a carbamate pesticide banned in France since 2008, represents a persistent risk of human poisoning. There is no up-to-date picture of aldicarb intentional...
Aldicarb, a carbamate pesticide banned in France since 2008, represents a persistent risk of human poisoning. There is no up-to-date picture of aldicarb intentional poisonings associated to detailed diagnosis and clinical management of cases to evaluate the effect of its ban on intoxication risk reduction, including suicide. This retrospective epidemiological study describes cases of suicidal intoxication from the Hauts-de-France region between 2012 and 2021 and illustrates this situation through one analytically documented case. 60 cases were collected, mostly presenting a pathognomonic symptomatology. Thirty-five victims presented a muscarinic syndrome (58.3%), 14 a nicotinic syndrome (23.3%), and 37 a central nervous system impairment (61.7%). Hospitalization was necessary for 44 cases (73.3%), with 2 fatal evolutions. Diagnosis was based on the blood cholinesterase activities. Among the 25 cases with toxicology results, 45.8% presented a moderate decrease of acetylcholinesterase activity, whereas 87.5% presented a strong decrease of butyrylcholinesterase activity. Blood or urine detection of aldicarb and its metabolites may be considered in therapeutic management, although their quantification is unlikely to change the emergency medical care. Our study updates epidemiology of aldicarb poisoning at a regional level, highlighting a persisting health threatening situation with banned pesticides. More than 65% of national cases occurred in this agricultural area, aldicarb remaining available from storage of previously purchased products. Robust evidence is presented that acute poisoning is an ongoing major global public health challenge. There is a need for continued international efforts in risk reduction, knowledge, and information strategies.
PubMed: 35782643
DOI: 10.1093/toxres/tfac031 -
Frontiers in Bioengineering and... 2021In this study, was grown on municipal biosolids (BS) as the substrate to produce laccase for the removal of pesticides (fungicides, herbicides, and insecticides) from...
In this study, was grown on municipal biosolids (BS) as the substrate to produce laccase for the removal of pesticides (fungicides, herbicides, and insecticides) from wastewater. Among the various types of BS tested, sterilized biosolids were the most promising substrate for laccase production by with a maximal laccase activity (162.1 ± 21.1 U/g dry substrate), followed by hygenized biosolids (96.7 ± 17.6 U/g dry substrate), unsterilized biosolids (UBS) (31.9 ± 1.2 U/g dry substrate), and alkali-treated biosolids (8.2 ± 0.4 U/g dry substrate). The ultrasound-assisted extraction of this enzyme from fermented UBS was carried out with 0.1 M phosphate buffer at pH 7.0, which increased the enzyme activity of the crude extract by 30%. To test the catalytic potential of the biocatalyst in real matrices, 1 U/ml of recovered crude laccase extract was applied for 24 h for the removal of 29 pesticides (nine fungicides, 10 herbicides, and 10 insecticides) either separately or as a mixture from spiked biologically treated wastewater effluent. When treated with crude enzyme extract, high-priority herbicides metolachlor and atrazine were completely removed, while 93%-97% of the insecticides aldicarb, spinosad, and azinphos-methyl and up to 91% of kresoxim-methyl were removed. Promising results were obtained with BS-derived crude enzyme extract exhibiting improved pesticides removal, which may be due to the mediator effect resulting from the catalytic transformation of other molecules in the cocktail. The results demonstrated a promising integrated bioprocess for the removal of pesticides in wastewater using crude laccase obtained from BS.
PubMed: 35223809
DOI: 10.3389/fbioe.2021.770435 -
The Journal of Biological Chemistry Jan 2022Complex biological functions within organisms are frequently orchestrated by systemic communication between tissues. In the model organism Caenorhabditis elegans, the...
Complex biological functions within organisms are frequently orchestrated by systemic communication between tissues. In the model organism Caenorhabditis elegans, the pharyngeal and body wall neuromuscular junctions are two discrete structures that control feeding and locomotion, respectively. Separate, the well-defined neuromuscular circuits control these distinct tissues. Nonetheless, the emergent behaviors, feeding and locomotion, are coordinated to guarantee the efficiency of food intake. Here, we show that pharmacological hyperactivation of cholinergic transmission at the body wall muscle reduces the rate of pumping behavior. This was evidenced by a systematic screening of the effect of the cholinesterase inhibitor aldicarb on the rate of pharyngeal pumping on food in mutant worms. The screening revealed that the key determinants of the inhibitory effect of aldicarb on pharyngeal pumping are located at the body wall neuromuscular junction. In fact, the selective stimulation of the body wall muscle receptors with the agonist levamisole inhibited pumping in a lev-1-dependent fashion. Interestingly, this response was independent of unc-38, an alpha subunit of the nicotinic receptor classically expressed with lev-1 at the body wall muscle. This implies an uncharacterized lev-1-containing receptor underpins this effect. Overall, our results reveal that body wall cholinergic transmission not only controls locomotion but simultaneously inhibits feeding behavior.
Topics: Aldicarb; Animals; Caenorhabditis elegans; Caenorhabditis elegans Proteins; Cholinesterase Inhibitors; Feeding Behavior; Levamisole; Neuromuscular Junction; Signal Transduction
PubMed: 34864060
DOI: 10.1016/j.jbc.2021.101466 -
Comparative Biochemistry and... Feb 2022Acetylcholinesterase (AChE; EC 3.1.1.7) from aquatic organisms have been used to evaluate the exposure of specimens to pesticides and heavy metals at sublethal levels in...
Acetylcholinesterase from the charru mussel Mytella charruana: kinetic characterization, physicochemical properties and potential as in vitro biomarker in environmental monitoring of mollusk extraction areas.
Acetylcholinesterase (AChE; EC 3.1.1.7) from aquatic organisms have been used to evaluate the exposure of specimens to pesticides and heavy metals at sublethal levels in environmental samples. AChE of Mytella charruana was extracted to characterize its physicochemical and kinetic properties as well as the effect of organophosphate (dichlorvos, diazinon, chlorpyrifos, methyl-parathion and temephos), carbamates (carbaryl, carbofuran and aldicarb), benzoylureas (diflubenzuron and novaluron), pyrethroid (cypermethrin) and juvenile hormone analog - JHA (pyriproxyfen) and the effect of metal ions: Hg, Cd, Pb, As, Cu and Zn, in order to evaluate the potential of the enzyme as biomarker. The optimum pH of M. charruana AChE was 8.5 and the maximum activity peak occurred at 48 °C, being highly thermostable maintaining 97.8% of its activity after incubation at 60 °C. The Michaelis-Menten constants (k) for the substrates acetylthiocholine and propionylthiocholine were 2.8 ± 1.26 and 4.94 ± 6.9 mmol·L, respectively. The V values for the same substrates were 22.6 ± 0.90 and 10.2 ± 4.94 mU·mg, respectively. Specific inhibition results suggest an AChE presenting active site with dimensions between those of AChE and butyrylcholinesterase (BChE). The IC values related to the effect of the pesticides on the enzyme showed higher inhibitory power of temephos (0.17 μmol·L), followed by aldicarb (0.19 μmol·L) and diflubenzuron (0.23 μmol·L). Metal ions inhibited M. charruana enzyme in the following order: Hg > Pb > Cd > As > Cu > Zn. These data suggest that the enzyme showed potential as in vitro biomarker of the exposure to temephos, mercury, zinc and copper.
Topics: Acetylcholinesterase; Animals; Biomarkers; Bivalvia; Environmental Monitoring; Gene Expression Regulation, Enzymologic; Metals, Heavy; Pesticides; Water Pollutants, Chemical
PubMed: 34744030
DOI: 10.1016/j.cbpc.2021.109225