MacDonald D et al. (2004) J Ethnopharmacol "Ascaridole-less infusions of Chenopodium ambrosioides contain a nematocide(s) that is(are) not toxic to mammalian smooth muscle."

Rosenfeld J, Rangachari PK, Sorger G, Warren C, MacDonald D, VanCrey K, Harrison P

[J Ethnopharmacol, 2004]

Infusions of Chenopodium ambrosioides (L.) have been used for centuries in the Americas as a popular remedy against intestinal worm infections. The essential oil of Chenopodium ambrosioides contains high levels of ascaridole, which is a potent anthelmintic, but which has also been responsible for human fatalities, leading to its disuse. Almost 90% of the nematocidal activity of Chenopodium ambrosioides infusions was due to a hydrophilic component different from ascaridole. Synthetic ascaridole and the ascaridole from infusions, extracted into hexane, caused a reduction of carbachol-induced contractions in rat gastrointestinal smooth muscle at concentrations required to kill Caenorhabditis elegans (L.). The herbal infusion and the ascaridole-free hexane-extracted aqueous residue of the above infusion, at nematocidal concentrations, had no detectable effect on smooth muscle contraction in the above system. It would appear that the traditional form of usage of Chenopodium ambrosioides infusions as a vermifuge is safer than the use of the herb's essential oil.

MacDonald, Lindsay D. et al. (2009) International Worm Meeting "Protein degradation in the C. elegans germ line can regulate entry into meiosis."

Hansen, Dave, MacDonald, Lindsay D., Knox, Aaron

[International Worm Meeting, 2009]

In any population of stem cells a delicate balance between proliferation and differentiation must be maintained to ensure proper stem cell function and tissue homeostasis. In the C. elegans germ line, this balance is controlled by the GLP-1/Notch signaling pathway. GLP-1/Notch signaling promotes proliferation, at least in part, by negatively regulating two redundant downstream pathways (GLD-1/NOS-3, GLD-2/GLD-3), which inhibit proliferation or promote meiotic entry. We have recently identified a weak loss of function allele of one of the proteasome alpha subunits (pas-5(oz237)), that enhances an overproliferation phenotype in the C. elegans germ line. Through genetic epistasis we have found that the proteasome has two separate functions in regulating the decision between proliferation and meiotic entry. First, the proteasome acts as a general negative regulator of Notch signaling. sel-10, an E3 ubiquitin ligase, has been implicated in GLP-1 INTRA degradation by targeting INTRA to the proteasome. However, when proteasome function is reduced and SEL-10 is no longer present in the cell, an enhancement of Notch signaling is still seen, suggesting a second E3 ubiquitin ligase is required for proper ubiquitination and subsequent degradation of INTRA. Second, the proteasome is involved in negatively regulating proliferation downstream of GLP-1/Notch signaling, preferentially in the GLD-1/NOS-3 pathway. GLD-1 is a KH domain containing protein that has been shown to bind the 3''UTRs of specific transcripts to prevent their translation to promote meiotic entry. These findings lead us to a model in which the proteasome is responsible for degrading proteins that are also regulated by GLD-1 to more efficiently remove them from the cell and promote entry into meiosis as the cells move out of the mitotic zone of the gonad. In order to identify the proteins whose degradation is necessary for proper entry into meiosis we are performing an RNAi screen of all substrate recognition subunits (SRS). These SRS''s are responsible for binding to the specific target protein to allow that protein to be ubiquitinated and subsequently degraded by the proteasome. If we knockdown the function of the SRS, its target protein will not be degraded by the proteasome, resulting in an overproliferation phenotype similar to that seen in the pas-5(oz237) mutant. If we can identify the SRS we can then use biochemical methods to identify the protein that is required for proliferation and whose degradation is required for entry into meiosis.

Lindsay D. MacDonald et al. (2007) International Worm Meeting "Mutation in pas-5, subunit of 20S proteosome, enhances overproliferation in the germline."

Dave Hansen, Lindsay D. MacDonald, Malcolm Williamson, Aaron Knox

[International Worm Meeting, 2007]

The GLP-1/Notch signaling pathway controls the proliferation vs. meiotic entry decision in the germline. GLP-1/Notch signaling promotes proliferation by negatively regulating two redundant downstream pathways (GLD-1/GLD-2), which inhibit proliferation and/or promote meiotic entry. To identify genes that function in the GLD-1 pathway a mutant screen in a gld-2(0) background was performed. The oz237 allele was identified in this screen as being synthetically tumourous with gld-2(0). The oz237 single mutant is sterile (Mog) and has a protruding vulva. We have mapped oz237 to a 40 kB region between lst-1 and tli-1 on Chromosome I. There are 14 genes in this region; we used RNAi and sequenced candidate genes to determine the gene corresponding to the oz237 allele. A mutation was discovered in the coding region of pas-5 (proteasome alpha subunit). An extrachromasomal array containing full length WT pas-5 can partially rescue the sterile phenotype in the oz237 mutants, suggesting the lesion in pas-5 is the oz237 allele. oz237 is likely a partial loss of function allele since RNAi to pas-5 causes embryonic lethality, and oz237/(0) causes a more severe phenotype than seen in the oz237/oz237 mutant worms. To help identify the protein or proteins that are not being degraded properly in the oz237 mutant we first determined if it interacts with other Notch regulated cell fate decisions. pas-5 may act as a general negative regulator of Notch signaling since the oz237 mutant can enhance the multi-vulva phenotype of two weak gain of function mutations of lin-12. We also performed epistasis experiments with other regulators of the proliferation vs. differentiation decision in the germline. Along with being synthetic tumourous with gld-2(0), oz237 is also synthetic tumourous with gld-3(0). When GLP-1 activity is removed from the gld-2 oz237 synthetically tumourous double mutant, the animals are no longer tumourous, but rather have the premature meiotic entry Glp phenotype. This interaction suggests pas-5 may function upstream of Notch signaling. However when GLP-1 activity is removed from the oz237; gld-3 double mutant the germline remains tumourous. This suggests that gld-2 and gld-3 have separate functions in relation to the oz237 mutation when involved in the mitosis vs. meiosis decision in the germline.

Sakoguchi H et al. (2016) Biosci Biotechnol Biochem "Screening of biologically active monosaccharides: growth inhibitory effects of d-allose, d-talose, and l-idose against the nematode Caenorhabditis elegans."

Izumori K, Yoshihara A, Sakoguchi H, Sato M

[Biosci Biotechnol Biochem, 2016]

We compared the growth inhibitory effects of all aldohexose stereoisomers against the model animal Caenorhabditis elegans. Among the tested compounds, the rare sugars d-allose (d-All), d-talose (d-Tal), and l-idose (l-Ido) showed considerable growth inhibition under both monoxenic and axenic culture conditions. 6-Deoxy-d-All had no effect on growth, which suggests that C6-phosphorylation by hexokinase is essential for inhibition by d-All.

Sakoguchi H et al. (2016) Bioorg Med Chem Lett "Growth inhibitory effect of d-arabinose against the nematode Caenorhabditis elegans: Discovery of a novel bioactive monosaccharide."

Shintani T, Izumori K, Sato M, Sakoguchi H, Okuma K, Yoshihara A

[Bioorg Med Chem Lett, 2016]

Biological activities of unusual monosaccharides (rare sugars) have largely remained unstudied until recently. We compared the growth inhibitory effects of aldohexose stereoisomers against the animal model Caenorhabditis elegans cultured in monoxenic conditions with Escherichia coli as food. Among these stereoisomers, the rare sugar d-arabinose (d-Ara) showed particularly strong growth inhibition. The IC50 value for d-Ara was estimated to be 7.5mM, which surpassed that of the potent glycolytic inhibitor 2-deoxy-d-glucose (19.5mM) used as a positive control. The inhibitory effect of d-Ara was also observed in animals cultured in axenic conditions using a chemically defined medium; this excluded the possible influence of E. coli. To our knowledge, this is the first report of biological activity of d-Ara. The d-Ara-induced inhibition was recovered by adding either d-ribose or d-fructose, but not d-glucose. These findings suggest that the inhibition could be induced by multiple mechanisms, for example, disturbance of d-ribose and d-fructose metabolism.

Yoshihara A et al. (2019) Bioorg Med Chem Lett "Growth inhibition by 1-deoxy-d-allulose, a novel bioactive deoxy sugar, screened using Caenorhabditis elegans assay."

Sakoguchi H, Fleet GWJ, Izumori K, Shintani T, Sato M, Yoshihara A

[Bioorg Med Chem Lett, 2019]

The biological activities of deoxy sugars (deoxy monosaccharides) have remained largely unstudied until recently. We compared the growth inhibition by all 1-deoxyketohexoses using the animal model Caenorhabditis elegans. Among the eight stereoisomers, 1-deoxy-d-allulose (1d-d-Alu) showed particularly strong growth inhibition. The 50% inhibition of growth (GI₅₀) concentration by 1d-d-Alu was estimated to be 5.4mM, which is approximately 10 times lower than that of d-allulose (52.7mM), and even lower than that of the potent glycolytic inhibitor, 2-deoxy-d-glucose (19.5mM), implying that 1d-d-Alu has a strong growth inhibition. In contrast, 5-deoxy- and 6-deoxy-d-allulose showed no growth inhibition of C. elegans. The inhibition by 1d-d-Alu was alleviated by the addition of d-ribose or d-fructose. Our findings suggest that 1d-d-Alu-mediated growth inhibition could be induced by the imbalance in d-ribose metabolism. To our knowledge, this is the first report of biological activity of 1d-d-Alu which may be considered as an antimetabolite drug candidate.

Katane M et al. (2020) Biochim Biophys Acta Proteins Proteom "Biochemical characterization of d-aspartate oxidase from Caenorhabditis elegans: Its potential use in the determination of free D-glutamate in biological samples."

Katane M, Sekine M, Nakayama K, Homma H, Kuwabara H, Saitoh Y, Miyamoto T

[Biochim Biophys Acta Proteins Proteom, 2020]

d-Aspartate oxidase (DDO) is a flavin adenine dinucleotide (FAD)-containing flavoprotein that stereospecifically acts on acidic D-amino acids (i.e., free d-aspartate and D-glutamate). Mammalian DDO, which exhibits higher activity toward d-aspartate than D-glutamate, is presumed to regulate levels of d-aspartate in the body and is not thought to degrade D-glutamate in vivo. By contrast, three DDO isoforms are present in the nematode Caenorhabditis elegans, DDO-1, DDO-2, and DDO-3, all of which exhibit substantial activity toward D-glutamate as well as d-aspartate. In this study, we optimized the Escherichia coli culture conditions for production of recombinant C. elegans DDO-1, purified the protein, and showed that it is a flavoprotein with a noncovalently but tightly attached FAD. Furthermore, C. elegans DDO-1, but not mammalian (rat) DDO, efficiently and selectively degraded D-glutamate in addition to d-aspartate, even in the presence of various other amino acids. Thus, C. elegans DDO-1 might be a useful tool for determining these acidic D-amino acids in biological samples.

Shintani T et al. (2019) J Appl Glycosci (1999) "D-Allose, a Stereoisomer of D-Glucose, Extends the Lifespan of Caenorhabditis elegans via Sirtuin and Insulin Signaling."

Izumori K, Sakoguchi H, Yoshihara A, Shintani T, Sato M

[J Appl Glycosci (1999), 2019]

D-Allose (D-All), C-3 epimer of D-glucose, is a rare sugar known to suppress reactive oxygen species generation and prevent hypertension. We previously reported that D-allulose, a structural isomer of D-All, prolongs the lifespan of the nematode Caenorhabditis elegans. Thus, D-All was predicted to affect longevity. In this study, we provide the first empirical evidence that D-All extends the lifespan of C. elegans. Lifespan assays revealed that a lifespan extension was induced by 28 mM D-All. In particular, a lifespan extension of 23.8 % was achieved (p< 0.0001). We further revealed that the effects of D-All on lifespan were dependent on the insulin gene daf-16 and the longevity gene sir-2.1, indicating a distinct mechanism from those of other hexoses, such as D-allulose, with previously reported antiaging effects.

Sato M et al. (2008) J Nat Med "Potential anthelmintic: D-psicose inhibits motility, growth and reproductive maturity of L1 larvae of Caenorhabditis elegans."

Izumori K, Yamasaki T, Kurose H, Sato M

[J Nat Med, 2008]

No anthelmintic sugars have yet been identified. Eight ketohexose stereoisomers (D- and L-forms of psicose, fructose, tagatose and sorbose), along with D-galactose and D-glucose, were examined for potency against L1 stage Caenorhabditis elegans fed Escherichia coli. Of the sugars, D-psicose specifically inhibited the motility, growth and reproductive maturity of the L1 stage. D-Psicose probably interferes with the nematode nutrition. The present results suggest that D-psicose, one of the rare sugars, is a potential anthelmintic.

Yasuaki Saitoh et al. (2010) East Asia Worm Meeting "Study on physiological functions of D-amino acid degradative enzymes in nematode Caenorhabditis elegans"

Yousuke Seida, Kazuhiro Maeda, Tomonori Kawata, Masumi Katane, Hiroyuki Kobuna, Takao Inoue, Yasuaki Saitoh, Hiroyuki Arai, Yasuhito Nakagawa, Masae Sekine, Taro Sakamoto, Hiroshi Homma, Takemitsu Furuchi

[East Asia Worm Meeting, 2010]

Among free D-amino acids existing in living organisms, D-serine (D-Ser) and D-aspartate (D-Asp) are the most actively studied. D-Ser has been proposed as a neuromodulator that regulates L-glutamate-mediated activation of the N-methyl-D-Asp (NMDA) receptor by acting as a co-agonist. On the other hand, several lines of evidence suggest that D-Asp plays important roles in regulating developmental processes, hormone secretion and steroidogenesis. D-Amino acid oxidase (DAO) and D-Asp oxidase (DDO) are known as stereospecific degradative enzymes that catalyze the oxidative deamination of D-amino acids. DAO displays broad substrate specificity and acts on several neutral and basic D-amino acids, while DDO is highly specific for acidic D-amino acids. DAO and DDO are presumed to regulate endogenous D-Ser and D-Asp levels, respectively, as well as mediate the elimination of accumulated exogenous D-amino acids in various organs. Previously, we demonstrated that nematode Caenorhabditis elegans, a multicellular model animal has at least one active DAO gene and three active DDO genes, while it had been thought that most organisms bear only one copy of each DAO and DDO gene. In addition, our previous study revealed that the spatiotemporal distributions of these enzymes in the body of C. elegans are different from one another. In this study, to elucidate the physiological roles of the C. elegans DAO and DDOs, we characterized several phenotypes of four C. elegans mutants in which each gene is partially deleted and inactivated. We also determined free D-amino acid contents in several worm samples using high-performance liquid chromatography (HPLC) techniques. We will report the phenotypes of the C. elegans mutants in comparison with those of wild-type C. elegans, as well as alterations in D-amino acid levels within the body.