MacGregor D et al. (1993) International C. elegans Meeting "UK: A search for neurotransmitter transporters in C. elegans."

Lee DC, MacGregor D, Coates D, Isaac RE

[International C. elegans Meeting, 1993]

MacGregor D et al. (1995) International C. elegans Meeting "CONTROL OF GENE T23G5.5 EXPRESSION BY UPSTREAM SEQUENCES."

Coates D, Lynch AS, Hope IA, MacGregor D, Isaac RE

[International C. elegans Meeting, 1995]

The gene T23G5.5 shares homology with the family of neurotransmitter transporters which possess 12 trans-membrane regions. This family of genes is highly conserved throughout the animal kingdom. We have created a fusion between a lacZ reporter gene and 6.4kb of DNA upstream of T23G5.5. The fusion occurs in the middle of the ninth predicted exon of T23G5.5. The 6.4kb upstream region also contains the gene T23G5.6 and its upstream region. When injected into C. elegans this construct shows nuclear localized expression in 6 cells of the head ganglia from the L1 stage onwards. A further component manifests itself during L2, when a bilaterally symmetric pair of nuclei in the posterior lateral ganglia begin expression. This pair of neurons can be identified as being either PDEL/R, PDEsoL/R or PVDL/R. Sequential deletion of the upstream region has shown that a 640bp region directly upstream of the initiation codon is sufficient and necessary for the expression pattern. We are currently studying this region in greater depth.

MacGregor D et al. (1997) International C. elegans Meeting "MEMBERS OF THE NEUROTRANSMITTER TRANSPORTER GENE FAMILY IN C. ELEGANS"

James CL, Coates D, MacGregor D, Isaac E

[International C. elegans Meeting, 1997]

The termination of synaptic transmission is often mediated by the action of neurotransmitter transporter proteins. These transporters remove neurotransmitter from the synaptic cleft into pre-synaptic neurons and glial cells. This action is vital for the ordered functioning of the nervous system. The members of two gene families are currently thought to be responsible for this action. A third gene family is responsible for uptake of neurotransmitter into the synaptic vesicles. We are interested in members of the sodium- and chloride-dependent neurotransmitter transporter gene family. These transporters are of particular interest as they are the targets of a number of drugs, such as cocaine and Prozac, which have profound effects on the behaviour of higher mammals. We present here data we have obtained on the cloning of several neurotransmitter transporter protein genes from C. elegans. The genes are T23G5.5, a putative dopamine transporter, T13B5.1, T28F3.? and F55H12.1. All the transporter cDNA!s we have cloned are members of the gene family of sodium- and chloride-dependent neurotransmitter transporters. The members of this family all share a number of motifs; twelve putative trans-membrane domains, a large extracellular loop which contains sites for glycosylation and a conserved motif (GLGNIWRFPXXXYNGGG) between trans-membrane domains 1 and 2, thought to be important for transport function. We present the cDNA sequences of these four transporter genes and highlight the features which suggest they are members of this gene family. We also discuss the genomic structure of these genes and comment on the evolution of this gene family in C. elegans. We also present expression patterns for several of these genes and uptake studies carried out in a COS cell expression system in order to understand the roles these proteins may be playing in the nervous system of C. elegans.

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."

Sato M, Sakoguchi H, Yoshihara A, Izumori K

[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, Yoshihara A, Sato M, Izumori K, Okuma K, Sakoguchi H

[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.

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."

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

[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.

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."

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

[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.

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

[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."

Kurose H, Izumori K, Yamasaki T, 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.

Saitoh, Yasuaki et al. (2013) International Worm Meeting "D-Aspartate oxidase is involved in the caloric restriction-induced lifespan extension in C. elegans."

Arai, Hiroyuki, Furuchi, Takemitsu, Okutsu, Mari, Homma, Hiroshi, Saitoh, Yasuaki, Katane, Masumi, Inoue, Takao, Sekine, Masae, Sakamoto, Taro

[International Worm Meeting, 2013]

Among free D-amino acids existing in living organisms, D-serine (D-Ser) and D-aspartate (D-Asp) are the most intensively studied. In mammals, D-Ser has been proposed as a neuromodulator that regulates L-glutamate (L-Glu)-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 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. Mammalian DAO and DDO are presumed to regulate endogenous D-Ser and D-Asp levels, respectively. Previously, we demonstrated that D-Ser, D-Asp, D-Glu and D-alanine (D-Ala) are present in nematode Caenorhabditis elegans, a multicellular model animal. We also found that C. elegans has at least one active DAO gene and three active DDO genes (DDO-1, DDO-2 and DDO-3), and that the spatiotemporal distributions of these enzymes in the body of C. elegans differ from one another. Furthermore, our previous study showed that alterations of brood size and hatching rate are observed in four C. elegans mutants lacking each gene for the DAO and DDOs. Interestingly, lifespan extension was observed in the DDO-3 mutant. To characterize the mechanism of lifespan extension in the DDO-3 mutant, we performed genetic epistasis experiments to test interactions between the DDO-3 gene and other known longevity pathways. The results suggest that DDO-3 is involved in caloric restriction-induced lifespan extension but not in insulin/IGF signaling pathway, NAD/sir2 pathway nor mitochondrial electron transport system. We also found that D-Glu and L-tryptophan (L-Trp) accumulate throughout life in the DDO-3 mutant. Now we are investigating the relationship between aging and the accumulations of D-Glu and L-Trp.