Nakajima T et al. (2000) Japanese Worm Meeting "A possible candidate of the DAF-2 ligands: structure, expression, and physiological functions"

Kawano T, Kimura Y, Ito Y, Nakajima T, Ishiguro M, Takuwa K

[Japanese Worm Meeting, 2000]

The DAF-2 protein, a C. elegans homolog of an insulin/IGF-1 receptor, is involved in dauer formation, fat metabolism, and a long life span (1). In the previous meeting, we reported two distinct genes encoding insulin/IGF-1-like peptides designated Ceinsulin-1 and Ceinsulin-2 (2). In this meeting, we will make a short talk of the Ceinsulin-1 regarding its structure, expression, and physiological functions. The precursor protein of Ceinsulin-1 consists of 95 amino acid residues and seems to include a signal peptides, B and A chains, and a so-call C peptide that could be removed out through post-translational proteolysis like the preproinsulin. First, to detect the mature peptide, a polyclonal antibody against the putative B chain was prepared and used for analyzing C. elegans protein by Western blotting. The result showed that the mature Ceinsulin-1 should consist of one polypeptide like IGFs. However, the predicted tertiary structure of Ceinsulin-1 was more similar to the crystal structure of insulin than that of IGF-1, suggesting that the Ceinsulin-1 should be a hybrid molecule of insulin and IGF-1. Next, we analyzed expression pattern of the peptide at several developmental stages using RT-PCR and Western blotting. Interestingly, the transcriptional and translational patterns were not always coincident with each other, suggesting that the peptide biosynthesis could be regulated post-transcriptionally with some degrees. Details will be shown in the meeting. Finally, an RNA-i experiment was carried out to elicit physiological functions of the Ceinsulin-1. The RNAi led to reduction of the corresponding transcript and an extended life span. The inhibition of the Ceinsulin-1 biosynthesis exhibited no drastic effects on entry into and recovery from the dauer stage. Now, the effect on fat accumulation is examined. Anyhow, other insulin/IGF-1 peptides may be involved in the dauer formation and fat metabolism. References 1. Kimura, K.D., et al. (1997) Science 227, 942-946. 2. Kawano, T., et al. (1999) Peptide Science 1998: M. Kondo (Ed.) pp117-120.

Kawano T et al. (1996) Biochem Biophys Res Commun "Molecular cloning of a cDNA for the glutamate transporter of the nematode Caenorhabditis elegans."

Kawano T, Takuwa K, Nakajima T

[Biochem Biophys Res Commun, 1996]

We cloned a cDNA for the glutamate transporter of the nematode Caenorhabditis elegans. The nucleotide sequence and Northern blotting indicated that the glutamate transporter gene is transcribed as a polycistronic mRNA in C. elegans and that subsequent trans-splicing yields two distinct monocistronic mRNAs for the glutamate transporter and the ATP synthase c subunit, respectively. The yields of these monocistronic mRNAs were quite different, suggesting that glutamate transport in C. elegans is regulated in the post-transcriptional phase. The glutamate transporter of C. elegans shows about 50-60% sequence similarity with those of mammals. This is the first description of invertebrate glutamate transporters.

Samuel ADT et al. (2003) J Neurosci "Synaptic activity of the AFD neuron in Caenorhabditis elegans correlates with thermotactic memory."

Murthy VN, Samuel ADT, Silva RA

[J Neurosci, 2003]

Thermotactic behavior in Caenorhabditis elegans is sensitive to both a worm's ambient temperature (T-amb) and its memory of the temperature of its cultivation (T-cult). The AFD neuron is part of a neural circuit that underlies thermotactic behavior. By monitoring the fluorescence of pH-sensitive green fluorescent protein localized to synaptic vesicles, we measured the rate of the synaptic release of AFD in worms cultivated at temperatures between 15 and 25degreesC, and subjected to fixed, ambient temperatures in the same range. We found that the rate of AFD synaptic release is high if either T-amb > T-cult or T-amb > T-cult, but AFD synaptic release is low if T-amb congruent to T-cult. This suggests that AFD encodes a direct comparison between T-amb and T-cult.

Kawano T et al. (1997) Biosci Biotechnol Biochem "Structure and activity of a new form of the glutamate transporter of the nematode Caenorhabditis elegans."

Kawano T, Takuwa K, Nakajima T

[Biosci Biotechnol Biochem, 1997]

A Caenorhabditis elegans cDNA for a glutamate transporter was cloned and examined in this study. The predicted protein is 11 residues shorter at the N-terminus than Ceglut-1, which we previously reported. The protein, when expressed in Xenopus laevis oocytes, showed much higher glutamate transport activity than Ceglut-1, suggesting that the N-terminal sequence is critical in glutamate transport.

Kawano T et al. (1998) Worm Breeder's Gazette "Insulin-like peptides of C. elegan"

Takuwa K, Kimara Y, Nakajima T, Kawano T

[Worm Breeder's Gazette, 1998]

When we searched a peptide homologous with A-chain of Bombyxin a silkworm insulin-like peptide on the C. elegans genome project, we found two species of putativ e peptides that share four cystein residues with Bombyxin A-chain. Then we carried out RACE to obtain the full-length cDNA sequences. The predicted precursor proteins consist of 95 and 108 amino acid residues and contain a signal peptide, B-cahin, C-peptide, and A-chain in the order like those of Bombyxin and human insulin. The deduced mature peptides consist of 50 and 49 residues (named Ceinsulin-1 and Ceinsulin-2) and include six cystein residues identical with those of Bombyxin and human insulin. Ceinsulin-1 A-chain: QLQTICCQVGCNVEDLLAYCAPI B-chain: QQADGRMKMCPPGGSTFTMAWSMSCSM Ceinsulin-2 A-chain: TMNMCCETGCEFTDIFAICNPFG B-chain: AKHGSLKLCPPGGASFLDAFNLICPM Now we try to elicit three dimensional structures of the C. elegans insulin-like peptides using computer chemistry to compare with Bombyxin and human insulin. The GFP fusion proteins will reveal when and where the C. elegans genes encoding insulin-like peptides are expressed

Bommireddy R et al. (2007) Trends Mol Med "TGFbeta1 and Treg cells: alliance for tolerance."

Bommireddy R, Doetschman T

[Trends Mol Med, 2007]

Transforming growth factor beta1 (TGFbeta1), an important pleiotropic, immunoregulatory cytokine, uses distinct signaling mechanisms in lymphocytes to affect T-cell homeostasis, regulatory T (T(reg))-cell and effector-cell function and tumorigenesis. Defects in TGFbeta1 expression or its signaling in T cells correlate with the onset of several autoimmune diseases. TGFbeta1 prevents abnormal T-cell activation through the modulation of Ca(2+)-calcineurin signaling in a Caenorhabditis elegans Sma and Drosophila Mad proteins (SMAD)3 and SMAD4-independent manner; however, in T(reg) cells, its effects are mediated, at least in part, through SMAD signaling. TGFbeta1 also acts as a pro-inflammatory cytokine and induces interleukin (IL)-17-producing pathogenic T-helper cells (T(h) IL-17 cells) synergistically during an inflammatory response in which IL-6 is produced. Here, we will review TGFbeta1 and its signaling in T cells with an emphasis on the regulatory arm of immune tolerance.

Agulnik SI et al. (1995) Genomics "Conservation of the T-box gene family from Mus musculus to Caenorhabditis elegans."

Bollag RJ, Silver LM, Agulnik SI

[Genomics, 1995]

Recently, a novel family of genes with a region of homology to the mouse T locus, which is known to play a crucial, and conserved, role in vertebrate development, has been discovered. The region of homology has been named the T-box. The T-box domain of the prototypical T locus product is associated with sequence-specific DNA binding activity. In this report, we have characterized four members of the T-box gene family from the nematode Caenorhabditis elegans. All lie in close proximity to each other in the middle of chromosome III. Homology analysis among all completely sequenced T-box products indicates a larger size for the conserved T-box domain (166 to 203 residues) than previously reported. Phylogenetic analysis suggests that one C. elegans T-box gene may be a direct ortholog of the mouse Tbx2 and Drosophila omb genes. The accumulated data demonstrate the ancient nature of the T-box gene family and suggest the existence of at least three separate T-box-containing genes in a common early metazoan ancestor to nematodes and vertebrates.

Ju T et al. (2006) Glycobiology "Identification of core 1 O-glycan T-synthase from Caenorhabditis elegans."

Ju T, Zheng Q, Cummings RD

[Glycobiology, 2006]

The common O-glycan core structure in animal glycoproteins is the core 1 disaccharide Galbeta1-3GalNAcalpha1-Ser/Thr, which is generated by addition of Gal to GalNAcalpha1-Ser/Thr by core 1 UDP-Gal:GalNAcalpha1-Ser/Thr beta1,3-galactosyltransferase (core 1 beta3-Gal-T or T-synthase, EC2.4.1.122)(2). Although O-glycans play important roles in vertebrates, much remains to be learned from model organisms such as the free-living nematode Caenorhabditis elegans, which offer many advantages in exploring O-glycan structure/function. Here we report the cloning and enzymatic characterization of T-synthase from C. elegans (Ce-T-synthase). A putative C. elegans gene for T-synthase, C38H2.2, was identified in GenBank by a BlastP search using the human T-synthase protein sequence. The full-length cDNA for Ce-T-synthase, which was generated by PCR using a C. elegans cDNA library as the template, contains 1,170 bp including the stop TAA. The cDNA encodes a protein of 389 amino acids with typical type-II membrane topology and a remarkable 42.7% identity to the human T-synthase. Ce-T-synthase has 7 Cys residues in the lumenal domain including 6 conserved Cys residues in all of the orthologs. The Ce-T-synthase has 4 potential N-glycosylation sequons, whereas the mammalian orthologs lack N-glycosylation sequons. Only one gene for Ce-T-synthase was identified in the genome-wide search and it contains 8 exons. Promoter analysis of the Ce-T-synthase using green fluorescent protein constructs show that the gene is expressed at all developmental stages and appears to be in all cells. Unexpectedly, only minimal activity was recovered in the recombinant, soluble Ce-T-synthase secreted from a wide variety of mammalian cell lines, whereas robust enzyme activity was recovered in the soluble Ce-T-synthase expressed in Hi-5 insect cells. Vertebrate T-synthase requires the molecular chaperone Cosmc, but our results show that Ce-T-synthase does not require Cosmc, and might require invertebrate-specific factors for formation of the optimally active enzyme. These results show that the Ce-T-synthase is a functional ortholog to the human T-synthase in generating core 1 O-glycans and opens new avenues to explore O-glycan function in this model organism.

Yoshida E et al. (1998) Gene "Identification of N-terminal minimal transactivation domain of CBP, p300 and caenorhabditis elegans homologues."

Nakajima T, Fukamizu A, Murakami K, Yoshida E

[Gene, 1998]

CBP/p300 is a multidomain transcriptional cofactor that acts in junction with other factors to regulate transcription. To elucidate the domain function of CBP, we fused its dissected fragments to Gal4 DNA-binding domain and transfected the deletion mutants into several lines. First, we found that the minimal transactivation domain (MTD) at the N-terminal portion maps to between 344 and 451 aa, and shows activity in a cell-type dependent manner. Second, we cloned C. elegans homologues corresponding to the MTD by RT-PCR and identified the three related products, two of which exhibited weak transcriptional activity. Finally, by means of the yeast two hybrid screening using MTD as a bait, we cloned hypoxia-inducible factor (HIF) 1a and Stat2 cDNAs. These results suggested a functional role of MTD located at the N-terminal region of CBP/p300 in connecting to transcriptional factors.

Muir RE et al. (2007) Int J Syst Evol Microbiol "Leucobacter chromiireducens subsp. solipictus subsp. nov., a pigmented bacterium isolated from the nematode Caenorhabditis elegans, and emended description of L. chromiireducens."

Muir RE, Tan MW

[Int J Syst Evol Microbiol, 2007]

A yellow-pigmented, Gram-positive, aerobic, non-motile, non-spore-forming, irregular rod-shaped bacterium (strain TAN 31504(T)) was isolated from the bacteriophagous nematode Caenorhabditis elegans. Based on 16S rRNA gene sequence similarity, DNA G+C content of 69.5 mol%, 2,4-diaminobutyric acid in the cell-wall peptidoglycan, major menaquinone MK-11, abundance of anteiso- and iso-fatty acids, polar lipids diphosphatidylglycerol and phosphatidylglycerol and a number of shared biochemical characteristics, strain TAN 31504(T) was placed in the genus Leucobacter. DNA-DNA hybridization comparisons demonstrated a 91 % DNA-DNA relatedness between strain TAN 31504(T) and Leucobacter chromiireducens LMG 22506(T) indicating that these two strains belong to the same species, when the recommended threshold value of 70 % DNA-DNA relatedness for the definition of a bacterial species by the ad hoc committee on reconciliation of approaches to bacterial systematics is considered. Based on distinct differences in morphology, physiology, chemotaxonomic markers and various biochemical characteristics, it is proposed to split the species L. chromiireducens into two novel subspecies, Leucobacter chromiireducens subsp. chromiireducens subsp. nov. (type strain L-1(T)=CIP 108389(T)=LMG 22506(T)) and Leucobacter chromiireducens subsp. solipictus subsp. nov. (type strain TAN 31504(T)=DSM 18340(T)=ATCC BAA-1336(T)).