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1.  GEI-8, a Homologue of Vertebrate Nuclear Receptor Corepressor NCoR/SMRT, Regulates Gonad Development and Neuronal Functions in Caenorhabditis elegans 
PLoS ONE  2013;8(3):e58462.
NCoR and SMRT are two paralogous vertebrate proteins that function as corepressors with unliganded nuclear receptors. Although C. elegans has a large number of nuclear receptors, orthologues of the corepressors NCoR and SMRT have not unambiguously been identified in Drosophila or C. elegans. Here, we identify GEI-8 as the closest homologue of NCoR and SMRT in C. elegans and demonstrate that GEI-8 is expressed as at least two isoforms throughout development in multiple tissues, including neurons, muscle and intestinal cells. We demonstrate that a homozygous deletion within the gei-8 coding region, which is predicted to encode a truncated protein lacking the predicted NR domain, results in severe mutant phenotypes with developmental defects, slow movement and growth, arrested gonadogenesis and defects in cholinergic neurotransmission. Whole genome expression analysis by microarrays identified sets of de-regulated genes consistent with both the observed mutant phenotypes and a role of GEI-8 in regulating transcription. Interestingly, the upregulated transcripts included a predicted mitochondrial sulfide:quinine reductase encoded by Y9C9A.16. This locus also contains non-coding, 21-U RNAs of the piRNA class. Inhibition of the expression of the region coding for 21-U RNAs leads to irregular gonadogenesis in the homozygous gei-8 mutants, but not in an otherwise wild-type background, suggesting that GEI-8 may function in concert with the 21-U RNAs to regulate gonadogenesis. Our results confirm that GEI-8 is the orthologue of the vertebrate NCoR/SMRT corepressors and demonstrate important roles for this putative transcriptional corepressor in development and neuronal function.
doi:10.1371/journal.pone.0058462
PMCID: PMC3590189  PMID: 23484030
2.  Novel structural arrangement of nematode cystathionine β-synthases: characterization of Caenorhabditis elegans CBS-1 
Biochemical Journal  2012;443(Pt 2):535-547.
CBSs (cystathionine β-synthases) are eukaryotic PLP (pyridoxal 5 *-phosphate)-dependent proteins that maintain cellular homocysteine homoeostasis and produce cystathionine and hydrogen sulfide. In the present study, we describe a novel structural arrangement of the CBS enzyme encoded by the cbs-1 gene of the nematode Caenorhabditis elegans. The CBS-1 protein contains a unique tandem repeat of two evolutionarily conserved catalytic regions in a single polypeptide chain. These repeats include a catalytically active C-terminal module containing a PLP-binding site and a less conserved N-terminal module that is unable to bind the PLP cofactor and cannot catalyse CBS reactions, as demonstrated by analysis of truncated variants and active-site mutant proteins. In contrast with other metazoan enzymes, CBS-1 lacks the haem and regulatory Bateman domain essential for activation by AdoMet (S-adenosylmethionine) and only forms monomers. We determined the tissue and subcellular distribution of CBS-1 and showed that cbs-1 knockdown by RNA interference leads to delayed development and to an approximately 10-fold elevation of homocysteine concentrations in nematode extracts. The present study provides the first insight into the metabolism of sulfur amino acids and hydrogen sulfide in C. elegans and shows that nematode CBSs possess a structural feature that is unique among CBS proteins.
doi:10.1042/BJ20111478
PMCID: PMC3316156  PMID: 22240119
cystathionine β-synthase (CBS); Caenorhabditis elegans; domain architecture; homocysteine; hydrogen sulfide; knockdown; AdoMet, S-adenosylmethionine; BN, blue native; BS3, bis(sulfosuccinimidyl) suberate; CBS, cystathionine β-synthase; CGL, cystathionine γ-lyase; DTT, dithiothreitol; EST, expressed sequence tag; GFP, green fluorescent protein; LC–MS/MS, liquid chromatography–tandem MS; PLP, pyridoxal 5*-phosphate; RNAi, RNA interference; RT, reverse transcription; SEC, size-exclusion chromatography; UTR, untranslated region; WT, wild-type
3.  Diversification of fasting regulated transcription in a cluster of duplicated nuclear hormone receptors in C. elegans 
Gene expression patterns : GEP  2010;10(6):227-236.
The genome of C. elegans encodes more than 280 nuclear hormone receptors (NHRs) in contrast to the 48 NHRs in humans and 18 NHRs in Drosophila. The majority of the C. elegans NHRs are categorized as supplementary nuclear receptors (supnrs) that evolved by successive duplications of a single ancestral gene. The evolutionary pressures that lead to the expansion of NHRs in nematodes, as well as the function of the majority of supnrs, are not known. Here, we have studied the expression of seven genes organized in a cluster on chromosome V: nhr-206, nhr-208, nhr-207, nhr-209, nhr-154, nhr-153 and nhr-136. Reverse transcription – quantitative PCR and analyses using transgenic lines carrying GFP fusion genes with their putative promoters revealed that all seven genes of this cluster are expressed and five have partially overlapping expression patterns including in the pharynx, intestine, certain neurons, the anal sphincter muscle, and male specific cells. Four genes in this cluster are conserved between C. elegans and C. briggsae whereas three genes are present only in C. elegans, the apparent result of a relatively recent expansion. Interestingly, we find that a subset of the conserved and non-conserved genes in this cluster respond transcriptionally to fasting in tissue-specific patterns. Our results reveal the diversification of the temporal, spatial, and metabolic gene expression patterns coupled with evolutionary drift within supnr family members.
doi:10.1016/j.gep.2010.05.001
PMCID: PMC2910203  PMID: 20460175
Caenorhabditis elegans; Caenorhabditis briggsae; nuclear hormone receptor; gene expression; fasting
4.  Characterization of gana-1, a Caenorhabditis elegans gene encoding a single ortholog of vertebrate α-galactosidase and α-N-acetylgalactosaminidase 
BMC Cell Biology  2005;6:5.
Background
Human α-galactosidase A (α-GAL) and α-N-acetylgalactosaminidase (α-NAGA) are presumed to share a common ancestor. Deficiencies of these enzymes cause two well-characterized human lysosomal storage disorders (LSD) – Fabry (α-GAL deficiency) and Schindler (α-NAGA deficiency) diseases. Caenorhabditis elegans was previously shown to be a relevant model organism for several late endosomal/lysosomal membrane proteins associated with LSDs. The aim of this study was to identify and characterize C. elegans orthologs to both human lysosomal luminal proteins α-GAL and α-NAGA.
Results
BlastP searches for orthologs of human α-GAL and α-NAGA revealed a single C. elegans gene (R07B7.11) with homology to both human genes (α-galactosidase and α-N-acetylgalactosaminidase) – gana-1. We cloned and sequenced the complete gana-1 cDNA and elucidated the gene organization.
Phylogenetic analyses and homology modeling of GANA-1 based on the 3D structure of chicken α-NAGA, rice α-GAL and human α-GAL suggest a close evolutionary relationship of GANA-1 to both human α-GAL and α-NAGA.
Both α-GAL and α-NAGA enzymatic activities were detected in C. elegans mixed culture homogenates. However, α-GAL activity on an artificial substrate was completely inhibited by the α-NAGA inhibitor, N-acetyl-D-galactosamine.
A GANA-1::GFP fusion protein expressed from a transgene, containing the complete gana-1 coding region and 3 kb of its hypothetical promoter, was not detectable under the standard laboratory conditions. The GFP signal was observed solely in a vesicular compartment of coelomocytes of the animals treated with Concanamycin A (CON A) or NH4Cl, agents that increase the pH of the cellular acidic compartment.
Immunofluorescence detection of the fusion protein using polyclonal anti-GFP antibody showed a broader and coarsely granular cytoplasmic expression pattern in body wall muscle cells, intestinal cells, and a vesicular compartment of coelomocytes.
Inhibition of gana-1 by RNA interference resulted in a decrease of both α-GAL and α-NAGA activities measured in mixed stage culture homogenates but did not cause any obvious phenotype.
Conclusions
GANA-1 is a single C. elegans ortholog of both human α-GAL and α-NAGA proteins. Phylogenetic, homology modeling, biochemical and GFP expression analyses support the hypothesis that GANA-1 has dual enzymatic activity and is localized in an acidic cellular compartment.
doi:10.1186/1471-2121-6-5
PMCID: PMC548690  PMID: 15676072

Results 1-4 (4)