Dietary Restriction Induced Longevity is Mediated by Nuclear Receptor NHR-62 in Caenorhabditis elegans

Dietary restriction (DR) extends lifespan in a wide variety of species, yet the underlying mechanisms are not well understood. Here we show that the C. elegans HNF4a- related nuclear hormone receptor NHR-62 is required for metabolic and physiologic responses associated with DR-induced longevity. nhr-62 mediates the longevity of eat- 2 mutants, a genetic mimetic of dietary restriction, and blunts the longevity response of DR induced by bacterial food dilution at low nutrient levels. Metabolic changes associated with DR, including decreased Oil Red O staining, increased autophagy, and changes in fatty acid composition are partly reversed by mutation of nhr-62. Expression profiles reveal that several hundred genes induced by DR depend on the activity of NHR-62, including a putative lipase required for the DR response. This study provides critical evidence that nuclear hormone receptors regulate the DR response, suggesting hormonal and metabolic control of life span.

Phytoremediation assessment of Gomphrena globosa and Zinnia elegans grown in arsenic-contaminated hydroponic conditions as a safe and feasible alternative to be applied in arsenic-contaminated soils of the Bengal Delta

Several agricultural fields show high contents of arsenic because of irrigation with arsenic- contaminated groundwater. Vegetables accumulate arse- nic in their edible parts when grown in contaminated soils. Polluted vegetables are one of the main sources of arsenic in the food chain, especially for people living in rural arsenic endemic villages of India and Bangladesh. The aim of this study was to assess the feasibility of floriculture in the crop rotation system of arsenic en- demic areas of the Bengal Delta. The effects of different arsenic concentrations (0, 0.5, 1.0, and 2.0 mg As L−1) and types of flowering plant (Gomphrena globosa and Zinnia elegans) on plant growth and arsenic accumula- tion were studied under hydroponic conditions. Total arsenic was quantified using atomic absorption spec- trometer with hydride generation (HG-AAS). Arsenic was mainly accumulated in the roots (72 %), followed by leaves (12 %), stems (10 %), and flowers (<1 %). The flowering plants studied did not show as high phytoremediation capacities as other wild species, suchas ferns. However, they behaved as arsenic tolerant plants and grew and bloomed well, without showing any phytotoxic signs. This study proves that floriculture could be included within the crop rotation system in arsenic-contaminated agricultural soils, in order to im- prove food safety and also food security by increasing farmer’s revenue.

The FLP-side of nematodes.

The central role of FMRFamide-like peptides (FLPs) in nematode motor and sensory capabilities makes FLP signalling an appealing target for new parasiticides. Accumulating evidence has revealed an astounding level of FLP sequence conservation and diversity in the phylum Nematoda, and preliminary work has begun to identify the nematode FLP receptor complement in Caenorhabditis elegans, with a view to investigating their basic biology and therapeutic potential. However, much work is needed to clarify the functional aspects of FLP signalling and how these peptides exert their effects at the organismal level. Here, we summarize our current knowledge of nematode FLP signalling.

The mouse and human IGSF6 (DORA) genes map to the inflammatory bowel disease 1 locus and are embedded in an intron of a gene of unknown function.

We have previously characterized IGSF6 (DORA), a novel member of the immunoglobulin superfamily (IGSF) from human and rat expressed in dendritic and myeloid cells. Using a probe from the open reading frame of the rat cDNA, we isolated a cosmid which contains the entire mouse gene. By comparative analysis and reverse transcriptase polymerase chain reaction, we defined the intron/exon structure and the mRNA of the mouse gene and, with respect to human BAC clones, the human gene. The genes span 10 kb (mouse) and 12 kb (human), with six exons arranged in a manner similar to other members of the IGSF. All intron/exon boundaries follow the GT-AG rule. Expression of the mouse Igsf6 gene is restricted to cells of the immune system, particularly macrophages. Northern blot revealed a single mRNA of 2.5 kb, in contrast to the human gene which is expressed as two mRNAs of 1 and 2.5 kb. The human and mouse genes were localized to a locus associated with inflammatory bowel disease. Analysis of the flanking regions of the Igsf6 gene revealed the presence of an unrelated gene, transcribed from the opposite strand of the DNA and oriented such that the Igsf6 gene is encoded entirely within an intron. An identical organization is seen in human. This gene of unknown function is transcribed and processed, contains homologues in Caenorhabditis elegans and prokaryotes, and is expressed in most organs in the mouse.

Getting to the root of neuronal signalling in plant-parasitic nematodes using RNA interference

A variety of genes expressed in preparasitic second-stage juveniles (J2) of plant-parasitic nematodes appear to be vulnerable to RNA interference (RNAi) in vitro by coupling double-stranded (ds)RNA soaking with the artificial stimulation of pharyngeal pumping. Also, there is mounting evidence that the in planta generation of nematode-specific double-stranded RNAs (dsRNAs) has real utility in the control of these pests. Although neuronally-expressed genes in Caenorhabditis elegans are commonly refractory to RNAi, we have discovered that neuronally-expressed genes in plant-parasitic nematodes are highly susceptible to RNAi and that silencing can be induced by simple soaking procedures without the need for pharyngeal stimulation. Since most front-line anthelmintics that are used for the control of nematode parasites of animals and humans act to disrupt neuromuscular coordination, we argue that intercellular signalling processes associated with neurons have much appeal as targets for transgenic plant-based control strategies for plant-parasitic nematodes. FMRFamide-like peptides (FLPs) are a large family of neuropeptides which are intimately associated with neuromuscular regulation, and our studies on flp gene function in plant-parasitic nematodes have revealed that their expression is central to coordinated locomotory activities. We propose that the high level of conservation in nervous systems across nematodes coupled with the RNAi-susceptibility of neuronally-expressed genes in plant-parasitic nematodes provides a valuable research tool which could be used to interrogate neuronal signalling processes in nematodes.

rbcL analysis supports the tribe Gymnothamnieae Kajimura (Ceramiaceae, Rhodophyta)

The tribe Gymnothamnieae Kajimura was proposed for the monotypic ceramiacean genus Gymnothamnion J. Agardh, previously placed either in the Ptiloteae Cramer or the Antithamnieae Hommersand. A bisporangial isolate of G. elegans (Schousboe ex C. Agardh) J. Agardh from Morocco formed only bisporangia in culture. Its smaller uninucleate cells and sporangia than those of tetrasporophytes suggested that bisporophytes may be haploid as in another member of the Ceramiaceae, Aglaothamnion diaphanum L'Hardy-Halos et Maggs. Phylogenetic analyses of the gene for the large subunit of rubisco (rbcL) from Gymnothamnion and representatives of eight other tribes of the Ceramiaceae confirmed that the removal of Gymnothamnion from the Ptiloteae and the Antithamnieae was warranted. Whereas all tribes with two or more representatives in our analyses were moderately or robustly resolved, Gymnothamnion did not form a strong clade with any other taxa. Analysis of rbcL sequences failed to resolve relationships between tribes, probably due to saturation at the high levels of sequence divergence found. In addition to reproductive features previously reported and interpreted as primitive, G. elegans shows a primitive vegetative feature and it is suggested that Gymnothamnion may be one of the most basal of the taxa presently included in the Ceramiaceae.

Neuropeptide-like protein diversity in phylum Nematoda

This study reports the identification of nematode neuropeptide-like protein (nlp) sequelogs from the GenBank expressed sequence tag (EST) database, using BLAST (Basic Local Alignment Search Tool) search methodology. Search strings derived from peptides encoded by the 45 known Caenorhabatitis elegans nlp genes were used to identify more than 1000 ESTs encoding a total of 26 multi-species nlp sequelogs. The remaining 18 nlps (nlp-4, -16, -24 through -36, -39, -41 and -45) were identified only in C elegans, while the sole EST representative of nlp-23 was from Caenorhabditis remanei. Several ESTs encoding putative antibacterial peptides similar to those encoded by the C elegans genes nlp-24-33 were observed in several parasite species. A novel gene (nlp-46) was identified, encoding a single, amidated dodecapeptide (NIA[I/T]GR[G/A]DG[F/L]RPG) in eight species. Secretory signal peptides were identified in at least one species representing each nlp sequelog, confirming that all 46 nematode nlp genes encode secretory peptides. A random sub-set of C elegans NLPs was tested physiologically in Ascaris suum ovijector and body wall muscle bioassays. None of the peptides tested were able to modulate ovijector activity, while only three displayed measurable myoactivity on somatic body wall muscle. AFAAGWNRamide (from nlp-23) and AVNPFLDSIamide (nlp-3) both produced a relaxation of body wall muscle, while AIPFNGGMYamide (nlp-10) induced a transient contraction. Numerical analyses of nip-encoding ESTs demonstrate that nlp-3, -13, -14, -15 and -18 are amongst the most highly represented transcripts in the dataset. Using available bioinformatics resources, this study delineates the nlp complement of phylum Nematoda, providing a rich source of neuropeptide ligands for deorphanisation of nematode neuropeptide receptors. (C) 2008 Australian Society for Parasitology Inc. Published by Elsevier Ltd. All rights reserved.

Nociception or pain in a decapod crustacean?

Nociception is the ability to perceive a noxious stimulus and react in a re flexive manner and occurs across a wide range of taxa. However, the ability to experience the associated aversive sensation and feeling, known as pain, is not widely accepted to occur in nonvertebrates. We examined the responses of a decapod crustacean, the prawn, Palaemon elegans, to different noxious stimuli applied to one antenna to assess reflex responses (nociception) and longer-term, specifically directed behavioural responses that might indicate pain. We also examined the effects of benzocaine, a local anaesthetic, on these responses. Noxious stimuli elicited an immediate reflex tail flick response, followed by two prolonged activities, grooming of the antenna and rubbing of the antenna against the side of the tank, with both activities directed specifically at the treated antenna. These responses were inhibited by benzocaine; however, benzocaine did not alter general swimming activity and thus the decline in grooming and rubbing is not due to general anaesthesia. Mechanical stimulation by pinching also resulted in prolonged rubbing, but this was not inhibited by benzocaine. These results indicate an awareness of the location of the noxious stimuli, and the prolonged complex responses indicate a central involvement in their organization. The inhibition by a local anaesthetic is similar to observations on vertebrates and is consistent with the idea that these crustaceans can experience pain.

RNAi Effector Diversity in Nematodes

While RNA interference (RNAi) has been deployed to facilitate gene function studies in diverse helminths, parasitic nematodes appear variably susceptible. To test if this is due to inter-species differences in RNAi effector complements, we performed a primary sequence similarity survey for orthologs of 77 Caenorhabditis elegans RNAi pathway proteins in 13 nematode species for which genomic or transcriptomic datasets were available, with all outputs subjected to domain-structure verification. Our dataset spanned transcriptomes of Ancylostoma caninum and Oesophagostomum dentatum, and genomes of Trichinella spiralis, Ascaris suum, Brugia malayi, Haemonchus contortus, Meloidogyne hapla, Meloidogyne incognita and Pristionchus pacificus, as well as the Caenorhabditis species C. brenneri, C. briggsae, C. japonica and C. remanei, and revealed that: (i) Most of the C. elegans proteins responsible for uptake and spread of exogenously applied double stranded (ds)RNA are absent from parasitic species, including RNAi-competent plant-nematodes; (ii) The Argonautes (AGOs) responsible for gene expression regulation in C. elegans are broadly conserved, unlike those recruited during the induction of RNAi by exogenous dsRNA; (iii) Secondary Argonautes (SAGOs) are poorly conserved, and the nuclear AGO NRDE-3 was not identified in any parasite; (iv) All five Caenorhabditis spp. possess an expanded RNAi effector repertoire relative to the parasitic nematodes, consistent with the propensity for gene loss in nematode parasites; (v) In spite of the quantitative differences in RNAi effector complements across nematode species, all displayed qualitatively similar coverage of functional protein groups. In summary, we could not identify RNAi effector deficiencies that associate with reduced susceptibility in parasitic nematodes. Indeed, similarities in the RNAi effector complements of RNAi refractory and competent nematode parasites support the broad applicability of this research genetic tool in nematodes.

An eye on RNAi in nematode parasites

RNA interference (RNAi) has revolutionised approaches to gene function determination. From a parasitology perspective, gene function studies have the added dimension of providing validation data, increasingly deemed essential to the initial phases of drug target selection, pre-screen development. Notionally advantageous to those working on nematode parasites is the fact that Caenorhabditis elegans research spawned RNAi discovery and continues to seed our understanding of its fundamentals. Unfortunately, RNAi data for nematode parasites illustrate variable and inconsistent susceptibilities which undermine confidence and exploitation. Now well-ensconced in an era of nematode parasite genomics, we can begin to unscramble this variation.

FMRFamide-like peptides in root knot nematodes and their potential role in nematode physiology

FMRFamide-like peptides (FLPs) are a diverse group of neuropeptides that are expressed abundantly in nematodes. They exert potent physiological effects on locomotory, feeding and reproductive musculature and also act as neuromodulators. However, little is known about the specific expression patterns and functions of individual peptides. The current study employed rapid amplification of cDNA ends-polymerase chain reaction (RACE-PCR) to characterize flp genes from infective juveniles of the root knot nematodes, Meloidogyne incognita and Meloidogyne minor. The peptides identified from these transcripts are sequelogs of FLPs from the free-living nematode, Caenorhabditis elegans; the genes have therefore been designated as Mi-flp-1, Mi-flp-7, Mi-flp-12, Mm-flp-12 and Mi-flp-14. Mi-flp-1 encodes five FLPs with the common C-terminal moiety, NFLRFamide. Mi-flp-7 encodes two copies of APLDRSALVRFamide and APLDRAAMVRFamide and one copy of APFDRSSMVRFamide. Mi-flp-12 and Mm-flp-12 encode the novel peptide KNNKFEFIRFamide (a longer version of RNKFEFIRFamide found in C. elegans). Mi-flp-14 encodes a single copy of KHEYLRFamide (commonly known as AF2 and regarded as the most abundant nematode FLP), and a single copy of the novel peptide KHEFVRFamide. These FLPs share a high degree of conservation between Meloidogyne species and nematodes from other clades, including those of humans and animals, perhaps suggesting a common neurophysiological role which may be exploited by novel drugs. FLP immunoreactivity was observed for the first time in Meloidogyne, in the circumpharyngeal nerve ring, pharyngeal nerves and ventral nerve cord. Additionally, in situ hybridization revealed Mi-flp-12 expression in an RIR-like neuron and Mi-flp-14 expression in SMB-like neurons, respectively. These localizations imply physiological roles for FLP-12 and FLP-14 peptides, including locomotion and sensory perception.

Exploring the neurotransmitter labyrinth in nematodes

Nematodes include both free-living species such as Caenorhabditis elegans and major parasites of humans, livestock and plants. The apparent simplicity and uniformity of their nervous system belies a rich diversity of putative signalling molecules,particularly neuropeptides. This new appreciation stems largely from the genome-sequencing project with C. elegans, which is due to be completed by the end of 1998. The project has provided additional insights into other aspects of nematode neurobiology, as have studies on the mechanism of action of anthelmintics. Here, progress on the identification, localization, synthesis and physiological actions of transmitters identified in nematodes is explored.

FMRFamide-related peptides (FaRPs) in nematodes: Occurrence and neuromuscular physiology

The occurrence of classical neurotransmitter molecules and numerous peptidic messenger molecules in nematode nervous systems indicate that although structurally simple, nematode nervous systems are chemically complex. Thus far, studies on one nematode neuropeptide family, namely the FMRFamide-related peptides (FaRPs), have revealed an unexpected variety of neuropeptide structures in both free-living and parasitic species. To date 23 nematode FaRPs have been structurally characterized including 12 from Ascaris suum, 8 from Caenorhabditis elegans, 5 from Panagrellus redivivus and 1 from Haemonchus contortus. Ten FaRP-encoding genes have been identified in Caenorhabditis elegans. However, the full complement of nematode neuronal messengers has yet to be described and unidentified nematode FaRPs await detection. Preliminary characterization of the actions of nematode neuropeptides on the somatic musculature and neurones of A. suum has revealed that these peptidic messengers have potent and complex effects. Identified complexities include the biphasic effects of KNEFIRFamide/KHEYLRFamide (AF1/2; relaxation of tone followed by oscillatory contractile activity) and KPNFIRFamide (PF4; rapid relaxation of tone followed by an increase in tone), the diverse actions of KSAYMRFamide (AF8 or PF3; relaxes dorsal muscles and contracts ventral muscles) and the apparent coupling of the relaxatory effects of SDPNFLRFamide/SADPNFLRFamide (PF1/PF2) to nitric oxide release. Indeed, all of the nematode FaRPs which have been tested on somatic muscle strips of A. suum have actions which are clearly physiologically distinguishable. Although we are a very long way from understanding how the actions of these peptides are co-ordinated, not only with those of each other but also with those of the classical transmitter molecules, to control nematode behaviour, their abundance coupled with their diversity of structure and function indicates a hitherto unidentified sophistication to nematode neuromuscular intergration.

KSAYMRFAMIDE - A NOVEL FMRFAMIDE-RELATED HEPTAPEPTIDE FROM THE FREE-LIVING NEMATODE, PANAGRELLUS-REDIVIVUS, WHICH IS MYOACTIVE IN THE PARASITIC NEMATODE, ASCARIS-SUUM

In nematodes, FMRFamide-related peptides (FaRPs) have been structurally characterised from the parasite, Ascaris suum, and from two free-living species, Panagrellus redivivus and Caenorhabditis elegans. While both FaRPs isolated from P. redivivus (PF1 and PF2) have been identified in C. elegans the two heptapeptides isolated from A. suum (AF1 and AF2) have until recently been considered unique to this parasitic species. We have recently isolated AF2 from P. redivivus and, during this study, an additional novel heptapeptide amide, Lys-Ser-Ala-Tyr-Met-Arg-Phe amide (KSAYMRFamide), was structurally characterised. A synthetic replicate of this peptide induced a rapid concentration-dependent muscle tension increase in an isolated A. suum somatic muscle preparation, with a threshold of approximately 0.1 mu M. These data suggest that the complement of FaRPs in parasitic and free-living nematodes may not be as radically different as preliminary studies would suggest, and that the absence of AF1, AF2 and KSAYMRFamide on the C. elegans FMRFamide-related peptide gene (flp-1) may imply the presence of at least two different FaRP genes in nematodes. (C) 1994 Academic Press, Inc.

A quorum-quenching approach to investigate the conservation of quorum-sensing-regulated functions within the Burkholderia cepacia complex

Taxonomic studies of the past few years have shown that the Burkholderia cepacia complex, a heterogeneous group of B. cepacia-like organisms, consists of at least nine species. B. cepacia complex strains are ubiquitously distributed in nature and have been used for biocontrol, bioremediation, and plant growth promotion purposes. At the same time, B. cepacia complex strains have emerged as important opportunistic pathogens of humans, particularly those with cystic fibrosis. All B. cepacia complex species investigated thus far use quorum-sensing (QS) systems that rely on N-acylhomoserine lactone (AHL) signal molecules to express certain functions, including the production of extracellular proteases, swarming motility, biofilm formation, and pathogenicity, in a population-density-dependent manner. In this study we constructed a broad-host-range plasmid that allowed the heterologous expression of the Bacillus sp. strain 240B1 AiiA lactonase, which hydrolyzes the lactone ring of various AHL signal molecules, in all described B. cepacia complex species. We show that expression of AiiA abolished or greatly reduced the accumulation of AHL molecules in the culture supernatants of all tested B. cepacia complex strains. Phenotypic characterization of wild-type and transgenic strains revealed that protease production, swarming motility, biofilm formation, and Caenorhabditis elegans killing efficiency was regulated by AHL in the large majority of strains investigated.