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[
International Worm Meeting,
2021]
Microbes shape many aspects of host physiology, development and predisposition to disease. Yet the complexity of host-associated communities (microbiomes) makes studying host-microbe interactions challenging. As such, we employed the Caenorhabditis elegans system to investigate molecular mediators of host-microbial interactions with a simplified 12-member microbiome (CeMbio). Previous studies showed the microbiome exerts significant influence on the physiology and development of C. elegans, but lack of molecular tools has limited the ability to identify the microbial factors responsible. To address this challenge, we developed resources to enable genetic manipulation of CeMbio strains. Thus far, we identified several genetic determinants of microbe-microbe interactions, host-specific association and impact on physiology in the dominant microbiome member Ochrobactrum. For genetic manipulation of the CeMbio strains, we utilized broad host range vectors to make an effective panel of fluorescent reporter strains. We used these strains to test pairwise interactions with other microbiome members both in vitro and within the C. elegans gut. Ochrobactrum growth was inhibited by three strains while synergistic with one strain in rich media. In the C. elegans gut, however, microbial interactions were often dramatically different. Notably, Myroides suppresses Ochrobactrum growth in vitro, while Ochrobactrum benefits from the presence of Myroides to colonize the gut. This suggests that host factors may be driving the enrichment for and interactions between members of its microbiome. To examine the molecular determinants of host association and competition, we developed tools for random transposon mutagenesis in Ochrobactrum and several CeMbio strains. We screened a 96-clone mutant library of Ochrobactrum for fitness changes relative to the wild-type in ability to colonize C. elegans hosts with and without Myroides. None of the mutants altered Myroides inhibition of Ochrobactrum in vitro, but several (12) mutants exhibited host association and inter-microbial competition defects. These mutants highlight specific metabolic pathways that Ochrobactrum relies on to colonize and compete for nutrients in association with C. elegans. Our work has demonstrated the ability to use broad host range molecular tools to manipulate CeMbio strains allowing us to visualize and identify molecular mechanisms underlying microbiome assembly and impact upon the C. elegans host.
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[
Elife,
2018]
A diverse array of species on the planet employ the Earth's magnetic field as a navigational aid. As the majority of these animals are migratory, their utility to interrogate the molecular and cellular basis of the magnetic sense is limited. Vidal-Gadea and colleagues recently argued that the worm <i>Caenorhabditis elegans</i> possesses a magnetic sense that guides their vertical movement in soil. In making this claim, they relied on three different behavioral assays that involved magnetic stimuli. Here, we set out to replicate their results employing blinded protocols and double wrapped coils that control for heat generation. We find no evidence supporting the existence of a magnetic sense in <i>C. elegans</i>. We further show that the Vidal-Gadea hypothesis is problematic as the adoption of a correction angle and a fixed trajectory relative to the Earth's magnetic inclination does not necessarily result in vertical movement.
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[
Science,
2000]
Protein interaction mapping using large-scale two-hybrid analysis has been proposed as a way to functionally annotate large numbers of uncharacterized proteins predicted by complete genome sequences. This approach was examined in Caenorhabditis elegans, starting with 27 proteins involved in vulval development. The resulting map reveals both known and new potential interactions and provides a functional annotation for approximately 100 uncharacterized gene products. A protein interaction mapping project is now feasible for C. elegans on a genome-wide scale and should contribute to the understanding of molecular mechanisms in this organism and in human diseases.AD - Massachusetts General Hospital Cancer Center, Charlestown, MA 02129, USA.FAU - Walhout, A JAU - Walhout AJFAU - Sordella, RAU - Sordella RFAU - Lu, XAU - Lu XFAU - Hartley, J LAU - Hartley JLFAU - Temple, G FAU - Temple GFFAU - Brasch, M AAU - Brasch MAFAU - Thierry-Mieg, NAU - Thierry-Mieg NFAU - Vidal, MAU - Vidal MLA - engID - 1 R21 CA81658 A 01/CA/NCIID - 1 RO1 HG01715-01/HG/NHGRIPT - Journal ArticleCY - UNITED STATESTA - ScienceJID - 0404511RN - 0 (Genetic Vectors)RN - 0 (Helminth Proteins)RN - 0 (LIN-35 protein)RN - 0 (LIN-53 protein)RN - 0 (Repressor Proteins)RN - 0 (Retinoblastoma Protein)SB - IM
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[
Worm Breeder's Gazette,
1999]
We are interested in investigating the function of nematode homologues of proteins known to be important for innate immunity in other organisms. Genetic studies with the mouse have shown that a single locus is involved in innate resistance against a range of pathogens including Mycobacterium bovis (BCG), M. intracellulare, M. lepraemurium, Salmonella typhirium and Leishmania. The corresponding gene codes for a protein Nramp1 (Natural resistance-associated macrophage protein; Vidal et al., 1993, Cell, 73, 469-485) that is specifically expressed in macrophages. A highly similar protein, Nramp2, is expressed ubiquitously and appears not to have a role in defense against infection. The Nramp proteins are members of a family of metal ion transporters, that includes the yeast Smf proteins.
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[
J Neurophysiol,
2015]
Although the ability to detect humidity (i.e., hygrosensation) represents an important sensory attribute in many animal species (including humans), the neurophysiological and molecular bases of such sensory ability remain largely unknown in many animals. Recently, Russell and colleagues (Russell J, Vidal-Gadea AG, Makay A, Lanam C, Pierce-Shimomura JT. Proc Natl Acad Sci USA 111: 8269-8274, 2014) provided for the first time neuromolecular evidence for the sensory integration of thermal and mechanical sensory cues which underpin the hygrosensation strategy of an animal (i.e., the free-living roundworm Caenorhabditis elegans) that lacks specific sensory organs for humidity detection (i.e., hygroreceptors). Due to the remarkable similarities in the hygrosensation transduction mechanisms used by hygroreceptor-provided (e.g., insects) and hygroreceptor-lacking species (e.g., roundworms and humans), the findings of Russell et al. highlight potentially universal mechanisms for humidity detection that could be shared across a wide range of species, including humans.
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[
International Worm Meeting,
2019]
Animals change their locomotion or gaits in response to environmental condition. In vertebrates, gait transition has been shown to be mediated by monoamines, which is conserved across many species including the nematode Caenorhabditis elegans (Vidal-Gadea et al., 2011). However, molecular mechanisms of gait transition are still unclear. C. elegans exhibits two gaits, swimming in liquids and crawling on dense gels. C. elegans genome contains evolutionarily conserved 28 DEG/ENaC channels, of which functions may be involved in mechanosensory transduction and locomotion (Goodman and Schwarz, 2003). We first hypothesized that mechanosensitive channels could act as a gait transition initiator and examined crawl-to-swim transition phenotype in DEG/ENaC mutants. We found that while
acd-5 mutants show normal crawling, transition from crawling to swimming upon liquid exposure is defective, suggesting roles of ACD-5 in gait transition. We are currently generating
acd-5 rescue lines and examining expression pattern of
acd-5.
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[
East Coast Worm Meeting,
1998]
The complete genome sequence of C. elegans should be available by the end of this year. It is estimated that the sequence will reveal approximately 13,000 ORFs, most of which correspond to predicted gene-products whose functions have yet to be determined. Hence, in addition to the powerful genetic and biochemical approaches used conventionally, it will be important to develop high-throughput strategies to help addressing the biological role of these gene-products. Since many cellular processes depend on protein-protein interactions, one approach used to elucidate functions consists in identifying physical interactions between proteins. Thus, our long-term goal is to generate a comprehensive protein interaction map for C. elegans, as a way to help characterizing regulatory networks and protein complexes in this model organism (see the abstract by Vidal et al.). We have selected the yeast two-hybrid system to identify potential interactions since on the one hand, its effectiveness has been demonstrated widely, and on the other, it is readily automatable. In this system, interaction between X and Y proteins, expressed as fusions with a DNA binding domain (DB) and an activation domain (AD) respectively, reconstitutes a functional transcription factor which in turn leads to a selectable phenotype. Each potential interaction is referred to as an Interaction Sequence Tag (IST), defined as a pair of short sequences obtained from cDNAs encoding interacting proteins in the two-hybrid system. We intend to release ISTs into AceDB in a format similar to the introduction of ESTs. We will describe the following aspects of the project: -the choice of the first subset of DB-X hybrid proteins used as "baits", -the semi-automated method used to generate the DB-X fusions, -the design and construction of an AD-cDNA library, -the semi-automated two-hybrid selections used to identify ISTs, and -the potential significance of the initial ISTs identified. Since ISTs merely represent hypotheses of interactions, we are also developing high throughput strategies to assess the biological significance of the potential interactions directly in worms (see the abstract by Vidal et al.). The combination of both the IST information available in AceDB and the functional assessment in vivo should be helpful in generating a comprehensive interaction map for C. elegans.
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[
Elife,
2018]
Many animals can orient using the earth's magnetic field. In a recent study, we performed three distinct behavioral assays providing evidence that the nematode <i>Caenorhabditis elegans</i> orients to earth-strength magnetic fields (<xref ref-type="bibr" rid="
bib28">Vidal-Gadea et al., 2015</xref>). A new study by Landler et al. suggests that <i>C. elegans</i> does not orient to magnetic fields (<xref ref-type="bibr" rid="
bib10">Landler et al., 2018</xref>). They also raise conceptual issues that cast doubt on our study. Here, we explain how they appear to have missed positive results in part by omitting controls and running assays longer than prescribed, so that worms switched their preferred migratory direction within single tests. We also highlight differences in experimental methods and interpretations that may explain our different results and conclusions. Together, these findings provide guidance on how to achieve robust magnetotaxis and reinforce our original finding that <i>C. elegans</i> is a suitable model system to study magnetoreception.
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Doucette-Stamm L, Lamesch PE, Reboul J, Temple GF, Hartley JL, Brasch MA, Hill DE, Vaglio P, Thierry-Mieg N, Shin-i T, Lee H, Moore T, Vandenhaute J, Kohara Y, Vidal M, Jackson C, Thierry-Mieg J, Tzellas N, Thierry-Mieg D, Hitti J
[
Nat Genet,
2001]
The genome sequences of Caenorhabditis elegans, Drosophila melanogaster and Arabidopsis thaliana have been predicted to contain 19,000, 13,600 and 25,500 genes, respectively. Before this information can be fully used for evolutionary and functional studies, several issues need to be addressed. First, the gene number estimates obtained in silico and not yet supported by any experimental data need to be verified. For example, it seems biologically paradoxical that C. elegans would have 50% more genes than Drosophilia. Second, intron/exon predictions need to be tested experimentally. Third, complete sets of open reading frames (ORFs), or "ORFeomes," need to be cloned into various expression vectors. To address these issues simultaneously, we have designed and applied to C. elegans the following strategy. Predicted ORFs are amplified by PCR from a highly representative cDNA library using ORF-specific primers, cloned by Gateway recombination cloning and then sequenced to generate ORF sequence tags (OSTs) as a way to verify identity and splicing. In a sample (n=1,222) of the nearly 10,000 genes predicted ab initio (that is, for which no expressed sequence tag (EST) is available so far), at least 70% were verified by OSTs. We also observed that 27% of these experimentally confirmed genes have a structure different from that predicted by GeneFinder. We now have experimental evidence that supports the existence of at least 17,300 genes in C. elegans. Hence we suggest that gene counts based primarily on ESTs may underestimate the number of genes in human and in other organisms.AD - Dana-Farber Cancer Institute and Department of Genetics, Harvard Medical School, Boston, Massachusetts, USA.FAU - Reboul, JAU - Reboul JFAU - Vaglio, PAU - Vaglio PFAU - Tzellas, NAU - Tzellas NFAU - Thierry-Mieg, NAU - Thierry-Mieg NFAU - Moore, TAU - Moore TFAU - Jackson, CAU - Jackson CFAU - Shin-i, TAU - Shin-i TFAU - Kohara, YAU - Kohara YFAU - Thierry-Mieg, DAU - Thierry-Mieg DFAU - Thierry-Mieg, JAU - Thierry-Mieg JFAU - Lee, HAU - Lee HFAU - Hitti, JAU - Hitti JFAU - Doucette-Stamm, LAU - Doucette-Stamm LFAU - Hartley, J LAU - Hartley JLFAU - Temple, G FAU - Temple GFFAU - Brasch, M AAU - Brasch MAFAU - Vandenhaute, JAU - Vandenhaute JFAU - Lamesch, P EAU - Lamesch PEFAU - Hill, D EAU - Hill DEFAU - Vidal, MAU - Vidal MLA - engID - R21 CA81658 A 01/CA/NCIID - RO1 HG01715-01/HG/NHGRIPT - Journal ArticleCY - United StatesTA - Nat GenetJID - 9216904SB - IM
-
[
European Worm Meeting,
2004]
Chromatin-modifying complexes are important for transcriptional control, but their roles in the regulation of development remains poorly understood. We are interested in elucidating how chromatin regulatory complexes function in developmental decisions. The synthetic multivulva (synMuv) genes negatively regulate vulval induction and a sub-set of these genes are components of histone deacetylase (HDAC) complexes (e.g., Rb and NuRD). HDACs are are thought to be recruited to target genes by sequence specific transcription factors, where they locally deacetylate histone H3 tails, and bring about transcriptional repression. We are trying to understand how chromatin-remodelling complexes are integrated into signalling and developmental pathways. To this end, we are currently constructing a C. elegans promoter DNA array which will be used in ChIP-chip (Chromatin immunoprecipitation with DNA Microarray) experiments to identify binding sites for chromatin factors and sequence specific binding proteins. We are spotting ~9000 intergenic PCR fragments generated for the promoterome project in the Vidal Lab. These PCR products are amplified upstream DNA sequences and cover approximately 1.5 - 2.2 kp long sequences from the translation start site. We are also comparing the gene expression profiles of synMuv mutants using the Afffymetrics gene system to look for change of gene expression in these mutants compared to wildtype.