Watts JL et al. (1993) International C. elegans Meeting "We're still trying to clone par-4."

Watts JL, Kemphues KJ

[International C. elegans Meeting, 1993]

JL Watts et al. (1999) International C. elegans Meeting "Examining the role of polyunsaturated fatty acids in growth and development of C. elegans"

JA Browse, JL Watts

[International C. elegans Meeting, 1999]

Polyunsaturated fatty acids (PUFAs) play important roles in the functioning of membranes and in the synthesis of signaling molecules. C. elegans can synthesize a wide range of polyunsaturated fatty acids using only saturated and monounsaturated fatty acids from E. coli as precursors. In order to study the role of PUFAs in development we have isolated a number of mutant lines exhibiting altered fatty acid compositions. Most of these fatty acid profile differences can be attributed to mutations in known desaturase genes which encode enzymes responsible for inserting double bonds into a fatty acid chain, or in genes involved in the elongation of 18-carbon to 20-carbon fatty acids. One mutant, fat-2 (qa1017) , is defective in D 12-desaturase activity, resulting in a block in the pathway of long chain PUFA biosynthesis. Whereas about 40% of membrane fatty acids in wild-type worms consist of PUFAs, these fatty acids are undetectable in fat-2 worms. The fat-2 mutants can be maintained as a homozygous line, however they grow very slowly, produce small broods, and exhibit a high degree of embryonic lethality. They are also uncoordinated and slightly dumpy. Feeding fat-2 worms small amounts of arachidonic acid rescues these defects, resulting in wild type movement and appearance, greater than 90% hatch rate, and a faster growth rate. We are currently examining the developmental defects of this strain in more detail in order to understand the role of PUFAs in growth, reproduction, and the nervous system of C. elegans .

Watts JL et al. (1991) International C. elegans Meeting "ANALYSIS OF ZYG-11 PROTEIN IN EARLY EMBRYOS OF C. ELEGANS"

Kemphues KJ, Watts JL

[International C. elegans Meeting, 1991]

zyg-11 is a maternally acting gene which is required for embryogenesis in C. elegans (Kemphues et al., Dev. Biol. 113, 449-460, 1986). The zyg-l l gene product is necessary for the proper completion of meiosis as well as for certain cytoplasmic and nuclear events of the first cell cycle. The sequence of zyg-11 does not reveal any significant homologies to other EMBL-GenBank sequences (Carter et al., Mol Gen Genet (1990) 221:72-80). Even though zyg-l 1 mutations show a strict maternal effect, zyg-l 1 RNA is not germline specific, as it is expressed in bn2 worms which lack a germ line. We have raised polyclonal antibodies against a trpE- zyg -11 fusion protein. Our antibody recognizes the fusion protein as well as a prominent band of approximately 90kD in N2 worm and embryo preps, corresponding to the predicted size of the zyg-11 protein. Affinity purified antibody stains the nuclear envelope of early embryos in a cell cycle dependent fashion and also stains muscle in larvae and adult worms. We are currently analyzing the localization and abundance of zyg-l l protein in zyg-ll mutants and bn2 worms.

Amanda Kahn-Kirby et al. (2004) West Coast Worm Meeting "Polyunsaturated Fatty Acids Modulate TRPV-dependent Chemosensory Signaling in vivo"

John Browse, Cornelia I Bargmann, Amanda Kahn-Kirby, Jami LM Dantzker, Jennifer Watts

[West Coast Worm Meeting, 2004]

The C. elegans osm-9 and ocr-2 genes encode predicted cation channels that are essential for proper signal transduction in a subset of chemosensory neurons. osm-9 and ocr-2 are members of the conserved TRP ion channel family; they are most closely related to the mammalian capsaicin receptor TRPV1 and the osmolarity-sensing channel TRPV4. Several mechanisms have been proposed for the intracellular modulation of TRP channels, including facilitation or inhibition by a panoply of lipid molecules. To date, genetic evidence for these models has been relatively scarce. Our in vivo genetic analysis of C. elegans mutants in polyunsaturated fatty acid (PUFA) synthesis pathways suggests a role for specific PUFAs in osm-9 and ocr-2 signal transduction. C. elegans mutants in PUFA synthesis pathways were isolated by a gas chromatography-based screen, as previously described (Watts and Browse 2002). Behavioral analysis established that a subset of mutants deficient in PUFA synthesis were strongly and specifically defective for chemosensory behaviors mediated by osm-9/ocr-2 signaling pathways, notably olfactory signaling in AWA neurons and avoidance signaling in ASH nociceptive neurons. Dietary supplementation with PUFAs resulted in acute rescue of behavioral defects. Direct exposure of animals to exogenous PUFAs resulted in a robust, TRPV-dependent avoidance behavior mediated by ASH, and in TRPV-dependent Ca 2+ transients in the ASH nociceptive neurons. Our behavioral data demonstrates that specific PUFAs contribute to chemosensory neuron function in C. elegans . We suggest that derivatives of these lipids are mobilized in sensory cilia by G-protein signaling pathways to modulate channel activity. Watts and Browse, PNAS 99 (2002): 5854-5859.

Amanda Kahn-Kirby et al. (2003) International Worm Meeting "In vivo genetic evidence for lipid modulation of sensory signal transduction pathways."

Cornelia I Bargmann, Amanda Kahn-Kirby, Jennifer Watts, John Browse

[International Worm Meeting, 2003]

What are the molecular mechanisms used by sensory neurons to detect external stimuli? In the C. elegans AWA and ASH neurons, the osm-9 and ocr-2 genes encode predicted cation channels that are essential for proper sensory signal transduction. osm-9 and ocr-2 are members of the conserved TRP family of sensory channels; they are most closely related to the mammalian channels TRPV1, the capsaicin receptor channel implicated in pain sensation, and the osmolarity-sensing channel TRPV4. Some TRP channels are opened directly by physical signals, whereas others respond to G-protein signaling pathways. The mechanism of TRP activation by G-proteins is unclear. Several mechanisms have been proposed for the intracellular modulation of TRP channels, including facilitation or inhibition by various lipid molecules. To date, genetic evidence for these models has been relatively scarce. Our in vivo genetic analysis of C. elegans mutants in polyunsaturated fatty acid (PUFA) synthesis pathways suggests a role for specific PUFAs in osm-9 and ocr-2 signal transduction. Mutants in PUFA synthesis pathways were isolated by a gas chromatography-based screen, as previously described (Watts and Browse 2002). Behavioral analysis established that fat-3 single mutants, which lack all 20-carbon PUFAs, and a fat-4 fat-1 double mutant, which is deficient for certain 20-carbon PUFAs, were strongly defective for chemotaxis to AWA-sensed odorants, but normal in their chemotaxis to AWC-sensed odorants. AWA, but not AWC, relies on osm-9 for its sensory activity. ASH-mediated avoidance of noxious stimuli, including nose touch and high osmolarity, was also diminished in fat-3 and fat-4 single mutants, and a fat-4 fat-1 double mutant. Dietary supplementation of adult animals with specific PUFAs resulted in acute rescue of the AWA and ASH behavioral defects, suggesting that the mutant phenotypes were not due to developmental abnormalities. Furthermore, direct exposure of animals to specific PUFAs resulted in a TRPV-dependent, ASH-mediated avoidance behavior. Our behavioral data demonstrates that certain PUFAs, including arachidonic acid and eicosapentanoic acid, contribute to AWA and ASH function. We speculate that derivatives of these lipids may be mobilized in sensory cilia by G-protein signaling pathways to modulate channel activity. Watts and Browse, PNAS 99 (2002): 5854-5859.

Watts, Jason et al. (2015) International Worm Meeting "Loss of the C. elegans holocarboxylase synthetase homolog, MEL-3 disrupts anterior-posterior polarity in the embryo and causes larval arrest in a diet dependent manner."

Watts, Jennifer, Kemphues, Kenneth, Watts, Jason, Morton, Diance

[International Worm Meeting, 2015]

The establishment of anterior-posterior polarity occurs rapidly, and is tightly coordinated with meiosis and eggshell formation. The precise timing of the process requires that many of the macromolecules involved must be synthesized and in the correct location prior to fertilization. Lipids play a vital role during the oocyte-to-embryo transition. A precise lipid balance must be maintained to support signaling prior to fertilization as well as membrane structural integrity and eggshell formation after. Here, we demonstrate that MEL-3, the C. elegans homolog to mammalian holocarboxylase synthetase, is an essential player in lipid biosynthesis and in the events following fertilization of the embryo.We found that reduction of MEL-3 function disrupts anterior-posterior polarity and permeability barrier formation in the C. elegans embryo similarly to loss of de novo fatty acid synthesis. We confirmed MEL-3's crucial role in de novo fatty acid synthesis using stable isotope labeling. Further, by genetically manipulating the bacterial food source of C. elegans to limit the dietary fatty acid precursors available to worms, we show that MEL-3 function is essential for larval development when dietary malonyl-CoA is limited.Importantly, we can rescue larval development in mel-3 mutants with supplemental biotin; however, we cannot rescue embryonic phenotypes with biotin or fatty acids. The observation that the mutants can utilize exogenous biotin suggests that they retain some enzymatic function and that the worm can use dietary metabolites to sustain post-embryonic development even when MEL-3 function is reduced, but full function is essential for the embryo. This suggests that the worm diet cannot compensate for either the rapid rate of lipid synthesis required at the transition, or for the specific lipid products produced. We conclude that C. elegans has a specific requirement for fully functioning lipid biosynthesis machinery during the oocyte-to-embryo transition. .

Bhagwati P Gupta et al. (2002) West Coast Worm Meeting "Genetic analysis of the vulval mutants in C. briggsae"

Shahla Gharib, Bhagwati P Gupta, Paul W Sternberg, Jennifer X Li

[West Coast Worm Meeting, 2002]

To understand the evolution of developmental mechanisms, we are doing a comparative analysis of vulval patterning in C. elegans and C. briggsae. C. briggsae is closely related to C. elegans and has identical looking vulval morphology. However, recent studies have indicated subtle differences in the underlying mechanisms of development. The recent completion of C. briggsae genome sequence by the C. elegans Sequencing Consortium is extremely valuable in identifying the conserved genes between C. elegans and C. briggsae.

Oomura, S. et al. (2019) International Worm Meeting "Early embryogenesis of C. inopinata, a sibling species of C.elegans."

Matsumura, Y., Haruta, N., Hoshi, Y., Sugimoto, A., Oomura, S.

[International Worm Meeting, 2019]

C. inopinata is a newly discovered sibling species of C. elegans. Despite their phylogenetic closeness, they have many differences in morphology and ecology. For example, while C. elegans is hermaphroditic, C. inopinata is gonochoristic; C. inopinata is nearly twice as long as C. elegans. A comparative analysis of C. elegans and C. inopinata enables us to study how genomic changes cause these phenotypic differences. In this study, we focused on early embryogenesis of C. inopinata. First, by the microparticle bombardment method we made a C. inopinata line that express GFP::histone in whole body, and compared the early embryogenesis with C. elegans by DIC and fluorescent live imaging. We found that the position of pronuclei and polar bodies were different between these two species. In C. elegans, the female and male pronuclei first become visible in anterior and posterior sides, respectively, then they meet at the center of embryo. On the other hand, the initial position of pronuclei were more closely located in C. inopinata. Also, the polar bodies usually appear in the anterior side of embryo in C. elegans, but they appeared at random positions in C. inopinata. Therefore, we infer that C. inopinata may have a different polarity formation mechanism from that in C. elegans. We are also analyzing temperature dependency of embryogenesis in C. inopinata, whose optimal temperature is ~7 degree higher than that in C. elegans.

Karin Kiontke et al. (2008) Development & Evolution Meeting "New Phylogeny Of Caenorhabditis Including Recently Discovered Species"

David Fitch, Karin Kiontke, Marie-Anne FELIX

[Development & Evolution Meeting, 2008]

Recently, seven new Caenorhabditis have been discovered, bringing the number of Caenorhabditis species in culture to 17, 10 of which are undescribed. To elucidate the relationships of the new species to the five species with sequenced genomes, we have used sequence data from two rRNA genes and several protein-coding genes for reconstructing the phylogenetic tree of Caenorhabditis. Four new species (spp. 5, 9, 10, 11) group within the so-called Elegans group of Caenorhabditis, with C. elegans being the first branch. Whereas none of them is likely to be the sister species of C. elegans, we now know of two close relatives of C. briggsae-C. sp. 5 and C. sp. 9. C. sp. 9 can hybridize with C. briggsae in the laboratory [see abstract by Woodruff et al.]. Of the remaining new species, C. sp. 7 branches off between C. elegans and C. japonica. This species is easier to cultivate than C. japonica and may be a better candidate for comparative experimental work. Two of the new species branch off before C. japonica as sister species of C. sp. 3 and C. drosophilae+C. sp. 2, respectively. Only one of the new species, C. sp. 11, is hermaphroditic. The position of C. sp. 11 in the phylogeny suggests that hermaphroditism evolved three times within the Elegans group. Two of the new species were isolated from rotting leaves and flowers, and five from rotting fruit. Rotting fruit is also the habitat in which C. elegans has been found to proliferate (Barriere and Felix, Genetics 2007) and from which C. briggsae, C. brenneri and C. remanei were repeatedly isolated. This suggests that the habitat of the stem species of Caenorhabditis after the divergence of the earliest branches (C. plicata, C. sonorae and C. sp. 1) was rotting fruit. The rate of discovery of new Caenorhabditis species has steadily increased since the description of C. elegans in 1899, with a leap in the last two years. There is no indication that we are even close to knowing all species in this genus.

Schartner, Caitlin M. et al. (2015) International Worm Meeting "X-chromosome evolution: divergence of X-sequence motifs that drive dosage compensation across Caenorhabditis species."

Meyer, Barbara J., Lo, Te-Wen, Schartner, Caitlin M.

[International Worm Meeting, 2015]

Dosage compensation (DC) across Caenorhabditis species exemplifies an essential process that has undergone rapid co-evolution of protein-DNA interactions central to its mechanism. In C. elegans, recruitment elements on X (rex sites) recruit a condensin-like DC complex (DCC) to hermaphrodite X chromosomes to balance gene expression between the sexes. Recruitment assays in vivo showed that C. elegans rex sites do not recruit the DCC of C. briggsae, and vice versa. To understand how DC complexes and X chromosomes evolved to use different X targeting sequences, we compared DCC subunits and binding sites in C. elegans to those in three species of the C. briggsae clade (15-30 MYR diverged): C. briggsae, its close relative C. nigoni (C. sp. 9), and C. tropicalis (C. sp. 11). By raising antibodies and introducing endogenous tags with TALENs or CRISPR/Cas9, we showed that homologs of both SDC-2, the pivotal X targeting factor, and DPY-27, a DCC-specific condensin subunit, bind X chromosomes of XX animals. Although the DCC shares key components across these four species, the binding sites differ. First, ChIP-seq studies in C. briggsae and C. nigoni identified DCC binding sites that are homologous across these close relatives but differ from C. elegans sites in sequence and location. Second, C. elegans sites use motifs enriched on X (MEX and MEXII) to drive DCC binding, but these motifs are not in C. briggsae or C. nigoni DCC sites and are not X-enriched. Third, we found an X-enriched motif at DCC binding sites of C. briggsae and C. nigoni that is not X-enriched in C. elegans. An oligo with the C. briggsae motif recruits the DCC in C. briggsae, but a similar oligo lacking the motif fails to recruit, establishing the importance of the motif. Fourth, another motif was found in C. briggsae and C. nigoni that shares a few nucleotides with MEX, but its functional divergence was shown by C. elegans recruitment assays. Fifth, two endogenous C. briggsae X-chromosome regions with strong C. elegans MEX motifs fail to recruit the C. briggsae DCC, as assayed by ChIP-seq and recruitment assays. None of these DCC motifs is enriched on the C. tropicalis draft X sequence, supporting further binding site divergence within the C. briggsae clade. Ongoing ChIP-seq studies in C. tropicalis will help determine how C. elegans and C. briggsae clade motifs are evolutionarily related. Comparison of DCC targeting mechanisms across these four species allows us to characterize a rarely captured event: the recent co-evolution of a protein complex and its rapidly diverged target sequences across an entire X chromosome.