Vancoppenolle B et al. (1996) European Worm Meeting "Comparison of the early embryonic cell lineage of Caenorhabdi- tis elegans and Pellioditis marina, two rhabditid nematodes."

Vancoppenolle B, Coomans AV, Schnabel R, Borgonie G

[European Worm Meeting, 1996]

The early embryonic cell lineage of Pellioditis marina, a marine rhabditid with relatively short developing time was traced using a 4D-microscope. Although the general pattern of cell divisions is congruent with the lineage described for Caenorhabditis elegans by Sulston and coworkers, striking differences can be observed concerning migrations, timing of divisions and cell deaths. The AB, MS and C lineage of P. marina differ from those of C. elegans both in the occurence of additional cell deaths as wel as in the abscence of certain cell deaths. Additionaly, Caap does not divide in accordance with the characteristic period of the rest of the C lineage. In contrast with C. elegans, the E founder cell in P. marina undergoes a migration before gastrulation and devides into Ea and Ep only after E has entered the interior of the embryo. D and P4 divide in a similar way as in C. elegans.

Vancoppenolle B et al. (1997) International C. elegans Meeting "COMPARISON OF THE EMBRYONIC CELL LINEAGE OF CAENORHABDITIS ELEGANS AND PELLIODITIS MARINA"

Borgonie G, Vancoppenolle B, Schnabel R, Coomans AV

[International C. elegans Meeting, 1997]

The early embryonic cell lineage of Pellioditis marina, a marine rhabditid with relatively short developing time (9hrs at 25!C) was traced using a 4D-microscope. Although the general pattern of cell division is congruent with the lineage described for Caenorhabditis elegans by Sulston and Co-workers, striking differences can be observed concerning migrations, timing of divisions and cell deaths. The AB, MS and C lineage of Pellioditis marina differ from those of Caenorhabditis elegans both in the occurence of additional cell deaths as well as in the abscence of certain cell deaths. Additionaly, Caap does not divide in accordance with the characteristic period for the rest of the C-lineage. In contrast with Caenorhabditis elegans, the E founder cell in Pellioditis marina undergoes a migration before gastrulation and divides into Ea and Ep only after E has entered the interior of the embryo. D and P4 divide in a similar way as in Caenorhabditis elegans.

Kuroyanagi, Hidehito et al. (2021) International Worm Meeting "Alternative splicing through m6A modification at a 3' splice site for SAM synthetase homeostasis"

Togo-Ohno, Marina, Fukamizu, Akiyoshi, Watabe, Eichi, KUROYANAGI, Hidehito, Hirota, Keiko, Ishigami, Yuma, Suzuki, Tsutomu, Suzuki, Yutaka

[International Worm Meeting, 2021]

Alternative splicing of precursor messenger RNAs (pre-mRNAs) contributes not only to proteome diversity but also to regulation of gene expression levels by generating mRNA isoforms with a premature termination codon (PTC). Such unproductively spliced mRNAs are unstable and almost undetectable due to an mRNA surveillance system termed nonsense-mediated mRNA decay (NMD). In order to elucidate a repertoire of mRNAs regulated by alternative splicing coupled with NMD (AS-NMD), we performed long-read RNA sequencing of poly(A)+ RNAs from an NMD-deficient mutant, smg-2, and obtained full-length sequences for mRNA isoforms from 259 high-confidence AS-NMD genes. Gene ontology (GO) analysis revealed enrichment of genes related to metabolism in addition to those related to RNA translation and processing. Among them are S-adenosyl-L-methionine (SAM) synthetase (sams) genes. SAM synthetase activity negatively autoregulates sams gene expression through AS-NMD. METT-10, the orthologue of human U6 snRNA methyltransferase METTL16, is required for the splicing regulation of the sams genes in vivo and specifically methylates in vitro the invariant AG dinucleotide at the distal 3' splice site (3'SS) used for the productive mRNAs. RNA immunoprecipitation with anti-m6A antibody and direct RNA sequencing with Nanopore technologies coupled with machine learning confirmed m6A modification of endogenous sams mRNAs. These results indicate that homeostasis of SAM synthetase in C. elegans is maintained by alternative splicing regulation through m6A modification at the 3'SS of the sams genes.

W Houthoofd et al. (2002) European Worm Meeting "Embryonic development of the free-living, marine nematode Pellioditis marina"

G Borgonie, A Coomans, B Vancoppenolle, K Jacobsen, W Houthoofd

[European Worm Meeting, 2002]

Until now only the embryonic cell lineage of the model organism Caenorhabditis elegans has been described (Sulston et al., 1983). The embryonic cell lineage of the free-living nematode Pellioditis marina has been traced from zygote up until the initiation of muscle contraction by means of 4D-microscopy, marking the second detailed description of the embryonic development of a nematode. P. marina is a close relative of C. elegans, but has adapted to a marine, brackish environment. The overall lineage resembles strongly on that of C. elegans, with a few small differences. The developmental tempo of the early embryogenesis (until division of E cell) is more then two times slower than C. elegans. But the primordial germline cell P4 is already present at the 15-cell stage (in C. elegans at the 24-cell stage). At the stage of muscle contraction (when most cells are established), P. marina has as many cells as C. elegans (571 cells) but less cell deaths (67 and 106 respectively). Tissue conservation varies from highly conserved to highly variable. The intestine, the primordial gonad and the body muscles are highly conserved in the two species, while the pharynx, the epidermis and the nervous system have a more variable configuration. The systematic position of Pellioditis remains unsolved, whether Caenorhabditis or Rhabditis is the closest relative. The early embryogenesis and the developmental timing are comparable with that of other Rhabditis species, while the overall cell lineage is almost identical with that of C. elegans. The latter is a strong argument to place P. marina close to C. elegans in the classification. In more primitive nematodes (like Halicephalobus sp.), sublineages form identical cells, which migrate to their exact location. C. elegans has adjusted these lineages to avoid these migrations (Borgonie et al., 2000). This could explain the chaotic' fate topology in the C. elegans cell lineage. P. marina falls in between: it has already adjusted the Caa-lineage to form two nerve cells, but still has migrations that are avoided in C. elegans.

Volker Braun et al. (2001) International C. elegans Meeting "Nematode embryonic cell lineages are computationally efficient"

Gaetan Borgonie, Kim Jacobsen, Armand Leroi, Hans-Peter Meinzer, Paul Agapow, Volker Braun, Wouter Houthoofd, Ricardo Azevedo, Markus Gumbel

[International C. elegans Meeting, 2001]

The fixed cell lineages of nematodes like Caenorhabditis elegans are thought to provide a particularly efficient way to build an organism. However, many aspects of the C.elegans embryonic lineage are not obviously efficient (e.g., the distribution of neurons). Here we test whether the embryonic lineages of three species of rhabditid nematodes, C. elegans, Pellioditis marina and Rhabditophanes sp., are computationally efficient in the way cell fates are specified. We define three measures of cell lineage computational efficiency: number of symmetry breaking events, number of determination events and number of sublineages. First, we find that the actual cell lineages of all species specify most cellular phenotypes, such as cell morphology, function, and position in the hatchling, significantly more efficiently than would be expected if these phenotypes were randomly distributed in the same lineage, regardless of the efficiency measure used. Second, we show that the topologies of the actual lineages, themselves, significantly improve the efficiency of cell fate specification compared to cell lineages with random topologies. Third, we find that the cell lineages of the three species, show comparable levels of computational efficiency, despite considerable differences in topology and cell fates assignments. Our results suggest that the embryonic lineages of rhabditid nematodes evolve to place the right cell in the right place in a computationally efficient way.

Ricardo B R Azevedo et al. (2002) East Coast Worm Meeting "Nematode embryonic cell lineages are computationally efficient"

Markus Gumbel, Ricardo B R Azevedo, Ute Platzer, Hans-Peter Meinzer, Kim Jacobsen, Armand M Leroi, Volker Braun, Gaetan Borgonie, Paul-Michael Agapow, Wouter Houthoofd

[East Coast Worm Meeting, 2002]

What is the minimal amount of information required to specify the cells of a metazoan? Based on ideas from algorithmic information theory and phylogenetics, we develop an algorithm for predicting the distribution of determination events in complete cell lineages. We assume that all such events are either cell autonomous or the outcome of permissive cell-cell interactions, and that the lineage is parsimoniously specified. Applying our algorithm to the complete embryonic lineage of Caenorhabditis elegans, we show that it predicts many known molecular events required to specify cell fates. We then show that less information is required to specify the actual C. elegans lineage than lineages simulated under null models. This is also true for two other species of rhabditid nematode, Pellioditis marina and Rhabditophanes sp., despite many interspecific differences in lineage topology and cell fate assignments. Only one cell fate was found to be inneficiently specified in all species: programmed cell death. Unlike normal cells, most apoptotic cells appear to have no particular function during development. However, we show that the computational efficiency of embryonic development would be increased if cell deaths did not occur all. Thus, selection for increased computational efficiency should lead to a reduction in the number of programmed cell deaths in embryonic cell lineages. Although many programmed cell deaths occur in the C. elegans embryonic lineage (17% of all cells), all of them occur in single-cell monoclones. This is a significantly higher proportion than that expected from permuted lineages and suggests that cell deaths have not accumulated neutrally in the cell lineages of the ancestors of C. elegans. That the absence of cell death monoclones containing two or more cells is due to selection and not due to an intrinsic constraint is demonstrated by the observation that they have been found in other species. Such cases, we suggest, arise frequently, but are then eliminated by reprogramming. Indeed, the main function of somatic cell death in these nematodes might be to eliminate redundant cells over the course of evolution. Our results strongly suggest that selection for computational efficiency moulds the evolution of nematode embryonic cell lineages. But even though nematode lineages are more efficient than random lineages, they are clearly not as efficient as they might be. Why not? The polyclonal origin of some cell fates might be due to the need to generate cells of the same type, such as neurons, in various parts of the embryo. This is supported by the observation that in all species studied here, the majority of cells are born close to their final position in the embryo. We speculate that, in C. elegans, P. marina and Rhabditophanes sp., the fitness cost of repeatedly specifying the same cell type may be less than the cost of additional cell migrations.

Dupuy, Denis (2010) C. elegans: Development and Gene Expression, EMBL, Heidelberg, Germany "High-throughput screening for alternative splicing regulation factors in vivo"

DUPUY, Denis

[C. elegans: Development and Gene Expression, EMBL, Heidelberg, Germany, 2010]

The aim of this our study is to systematically identify the trans-acting elements involved i alternative splicing regulation. Fluorescent reporters provide the opportunity to observe functional effects of a gene knock-down that may have a subtle phenotype that is undetectable with traditional observations. We are using a transgenic strain carrying a two-color reporter system in which two fluorescent reporters (GFP and RFP) are respectively fused to mutually exclusive alternatively spliced exons (Ohno, et al 2008) of the let-2 gene (exons 9 and 10). This strain has been subjected to a genome-wide RNAi screen for modifiers of the alternative splicing ratio. While previous RNAi screens have mostly consisted of visual observation of the worms by microscopy, we use the COPAS-Profiler to perform multidimensional quantitative analysis on the knocked-down worm populations in 96-well format. This enables us to detect not only variation in growth and fertility but also any modification of the relative expression of the two reporters i.e. modification of the alternative splicing balance. In this set up we are able to measure, for each individual RNAi experiment, the total number of worms, their sizes distribution, as well as collect longitudinal expression profiles in two fluorescent channels. Importantly, we are able to compare expression patterns within individual animals, thus providing an endogenous control for stochastic variations in overall expression level.

Ohno, H. et al. (2015) International Worm Meeting "In vivo imaging of diacylglycerol signaling in a taste receptor neuron involved in salt-concentration memory."

Iino, Y., Ohno, H.

[International Worm Meeting, 2015]

Adaptation to environmental conditions by modifying behavior often involves simple forms of learning and memory. We have previously reported that C. elegans worms can memorize ambient salt concentrations and are attracted to the previous salt concentration at which they have been fed, whereas they avoid it if they have been starved (Kunitomo et al., Nat. Commun., 2013; Ohno et al., Science, 2014). This behavioral plasticity has been suggested to be regulated by the diacylglycerol (DAG)/protein kinase C (PKC) pathway acting in a taste receptor neuron, ASER. Genetic manipulations that activate or inactivate this pathway in ASER result in worm migration to higher or lower salt concentrations, respectively (Adachi et al., Genetics, 2010; Kunitomo et al., Nat. Commun., 2013). To explore the role of DAG signaling in salt concentration memory, we have monitored the abundance of DAG in ASER of living worms. When the external salt concentration is increased or decreased from the concentration that has been experienced with food, DAG level is reduced or elevated, respectively, in presynaptic regions or regions adjacent to presynapses, where a novel protein kinase C (nPKC)-epsilon/eta ortholog TTX-4 (a.k.a. PKC-1) is localized. These DAG responses are largely diminished after starvation. These results raise the possibility that the activity of DAG signaling in the presynaptic region of ASER encodes the differences between past and present salt concentrations and thereby forms an integral component of the food-associated memory.

Ohno, Hayao et al. (2017) International Worm Meeting "Diacylglycerol encodes differences between past and current stimulus intensity."

Ohno, Hayao, Adachi, Takeshi, Iino, Yuichi, Sakai, Naoko

[International Worm Meeting, 2017]

C. elegans can memorize external salt concentrations and is attracted to the salt concentration at which it has been fed, whereas it avoids the previous salt concentration if it has been starved (Kunitomo et al., Nat. Commun., 2013; Ohno et al., Science, 2014). An important question is how worms compare the intensity of the past and current sensory stimuli and execute appropriate chemotactic behaviors. The plasticity of salt chemotaxis is regulated by the diacylglycerol (DAG)/protein kinase C (PKC) pathway and the insulin/PI3K pathway, both of which act in the ASER taste receptor neuron. Genetic manipulations that activate or inactivate the DAG/PKC pathway result in worm migration to higher or lower salt concentrations, respectively. The insulin/PI3K pathway is essential for the avoidance of the salt concentrations associated with starvation. We have monitored the abundance of DAG in the ASER presynaptic regions, where the nPKC-epsilon/eta ortholog TTX-4 (a.k.a. PKC-1) is localized. When the external salt concentration is increased or decreased from the concentration that has been experienced with food, DAG level is reduced or elevated, respectively, in a manner dependent on the TAX-4 CNG channel subunit and the EGL-8 phospholipase C beta . These DAG responses are largely diminished after starvation. An unbiased genetic screen and mutant analyses show that the DAG pathway acts downstream of the insulin/PI3K/Akt pathway. Our results suggest that the abundance of DAG in the ASER presynaptic region can encode differences between past and current salt concentrations and the insulin/PI3K pathway regulates the change of salt concentration preferences through modulation of the DAG dynamics.

Nagashima, T. et al. (2017) International Worm Meeting "Multiple isoforms of the DAF-16/FOXO transcription factor regulate taste avoidance learning."

Tomioka, M., Nagashima, T., Iino, Y.

[International Worm Meeting, 2017]

Most Animals can memorize various environmental conditions and alter behaviors according to past experiences. Although signaling pathways underlying learning and memory have been extensively studied using various organisms such as Drosophila and mice, it has been challenging to understand the functional role of signaling pathways in defined neurons. We focus on taste avoidance learning in C. elegans, in which animals learn to avoid salt concentrations encountered under starvation conditions. We have shown that the local action of the insulin-like signaling at the axon of the ASER gustatory neuron regulates taste avoidance learning (Ohno et al., 2014). The insulin-like signaling regulates longevity, development and several other phenomena through control of DAF-16, the FOXO transcription factor homolog. On the other hand, it remains unclear how DAF-16 functions in taste avoidance learning. To clarify the role of DAF-16 in learning and memory, we have analyzed the functional role of DAF-16 in taste avoidance learning. We found that daf-16 loss-of-function mutants show defects in taste avoidance learning: they showed defects in avoidance of high or low salt concentrations after starvation conditioning with high or low salt concentrations, respectively. It was previously reported that DAF-16 has several isoforms with different functions. Behavioral phenotypes of isoform-specific mutants and isoform-specific rescue experiments suggested that at least four DAF-16 isoforms, a-, b-, d- and f-isoforms, were functional in high-salt avoidance, while only the a-isoform of DAF-16 can support low-salt avoidance. Moreover, cell-specific rescue experiments showed that expression of DAF-16 (a-isoform) in the ASER neuron rescued the defect in high-salt avoidance but not low-salt avoidance. These results imply that the functional roles of DAF-16 might vary according to the salt concentrations. We are trying to clarify the molecular mechanism downstream of DAF-16 in taste avoidance learning.