JH Stear et al. (1999) International C. elegans Meeting "Identification of mutants affecting mitotic chromosome segregation in the embryo"

BJ Buchwitz, LL Moore, JH Stear, MB Roth

[International C. elegans Meeting, 1999]

We are interested in characterizing the centromere in C. elegans. As part of this work we have screened a collection of 250 temperature sensitive, embryonic lethal mutants by staining with DAPI. A large percentage (10%) of these mutants displayed alterations in the distribution of DNA in the embryo. We have used high resolution Deltavision microscopy to determine that twelve of these mutants exhibit defects in mitotic chromosome segregation. To further characterize these mutants we stained them with several different centromere antibodies. One class of the mutants shows appropriate individualization of mitotic chromosomes during prophase and congression to the metaphase plate, yet fails to exhibit appropriate deposition of the histone H3 variant ceCENPA. Another class shows defects in the transition from metaphase to anaphase. We have begun to map and clone a subset of these mutants, and are also in the process of performing a saturating screen to isolate additional mutants with similar phenotypes.

Stear JH et al. (2004) Mol Biol Cell "The Caenorhabditis elegans kinetochore reorganizes at prometaphase and in response to checkpoint stimuli."

Roth MB, Stear JH

[Mol Biol Cell, 2004]

Monitoring Editor: Joseph Gall Previous studies of the kinetochore in mammalian systems have demonstrated that this structure undergoes reorganizations following microtubule attachment or in response to activation of the spindle checkpoint. Here we show that the C. elegans kinetochore displays analogous rearrangements at prometaphase, when microtubule/chromosome interactions are being established, and following exposure to checkpoint stimuli such as nocodazole or anoxia. These reorganizations are characterized by a dissociation of several kinetochore proteins, including HCP-1/CeCENP-F, HIM-10/CeNuf2, SAN-1/CeMad3 and CeBUB-1, from the centromere. We further demonstrate that at metaphase, despite having dissociated from the centromere, these reorganized kinetochore proteins maintain their associations with the metaphase plate. Following checkpoint activation, these proteins are detectable as large "flares" that project out laterally from the metaphase plate. Disrupting these gene products via RNAi results in sensitivity to checkpoint stimuli, as well as defects in the organization of chromosomes at metaphase. These phenotypes suggest that these proteins, and by extension their reorganization during mitosis, are important for mediating the checkpoint response as well as directing the assembly of the metaphase plate.

Stear JH et al. (2002) Genes Dev "Characterization of HCP-6, a C. elegans protein required to prevent chromosome twisting and merotelic attachment."

Stear JH, Roth MB

[Genes Dev, 2002]

Previous studies of mitosis show that capture of single kinetochores by microtubules from both centrosomes (merotelic orientation) is a major cause of aneuploidy. We have characterized hcp-6, a temperature-sensitive chromosome segregation mutant in C. elegans that exhibits chromosomes attached to both poles via a single sister kinetochore. We demonstrate that the primary defect in this mutant is a failure to fully condense chromosomes during prophase. Although centromere formation and sister centromere resolution remain unaffected in hcp-6, the chromosomes lack the rigidity of wild-type chromosomes and twist around the long axis of the chromosome. As such, they are unable to establish a proper orientation at prometaphase, allowing individual kinetochores to be captured by microtubules from both poles. We therefore propose that chromosome rigidity plays an essential role in maintaining chromosome orientation to prevent merotelic capture.

Hansen EL et al. (1971) Experientia "Effect of insect hormones on nematodes in axenic culture."

Hansen EL, Buecher EJ

[Experientia, 1971]

Insect juvenile hormones (JH) or JH mimetics have been shown to affect development of nematodes: Trichinella spiralis larvae and fourth stage Phocanema decipiens were inhibited, and abnormal morphology was seen in Heterodera schactii. The effects of insect hormones and analogues on development of several free-living and parasitic nematodes cultured axenically are described in the present paper.

Sim C et al. (2013) Front Physiol "Insulin signaling and the regulation of insect diapause."

Denlinger DL, Sim C

[Front Physiol, 2013]

A rich chapter in the history of insect endocrinology has focused on hormonal control of diapause, especially the major roles played by juvenile hormones (JHs), ecdysteroids, and the neuropeptides that govern JH and ecdysteroid synthesis. More recently, experiments with adult diapause in Drosophila melanogaster and the mosquito Culex pipiens, and pupal diapause in the flesh fly Sarcophaga crassipalpis provide strong evidence that insulin signaling is also an important component of the regulatory pathway leading to the diapause phenotype. Insects produce many different insulin-like peptides (ILPs), and not all are involved in the diapause response; ILP-1 appears to be the one most closely linked to diapause in C. pipiens. Many steps in the pathway leading from perception of daylength (the primary environmental cue used to program diapause) to generation of the diapause phenotype remain unknown, but the role for insulin signaling in mosquito diapause appears to be upstream of JH, as evidenced by the fact that application of exogenous JH can rescue the effects of knocking down expression of ILP-1 or the Insulin Receptor. Fat accumulation, enhancement of stress tolerance, and other features of the diapause phenotype are likely linked to the insulin pathway through the action of a key transcription factor, FOXO. This review highlights many parallels for the role of insulin signaling as a regulator in insect diapause and dauer formation in the nematode Caenorhabditis elegans.

Fodor A et al. (1989) General & Comparative Endocrinology "Effects of precocene analogs on the nematode Caenorhabditis remanei (var. Bangaloreiensis). 2. Competitions with a juvenile hormone analog (methoprene)."

Timar T, Fodor A

[General & Comparative Endocrinology, 1989]

Fourteen 7-alkoxy-2,2-dimethylchromenes were synthetized and studied in JH competition experiments: prococenes (Ps) PI and PII, and synthetic analogs (PAs) including (i) three with both antiallatal and P-like activities: 7-ethoxy-PII (7-EPII); 7-(prop-2-ynyloxy)-2,2-dimethylchromene (PPI); and 6-methoxy-7-(prop-2-yynyloxy)-2,2-dimethylchromene (PPIII); (ii) six without antiallatal activity, exerting P-like activity in nematodes; and (iii) three without either antiallatal or P-like activity, but with a strong nematocidal effect. Within the dose range 8-1000 ug/ml, different concentrations of each PA were applied to nematode growth medium which did or did not contain 1000 ug methoprene (a juvenile hormone analog JHA)/ml. Plates inoculated with Caenorhabditis embryos were incubated and scored for developmentally affected survivors. The JHA did not compete with any PA mentioned as (iii). It competed moderately with some nonantiallatal PAs (8-Me-PPI, 8-MeO-PPI, and 3,4-diCl-PPI) with strong P-like and nematocidal activities. The JHA competed most efficiently with all Ps, antiallatal PAs, and two nonantiallatal PAs (PPII and thio-PI) which exerted severe P-like activities in nematodes. Parameters assumed to be indicators of the P-like (rather than nematocidal) activity of the PAs proved more sensitive to the JHA than those of nematocidal activity. Whether the JH-compensable P-like activity of some PAs can be regarded as a real anti-JH action needs further clarification.

Landis GN et al. (2020) J Gerontol A Biol Sci Med Sci "Metabolic Signatures of Life Span Regulated by Mating, Sex Peptide and Mifepristone/RU486 in Female Drosophila melanogaster."

Abdelmesieh M, Patel P, Wang T, Tower J, Wang L, Fan Y, Promislow DEL, Doherty DV, Lee S, Vroegop J, Wu J, Shen J, Landis GN, Yen CA, Wang I, Curran SP

[J Gerontol A Biol Sci Med Sci, 2020]

Mating and transfer of male Sex Peptide (SP), or transgenic expression of SP, causes inflammation and decreased life span in female Drosophila. Mifepristone rescues these effects, yielding dramatic increases in life span. Here targeted metabolomics data were integrated with further analysis of extant transcriptomic data. Each of seven genes positively correlated with life span were expressed in the brain or eye, and involved regulation of gene expression and signaling. Genes negatively correlated with life span were preferentially expressed in midgut and involved protein degradation, amino acid metabolism, and immune response. Across all conditions, life span was positively correlated with muscle breakdown product 1/3-methylhistadine and purine breakdown product urate, and negatively correlated with tryptophan breakdown product kynurenic acid, suggesting a SP-induced shift from somatic maintenance/turnover pathways to the costly production of energy and lipids from dietary amino acids. Some limited overlap was observed between genes regulated by mifepristone and genes known to be regulated by ecdysone, however, mifepristone was unable to compete with ecdysone for activation of an ecdysone-responsive transgenic reporter. In contrast, genes regulated by mifepristone were highly enriched for genes regulated by Juvenile Hormone (JH), and mifepristone rescued the negative effect of JH analog methoprene on life span in adult virgin females. The data indicate that mifepristone increases life span and decreases inflammation in mated females by antagonizing JH signaling downstream of male SP. Finally, mifepristone increased life span of mated, but not unmated, C. elegans, in two of three trials, suggesting possible evolutionary conservation of mifepristone mechanisms.

Fodor A et al. (1987) Worm Breeder's Gazette "competition of exogenous methoprene and precocenes in nematodes: the end of the story."

Timar T, Hosztafi S, Dinya Z, Fodor A, Kiss I

[Worm Breeder's Gazette, 1987]

We have accomplished a ten year long project aimed to learn whether competition between the juvenile hormone (JH) analogue methoprene (JHA) and precocenes (P's) (chromene derivatives capable to destruct the JH producing organ (CA) in sensitive insect species tissue specifically) in C. elegans (Fodor, et. al., Gen. Comp. Endcr. 46: p. 99 (1982)) can or cannot be explained by a comparable 'anti-JH' action of P's in nematodes. Neither JH or CA like organ has been discovered in nematodes so far. There are only a few indirect data showing that insect juvenile hormones may influence certain nematodes pathogenizing insects. We adopted a 'structure/activity' approach including design, synthesis and test P analogues on nematodes in the presence and absence of JHA. If (at least part of) those analogues which capable to destruct the CA of a sensitive insect (Locusta migratoria) were also effective in nematodes and their effect could be compensated by JHA exogenously, then this hormone (analogue) should play a physiological role in the P-poisoned nematodes. If those P's could be competed by JHA, which proved effective (as 'anti-JH' compounds) in insects, but those which exerted only aspecific toxicity could not be, then it would be logical to suggest, that P's are the same kind of 'suicide compounds' for nematodes as for insects. More than 200 P derivatives were synthesized (Tim r, Hosztafi) and tested on C. elegans (Fodor) and L. migratoria (Kiss). After a detailed quantitative structure/activity relation (QSAR) analysis ( Dinya, et. al., QSAR Strat. Des. Bioact. Compd. Proc. Eur. Symp. Struct.-Act. Relat. 5th (1984) Publ. 1985) several new P analogue were designed, synthesized and tested on L. migratoria and on C. remanei var. Bangaloriensis. (We choose this nematode strain because half of its population consists of males, therefore it is easy to distinguish male adultoids from other type of retarded worms.) Altogether, 121 molecules were retested C. remanei and 17 of them was found to exert some significant biological effect. These compounds were retested again several times both in the absence and in the presence of 1 mg/ml NGM dose of JHA: altogether, more that 144,000 C. remanei embryos were counted, treated and scored afterwards. The tests on nematodes were carried out as described in our attached paper. The most characteristic data concerning precocene activity in nematodes were the following: (1) LC50: the half lethal dose (in g/ml) at which half of the embryos develops to worms (calculated by probit analysis); (2) AD50: the dose ( g/ml) at which half of the embryos develops to normal adults; (3) EC50: the dose ( g/ml) at which half of the nematodes on the plates found as 'normal' fertile adults; (4) The maximum frequency of 'adultoid mini worms' during the experiments. [See Figures 1- 2] The main conclusions are the following: About structure/activity relations: (1) All the three (P1-P3) precocene is effective in nematodes and their effects can be compensated by exogenous JHA. (2) The longer the chain of the R7 substituent the less the effect of the compounds in nematodes. (3) The 7-proparglyoxy analogues are much more effective in nematodes than any other C7 substituted compound. (compare P1 to TT51; P2 to K460; P3 to TT80; TT56 to TT58 or 3,4-diCl-P1 (inactive) to FI121.) (4) The asymmetrically disubstituted analogues are much more effective than the symmetrically disubstituted ones (compare TT80 to K460). It is true, if R7 is longer than R6. (5) Me substitution at C5 position inactivates the originally potent P's (compare TT58 to TT51) but restore the activity of originally inactive (for instance, 7-sBuO-P1) analogues (compare it to TT56). 8-MeO substitution eliminate specific P activity (compare TT51 to K464). (6) Both 8-Me and 8-MeO substitution increase toxic rather than JH compatible biological activity of P's. 8-MeO analogues are more toxic than 8-Me ones, but the consequences of the action of 8-MeO compounds in nematodes can be cured more efficiently by JHA than those concerning 8-Me compounds (compare TT100 to K475). About JHA competition experiments: JHA competed the effects of all precocenes which effected both insects and nematodes. However, the data concerning K354 and FI121 show, that there are analogues which effective only in nematodes and their effects can also be cured by exogenous JHA. Although there are aspecifically toxic analogues (like K454 or 2,3,5-triMe-7 propargO-P1) which cannot be compensated by methoprene, we cannot conclude, that our data unambiguously support the idea of existence JH-like hormones in nematodes. It seems very probable, however, that JH-like compounds can interfere with the lethal metabolism of P's.

Fodor A et al. (1989) Gen Comp Endocrinol "Effects of precocene analogs on the nematode Caenorhabditis remanei (var. Bangaloreiensis). I. Structure/activity relations."

Sebok P, Varga E, Hosztafi S, Fodor A, Timar T, Kiss I, Soos J

[Gen Comp Endocrinol, 1989]

Precocenes (PI and PII) and 114 of their analogs (PAs) were synthetized and tested on C. remanei embryos for their precocene-like (P-like) activities resulting in unusual development at sublethal doses. The P-like activity was quantitated by plotting the probit of the percentage of the developmentally affected survivors against the (log) dose to obtain the EC plot and the half effective concentration (EC50). All five PAs (PI, PII, 7-ethoxy-PII, 7-(prop-2-ynyloxy)-PI, and 6-methoxy-7-(prop-2-ynyloxy)-PII) which exert both antiallatal activity in insects and P-like activity in nematodes are 7-alkoxy-substituted 2,2-dimethylchromenes. Both activities can be enhanced by an additional 6-MeO-substitution or by an asymmetric 6,7-dialkoxy-substitution, on condition that R-7 is longer than R-6. There are many more similarities than dissimilarities in the structural requirements needed for antiallatal and P-like activities. All but three nonantiallatal PAs effective in nematodes are 7-prop-2-ynyloxy-substituted; two are symmetrically 6,7-disubstituted, and one is heterosubstituted (thio-PI). All PAs with antiallatal but without P-like activity are 7-monosubstituted with a relatively long alkoxy group. Certain substitutions favor antiallatal activity and others P-like activity. The severe nematocidal effect of 6,7-methylenedioxy-2,2-dimethylchromene (inert in insects) is not accompanied by P-like activity. The present findings lend some indirect support to the supposition that JH-producing cells and/or JH-dependent function(s) might

Voegtlin, Thomas et al. (2011) International Worm Meeting "An Open-Source Neuromechanical Model of C. Elegans Locomotion."

Cohen, Netta, Voegtlin, Thomas

[International Worm Meeting, 2011]

A neuromechanical model of locomotion in C. elegans was recently proposed by Jordan H. Boyle [1]. One of the main results is that both swimming and crawling can be generated by a single neural circuit, reflexively modulated by the environment. This supports the known experimental results showing that different forms of C. elegans forward locomotion (e.g., swimming and crawling) can be described by a modulation of a single biomechanical gait [2]. The modelling result illustrates the importance and the potential of neuromechanical simulations for the analysis of the worm's behaviour. In order to continue this work, and to make it usable by a broader audience, we have developed a similar neuromechanical model of the worm using CLONES. CLONES (Closed Loop Neural Simulation) is an open source framework for neuromechanical simulations. CLONES implements a communication interface between a neural simulator, called BRIAN [3], and a physics engine for biomedical applications, called SOFA [4]. BRIAN and SOFA are open-source simulators that are easy to use and provide high performance. Our implementation of the worm's locomotion reproduces the neural model described in [1]. However, there are two key differences between the original physical model and our implementation. Firstly, Boyle's model considers that the body of the worm has zero mass (a low Reynolds number approximation). In contrast, the SOFA simulator allows us to integrate equations with mass and inertia. Secondly, the original model uses rigid rods of fixed length orthogonal to the body axis (approximating the incompressibility of the body due to high internal pressure). In SOFA rigid rods are modeled as springs of very high stiffness. The physical system simulated in SOFA is described using a XML syntax. The neural network model interpreted by BRIAN is written in Python, using MATLAB-like syntax. Thus, the model is completely interpreted, and it is possible to visualize/interact with the simulation during runtime. Physical environments containing obstacles or chemical concentration gradients can be defined easily. References 1. Boyle JH: C. elegans locomotion: an integrated approach. PhD thesis, university of Leeds, 2009 2. Berri S, Boyle JH, Tassieri M, Hope IA and Cohen N, Forward locomotion of the nematode C. elegans is achieved through modulation of a single gait HFSP J 3:186, 2009; 3. Goodman DF, Brette R: Brian: a simulator for spiking neural networks in Python. Front Neuroinform 2:5, 2008 4. Allard J, Cotin S, Faure F, Bensoussan PJ, Poyer F, Duriez C, Delingette H, Grisoni L: SOFA - an Open Source Framework for Medical Simulation. Medicine Meets Virtual Reality (MMVR'15), pp. 13-18, 2007.