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[
International Worm Meeting,
2003]
Autosomal recessive juvenile parkinsonism (AR-JP) is one of the most common forms of familial parkinsons disease characterized by selective loss of dopaminergic neurons in substantia nigra and the locus coeruleus. parkin is the causative gene of AR-JP. The human parkin gene encodes 465 amino acids with a ubiquitin-like domain in the amino-terminus and two RING finger motifs in the carboxy terminus. So far, various deletion mutations and point mutations have been discovered in patients of AR-JP, suggesting that the loss of function of Parkin is the cause of AR-JP. Recently we and others showed that Parkin has a ubiquitin-protein ligase activity and therefore suggested that the defect of protein degradation in the neurons of AR-JP patients (Shimura H. et al. Nat. Genet. 25, 302-5, 2000). To investigate the function of Parkin in vivo, we began to analyze the Ce-PARKIN of C. elegans. Two deletion mutations in parkin genes show no defect in their viabilities. The expression of Ce-PARKIN seems to be specific to subset of neurons. Therefore, we focused on the function of Ce-PARKIN in the neurons and the analysis is underway.
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[
International Worm Meeting,
2013]
The pharyngeal pumping in Caenorhabditis elegans is generated and controlled by the pharyngeal neurons and muscular cells. In this study, we proposed a simulation-based approach to estimate the mechanisms of oscillation generation in pharynx at cell level. To conduct the simulations, we previously developed a pharyngeal muscle model including 20 muscular cell models and 9 marginal cell models (Hattori, Y., et al, Artif. Life Robotics, 17, 2012). Output in each cell model was the membrane potential based on FitzHugh-Nagumo equations. These cell models were connected by gap junctions based on the actual connection structure of pharyngeal muscle in C. elegans. The gap junctions transmitted the outputs between cell models. The electropharyngeogram (EPG), which displays the summed activity of the electrophysiological responses of pharyngeal muscle cells and neurons, was used to measure the biological signals from pharyngeal pumping in C. elegans. In our simulation, we obtained the EPG using the outputs of individual cell models.
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Nitta, N., Matsuda, S., Yanase, S., Sakashita, T., Tamura, M., Suzuki, M., Ishii, N.
[
International Worm Meeting,
2015]
Oxidative stress threatens cellular constructs including lipids, proteins and nucleic acids, and causes or influence apoptosis, degenerative diseases and aging [1]. Recently, we found an abnormal posture in short-lived mutants such as
mev-1 and
daf-16, exposed to hyperoxia. In this study, we detected the hyperoxia-induced abnormal posture in
mev-1 and
daf-16 mutants compared with wild-type N2 and long-lived
age-1 strains. 4-day-old animals placed on a NGM agar plate with a fixed bacterial lawn were exposed to hyperoxia (90% of oxygen) overnight. After the exposure, some animals were immediately recorded as digital images with up to 400-fold magnification of the microscope. The images were analyzed using image-processing software, Wriggle Tracker and Move-tr/2D (Library Co., Ltd., Tokyo), by a previously published method [2]. The body line was skeletonized and evenly divided into 12 segments. Subsequently, X- and Y-coordinates of each point on the body were acquired. To evaluate the posture of the body, we introduced a novel standard, namely the 'posture index' as the absolute average of the relative angle between adjacent dividing points. As a result of this investigation, we can not only intuitively but also quantitatively and statistically understand a tendency of the behavior in various postural responses under stressful conditions in the short-lived mutants.References 1. Yanase S. and Ishii N. (2008) Hyperoxia exposure induced hormesis decreases mitochondrial superoxide radical levels via Ins/IGF-1 signaling pathway in a long-lived
age-1 mutant of Caenorhabditis elegans, J. Radiat. Res. 49: 211-218.2. Hattori Y. et al. (2012) Theoretical and evolutionary parameter tuning of neural oscillators with a double-chain structure for generating rhythmic signals, Neural Compt. 24: 635-675.
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[
International Worm Meeting,
2015]
The pharyngeal pumping in Caenorhabditis elegans is generated and controlled by the pharyngeal muscular cells and neurons. In this study, we proposed a simulation-based approach to estimate the control mechanisms of the pharyngeal pumping motion. To conduct the simulations, we previously developed a pharyngeal muscle model including 20 muscular cells and 9 marginal cells [1]. Output in each cell was represented by the membrane potential based on FitzHugh-Nagumo equations. These cells were connected by gap junctions based on the real connection structure of pharyngeal muscle in C. elegans. The gap junctions transmitted the outputs among cells. The electropharyngeogram (EPG), which shows the summed activity of the electrophysiological responses of pharyngeal muscle cells and neurons, was used to measure the biological signals from pharyngeal pumping in C. elegans. The model represented the EPG using the outputs of individual cells.We virtually input various electrical signals as neuronal signals to muscular cells in the model and calculated the EPG. As a result, some signal patterns induced the EPG which was similar to that observed in wild-type C. elegans. The cycle length of the EPG was synchronized with that of neuronal signals. These results suggest that neuronal signal could control the rhythm of pumping motion. Our approach shows potential for elucidating important signal factors (e.g., amplitude and cycle length) involved in the control of the pumping motion. Through a series of simulations, we confirmed the usefulness of the simulation-based approach for examining the control mechanisms of the pharyngeal pumping motion.[1] Hattori, Y., et al., Modeling of the pharyngeal muscle in Caenorhabditis elegans based on FitzHugh-Nagumo equations, Artificial Life and Robotics, Vol. 17, No. 2, pp. 173-179, 2012.
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[
International Worm Meeting,
2013]
Background and purpose: C. elegans is a good in vivo model system to examine radiobiological effects. We have previously examined the effects of ionizing radiation (IR) on locomotion in C. elegans using 'body bends' (the number of bends counted in the anterior body region at 20-s intervals) [1] and reported an IR-induced reduction in motility [2],[3]. However, the degree of motility in each region of the body has not been established. In the present study, we employed a video-based analysis and investigated the IR-induced effects on locomotion in more detail. Irradiation and video-based analysis: Young adult wild-type C. elegans were placed on a NGM plate with a bacterial lawn (food) and irradiated with graded single doses (0-1.5 kGy) of 60Co g-rays. Immediately after irradiation, animals were transferred to a NGM plate, either with or without food. The movements of five or more animals placed on each plate were video-recorded. The video images were analyzed off-line based on a previously published method [4]. Briefly, after binarization and denoising, the body line was skeletonized and evenly divided into 12 segments; X- and Y-coordinates of each point on the body were subsequently acquired. To evaluate the motility of each point on the body, the moving distance of each of 13 points over a 5 s period was calculated using the X- and Y-coordinates. In addition, we introduced a novel standard, namely the straight distance from head to tail, to evaluate the body form. Results: Under the -food condition, the moving distance of irradiated animals was reduced in a dose-dependent manner at each point on the body, and there was no difference between the effects on each region of the body. The dose-dependent reduction in locomotion was also observed in animals under the +food condition. Furthermore, we evaluated body form and found that IR-induced quantitative changes in body form. References: [1] Sawin, E.R., et al. (2000) Neuron 26, [2] Sakashita, T., et al. (2008) J. Radiat. Res. 49, [3] Suzuki, M., et al. (2009) J. Radiat. Res. 50, [4] Hattori, T., et al. (2012) Neural Comput. 24.
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[
Worm Breeder's Gazette,
1997]
A number of laboratories have recently described various members of a highly conserved serine/threonine protein kinase family that appears to play a role in chromosome segregation and mitotic spindle dynamics in several different organisms (1,2,3,4). A search of the C. elegans genome database revealed that there are at least two nematode proteins that are highly related to this protein kinase family. We refer to these proteins as AIR-1 and AIR-2 (Aurora/Ipl-1 related). To date, the subcellular location of only one member of this protein kinase family has been reported (3,4). The mammalian protein Iak-1 has been shown to be associated with the centrosomes of the mitotic spindle in tissue culture cells and with meiotic chromosomes in mouse spermatocytes (J. Schumacher, unpublished). By performing immunocytochemistry experiments on fixed C. elegans embryos with antisera raised against specific peptides, we have found that the AIR-1 protein, like its mammalian counterpart, is also found on centrosomes in mitotic cells. The protein is first detectable on centrosomes of the first mitotic division following pronuclear fusion in the one-cell embryo. It is clearly associated with duplicated centrosomes prior to their migration to opposite sides of the nucleus and is found on mitotic centrosomes up to the limits of resolution in two-fold stage embryos. The location of the AIR-2 protein mimics that of Iak-1 in meiotic cells. Like Iak-1 in spermatocytes, AIR-2 is also found on chromosomes undergoing meiotic divisions both in oocytes and spermatocytes. AIR-2 staining is diffuse throughout the cellularized oocytes of the proximal gonad, but becomes localized to the chromosomes in the oocyte that resides next to the spermatheca. The protein persists on these chromosomes throughout meiosis and remains associated with polar body chromatin following these divisions. AIR-2 is also found on meiotic chromosomes during spermatogenesis in C. elegans males. In addition, it is associated with mature sperm present in the spermatheca, but at this stage it doesn!t appear to be localized to the chromatin. Instead, it appears to surround the sperm, suggesting an association with the cellular membrane. In embryos, diffuse AIR-2 staining is found in the cytoplasm, but is also clearly localized to mitotic metaphase chromosomes. The protein may be present on chromosomes at other stages of mitosis, but is difficult to detect on less condensed chromatin. By telophase, AIR-2 is clearly localized to midbody microtubules, and a small dot of staining persists on the cell membrane once cytokinesis is complete. To disrupt the function of each of these proteins during embryogenesis, we injected antisense RNA corresponding to the entire cDNA of each gene into the gonads of C. elegans hermaphrodites. Injection of either RNA resulted in embryonic lethality and the specific loss of each protein as detected by immunocytochemistry. AIR-1 deficient embryos die with greater than 100 cells and are severely aneuploid. Analysis of younger embryos revealed a variety of chromosome segregation defects ranging from the loss of a single chromosome to the missegregation of every chromosome to one daughter cell in a particular division. Some cells also appeared to be severely polyploid and contain multiple centrosomes, suggesting multiple cell cycles that lack an intervening mitotic division. Disruption of AIR-2 resulted in the production of one-cell embryos that contained many nuclei and centrosomes, as well as polar bodies that continue to replicate and divide. One-cell embryos containing anywhere from one to greater than 20 nuclei were found with equally abnormal numbers of centrosomes. This dramatic phenotype again suggests the uncoupling of DNA replication and centrosome duplication from the completion of mitosis. References: 1) Chan, C.S. and Botstein, D. (1993). Isolation and characterization of chromosome-gain and increase-in-ploidy mutants in yeast. Genetics 135, 677-691 2) Glover, D. M., Leibowitz, M. H., McLean, D. A., and Parry, H. (1995). Mutations in aurora prevent centrosome separation leading to the formation of monopolar spindles. Cell 81, 95-105. 3) Gopalan, G., Chan C.S., and Donovan P.J. (1997). A novel mammalian, mitotic spindle-associated kinase is related to yeast and fly chromosome segregation regulators. J Cell Biol 138, 643-656 4) Kimura, M., Kotani, S., Hattori, T., Sumi, N., Yoshioka, T., Todokoro, K., and Okano, Y. Cell cycle-dependent expression and spindle pole localization of a novel human protein kinase, Aik, related to Aurora of Drosophila and yeast Ipl1. J Biol Chem 272, 13766-13771 Research sponsored by the National Cancer Institute, DHHS, under contract with ABL.