7 Papers found (0.009 seconds)

Christopher J Cronin et al. (2005) International Worm Meeting "The Combined UC San Diego/Caltech Quantitative Behavior Analysis System"

William R Schafer, Zhaoyang Feng, Kuang-man Huang, Paul W Sternberg, Christopher J Cronin

[

International Worm Meeting,

2005]

California Institute of Technology and University of California, San Diego have each developed practical, automated movement analysis systems to quantify aspects of worm behavior and morphology (see An imaging system for standardized quantitative analysis of C. elegans behavior, Zhaoyang Feng, et al, BMC Bioinformatics 2004, 5:115, An automated system for measuring parameters of nematode sinusoidal movement, Christopher J Cronin, et al, BMC Genetics 2005, 6:5, and previous meeting abstracts). We have consolidated our development efforts and merged our independent system designs to create a single, unified behavior analysis system. The combined system provides researchers with all of the quantification tools and database functionality of the individual UCSD and Caltech systems, but also establishes a common, open software and hardware foundation for continued development by Caltech, UCSD and the community. Current efforts include expanding system capabilities for the automated analysis of mating behavior with the challenge of tracking multiple moving individuals, both in contact with and separated from each other.

Kamal, Muntasir et al. (2021) International Worm Meeting "The Alimentary Cuticle of C. elegans Plays Multiple Roles in Mediating Xenobiotic Sensitivity"

Cummins, Carolyn, Kamal, Muntasir, Nguyen, Ken, Yip, Christopher, Knox, Jessica, Yeo, May, Roy, Peter, Szlapa, Karolina, Moshiri, Houtan, Won, Amy, Magomedova, Lilia, Hall, David, Bagg, Rachel, Han, Duhyun

[

International Worm Meeting,

2021]

Forward genetic screens with small molecules (SMs) in C. elegans carried out in our and other laboratories have repeatedly shown that some SMs aggregate in the anterior pharynx of worms as dark objects. Our group first identified the accumulation as at least partly, if not wholly composed of SM itself manifest as birefringent crystals and/or globular spherical objects. The crystallizing molecules are all lethal to worms. Our data indicates that the process of SM crystallization begins in the pharyngeal cuticle of the anterior pharynx, and over time, extends to the plasma membrane, ultimately causing extensive cellular damage. The crystallizing molecules can be grouped into 24 distinct structurally similar families (called scaffolds), and tend to be generally more hydrophobic and polar than molecules that do not aggregate. To understand the molecular basis of crystal accumulation, we screened 1.3 million randomly mutagenized wild-type F2 genomes and isolated 46 mutants that resist the lethal effects of a crystallizing SM, wact-190. This uncovered loss or reduction-of-function mutations in genes encoding components of a putative sphingomyelin synthesis pathway, SMS-5, TTM-5 and SPTL-2. Additionally, loss or reduction-of-function mutations in the ABC transporter PGP-14 also confer robust resistance to all crystallizing SMs. sms-5 and pgp-14 are moderately and highly expressed, respectively, in pharyngeal cells that line the pharynx cuticle. Both sms-5 and pgp-14 mutants show reduced accumulation of the hydrophobic, crystallizing SMs. Interestingly, both mutants show higher accumulation and sensitivity to a different set of lethal SMs with different physical properties relative to the wildtype. Together, these results indicate that (1) the anterior pharyngeal cuticle may be an important route of SM entry and a "sink" for many hydrophobic molecules, and (2) loss of SMS-5 or PGP-14 activity leads to altered barrier properties. Remarkably, sms-5 mutants are hypersensitive to cholesterol-deprived conditions, suggesting sms-5 is specifically required for accumulation of hydrophobic nutrients and other SMs in the pharynx cuticle and in turn into pharyngeal cells. PGP-14 maybe required for transporting a hydrophobic molecule related to SMS-5-catalyzed polar lipid synthesis into the developing cuticle. Taken together, our data suggest that SMS-5 and PGP-14 contribute to a lipid barrier in the anterior pharynx cuticle to limit access of small molecules to the animal.

Pena-Gonzalez, I. et al. (2013) International Worm Meeting "Suppressors of TDP-1 toxicity in Caenorhadbitis elegans."

Link, CD., Pena-Gonzalez, I.

[

International Worm Meeting,

2013]

Suppressors of TDP-1 toxicity in Caenorhadbitis elegans Ilana Pena-Gonzalez1 and Christopher D Link1, 2 1 Integrative Physiology, University of Colorado, Boulder, CO 80309, USA 2 Institute for Behavioral Genetics, University of Colorado, Boulder, CO 80309, USA RNA binding protein TDP-43 forms damaging aggregates in multiple neurodegenerative diseases. We have found that the deletion of tdp-1, the C. elegans ortholog of TDP-43, leads to increased accumulation of double stranded RNA. TDP-1 is not believed to bind to dsRNA itself; however the marked increase in dsRNA accumulation in worms deleted for tdp-1 implies TDP-1 normally participates in limiting the stability and structure of dsRNA. Although worms deleted for TDP-1 do not have a severe phenotype, overexpressed nuclear TDP-1 is toxic in worms. One possible explanation for this is that too much TDP-1 leads to excessive disruption of structured RNAs needed for normal RNA metabolism. Making use of this overexpression model we have established a heat shock induction system to help identify components associated with TDP-1's prevention of dsRNA accumulation. Identifying mutations that suppress the neurotoxic effects of overexpressed TDP-1 could help identify others factors contributing to the complex mechanism of disassembling dsRNA. A mutagenesis screen has identified four mutant strains that partially suppress the toxicity of overexpressed TDP-1. We have also tested suppressors of other toxic proteins. Further identifying the pieces of the mechanism regulating dsRNA breakdown could help better explain the pathogenic pathway of TDP-1 and consequently TDP-43 in disease.

Desse, V. et al. (2019) International Worm Meeting "SAX-7/L1CAM in the maintenance of neuronal architecture."

Perrat, P., Blanchette, C., Desse, V., Benard, C.

[

International Worm Meeting,

2019]

Whereas remarkable advances have been made in understanding neuronal development, the mechanisms that maintain the nervous system architecture lifelong are poorly understood. How does the nervous system established during embryogenesis subsequently maintain its integrity throughout life, despite the animal's growth, the addition of new neurons, maturation processes, body movements, and aging? The evolutionarily conserved gene sax-7, homologous to mammalian L1CAM, is one of the factors required for maintaining the structural integrity of ganglia and fascicles in C. elegans1. Thus, sax-7 not only contributes to nervous system development (roles in guidance and fasciculation of a subset of developing neurites)2, but is also required for maintaining the organization of neural architecture after its initial development. Indeed, some neural structures that initially develop normally become disorganized in animals lacking the function of sax-7. With heat shock assays, we show that expression of sax-7S(+) during larval stages is sufficient to rescue the maintenance defects of sax-7 mutants that were devoid of functional sax-7 during nervous system development. We are characterizing a true null allele (generated using CRISPR/Cas9), as well as the expression pattern of a functional SAX-7S::sfGFP (insertion in the endogenous locus using CRISPR/Cas9), and we find that SAX-7S is present in neurons, at the plasma membrane. We are exploring the structure-function relationship of SAX-7S, a transmembrane protein harboring extracellular Ig and FnIII domains, in the context of maintenance of the nervous system architecture. A conditional knockout of L1CAM in the adult mouse brain results in behavioral and learning defects, indicating that L1CAM plays a post-developmental role in mammals as well. Our studies are expected to help understand the role of SAX-7/L1CAM in conserved molecular mechanisms ensuring neuronal maintenance, which may go awry in neurodegenerative conditions. 1 Pocock, Benard et al 2008; Sasakura et al 2005; Wang, Chen et al 2005; Zallen et al 1999; Ramirez-Suarez et al 2019 2 Diaz-Balzac et al 2015; Salzberg et al 2013; Yip and Heiman 2018, Dong et al 2013; Zhu et al 2017

Gattegno, Tamar et al. (2013) International Worm Meeting "Menorah-menorah auto-fusion as a mechanism of PVD response to injury."

Gattegno, Tamar, Podbilewicz, Benjamin, Oren, Meital

[

International Worm Meeting,

2013]

The PVDs are two nociceptors localized along C. elegans lateral sides, enabling them to sense strong mechanical stimuli such as pain, temperature and posture. The PVDs develop through a dynamic process which involves extension and retraction of dendrite branches. Though we now better understand C. elegans PVD development, very little is known about PVD regeneration and degeneration following injury. In order to study the genetic pathway guiding PVD response to injury, we use laser microsurgery to severe the PVD and live imaging of GFP reporters. We found that when the PVDs are injured at the 1ry branch, AFF-1 mediated fusion occurs between the terminal branches to bypass the site of injury, followed by EFF-1 mediated pruning and arbor simplification. Degeneration of the distal stump occurred in animals that failed to undergo reconnection of the 1ry severed branch or menorah-menorah fusion. We used Kaede photo-convertible fluorescent protein expressed in the PVD to support the observation that injured PVD has reconnected through auto-fusion. We severed the PVD 1ry branch, recovered the worms and detected reconnection after 6-12 hours. The PVD cell body of recovered animals was than photo-converted from green to red and we analyzed the transfer of the Red-Kaede through the reconnected neuron. We found that PVD reconnection occurs through fusion. We then performed a similar experiment in lin-22 animals which have 2 to 5 extra PVD cell bodies (kindly provided by M. Heiman and C. Yip) to study whether adjacent PVDs connect by fusion. We found that following injury different PVDs can fuse as a response to injury. These experiments show that PVD neurons response to dendrotomy involves four steps: (1) Loss of self-avoidance or tiling. (2) Menorah-menorah fusion mediated by AFF-1 and/or reconnection of injured 1ry branches by auto-fusion. (3) Ectopic sprouting of dendrites from the PVD. (4) Simplification by branch retraction mediated by EFF-1. The outcome of the regeneration process is the recovery of the PVD morphology and if regeneration by auto-fusion fails the distal part degenerates.

Bauer CC et al. (1998) West Coast Worm Meeting "a, b AND g-FILAGENIN - NEW PROTEINS IN THE CORE'S OF THICK FILAMENTS"

Schmid MF, Ortiz I, Epstein HF, Liu FZ, Bauer CC

[

West Coast Worm Meeting,

1998]

The cores of muscle thick filaments are composed of an outer layer of paramyosin surrounding an inner tubular region containing additional core proteins. In order to study the structure of the thick filament core and the mechanisms and molecules involved in the formation of this length-regulated, multi-protein structural assemblage, we have isolated and characterized three novel proteins: a, b and g-filagenin. We obtained partial peptide sequences from each of the filagenin proteins and determined their predicted protein sequences from the C. elegans Genome Project's database and correlated this information with cDNA sequence obtained from RT-PCR or phage libraries. All three filagenins are novel proteins with no significant homologies to any known proteins (although homologues in parasitic nematodes have been found in the databases). All filagenins also possess the distinction of having calculated isoelectric points of over pH 10 and having a higher than average number of aromatic amino acids. Both b and g-filagenin are predicted to have secondary structures containing only beta strands and loops, whereas a-filagenin is composed of both alpha helices and beta strands. Structural studies of the core indicate that there are seven paramyosin subfilaments crosslinked by additional internal proteins. We tested our structural model with Western blotting using an affinity-purified antibody and have showed that a, b and g-filagenins are associated with the cores. All three filagenins were localized by immunofluorescence microscopy to the A bands of body wall muscles, but there was no detectable reaction with the pharynx, with the exception of a slight reaction with anti-a-filagenin antibody. b-Filagenin has been shown by immunoelectron microscopy to be assembled with paramyosin into the tubular cores of wild-type nematodes at the same periodicity of 72-nm as observed with paramyosin. In CB1214 paramyosin null mutants, b-filagenin assembles with myosin to form abnormal tubular filaments with a periodicity identical to that of wild type. These results verify that the filagenins are core proteins that co-assemble with either myosin or paramyosin in C. elegans to form tubular filaments. Further studies on nematode muscle thick filaments are intended to elucidate the mechanisms behind the formation and regulation of these stable assemblies of myosin and associated proteins. *Supported by grants from the Muscular Dystrophy Association, the National Science Foundation and a NRSA postdoctoral fellowship for Christopher Bauer.

Roy, Peter et al. (2021) International Worm Meeting "A C. elegans Motor-Centric Screening Pipeline Yields Novel and Selective Nematicidal Scaffolds"

[

International Worm Meeting,

2021]

Nematode parasites of humans, livestock and crops pose a significant burden to human health and welfare. Alarmingly, our arsenal of effective nematocidal compounds is being depleted. Parasitic nematodes of animals have rapidly evolved resistance to anthelmintic drugs, and traditional nematicides used for crop protection have been restricted or banned because of poor phylum-selectivity. Here, we present our C. elegans-based discovery pipeline focused on lethal molecules that also induce motor defects. This pipeline yielded multiple new and selective nematicidal small molecule scaffolds (i.e., structurally-related families of molecules). We show that one of these scaffolds, tentatively called the APPs, stimulates neurotransmitter release and immobilizes larvae of multiple nematode parasites of plants and mammals in vitro. At similar concentrations, APP-1 does little to model flies, fish, plants or human cells. Forward genetic screens of 100,000s of mutant genomes failed to yield resistant animals that resist the lethal effects of APP-1, suggesting that resistance to the APPs in the field may not be easily generated. Hence, the APPs represent a novel, selective and potentially useful addition to our nematocidal armament.