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[
International Worm Meeting,
2013]
Correct alignment and segregation of chromosomes during cell division is essential for proper development and growth. When chromosomes are improperly attached an intracellular signaling cascade, termed the spindle checkpoint, generates a "wait anaphase" signal which delays cell cycle progression until all chromosomes are properly aligned. This signal originates from unattached kinetochores, the macromolecular spindle microtubule-binding machines built on the centromeric regions of chromosomes. Central to the "wait anaphase" signal is the protein MDF-1 (Mad1 in human), but it is still unclear how MDF-1 is recruited to and activated at unattached kinetochores. To address this question, we performed a yeast two-hybrid screen with MDF-1 and a library of kinetochore constituents. We identified an interaction between MDF-1 and BUB-1, a conserved kinase implicated in spindle checkpoint signaling. We then performed a mutagenic yeast two-hybrid screen, which identified alleles of MDF-1 that no longer interact with BUB-1. Prior work had shown that BUB-1 is required for the kinetochore localization of MDF-1. Employing the MosSCI system, we generated MDF-1 mutants that do not interact with BUB-1 and found that the mutant forms of MDF-1 are compromised in their localization to unattached kinetochores. Consistent with this, the MDF-1 mutants that do not interact with BUB-1 were defective in generating a spindle checkpoint signal. As BUB-1 is a kinase, we generated kinase-dead mutants of BUB-1 and found that these mutants no longer interact with MDF-1 in the two-hybrid assay, suggesting that the kinase activity of BUB-1 is required for this interaction. Currently, we are investigating BUB-1 kinase-dead mutants in vivo. Finally, we performed a compensatory mutagenic yeast two-hybrid screen and found a mutation in BUB-1 that restores interaction with a MDF-1 mutant, strongly suggesting that MDF-1 and BUB-1 directly interact. Overall, our findings elucidate the recruitment of MDF-1 to unattached kinetochores via the kinase BUB-1, and show that the specific interaction underlying this recruitment is required for generation of the "wait anaphase" checkpoint signal.
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Kumar, Abhishek, Shah, Pavak, Wu, Yicong, Santella, Anthony, Colon-Ramos, Daniel, Marquina, Javier, Kovacevic, Ismar, Moyle, Mark, Gutierrez, Natasha, Mohler, William, Bao, Zhirong, Shroff, Hari, Christensen, Ryan
[
International Worm Meeting,
2015]
How neuronal networks develop during embryogenesis to generate a functional nervous system is a fundamental question in developmental biology and neuroscience. In C. elegans the entire wiring diagram of the adult nervous system ("the connectome") has been mapped. However, it is unclear how the nervous system is organized during embryogenesis to correctly form the connectome. Through a multidisciplinary collaboration, we have established an imaging and analysis pipeline to generate and share a 4D atlas of C. elegans neurodevelopment throughout embryogenesis. To this end, we created a light-sheet microscope capable of prolonged time-lapse imaging with subcellular resolution. The dual-view selective plane illumination microscope (diSPIM) is capable of constant volumetric 350nm isotropic imaging at speeds 10-1000x faster than conventional 4D imaging techniques. Also, we have collected existing and are generating novel C. elegans strains that label sparse subsets of neurons to facilitate segmentation and are pioneering strategies for heat-shocking/photoactivation to allow for efficient use of pan-neuronal promoters. Furthermore, we developed novel image analysis software (StarryNite) capable of systematically tracking and identifying every developing cell, as well as software for segmenting individual neurons in 4D, and linearizing the twisted C. elegans embryo. Finally, to allow for widespread dissemination of these large data sets, we created iOS and Android applications, and an Internet-based viewing program called CytoSHOW, which allows the scientific community access to all the neurodevelopmental data. We envision a dynamic workspace where any researcher can visualize, integrate, and interrelate the diverse scope of C. elegans neurological and developmental data.
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[
Elife,
2020]
Worms with increased levels of the epigenetic mark H3K9me2 have a longer lifespan that can be passed down to future generations.
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[
Curr Top Dev Biol,
2017]
PAR-1/MARK kinases are conserved serine/threonine kinases that are essential regulators of cell polarity. PAR-1/MARK kinases localize and function in opposition to the anterior PAR proteins to control the asymmetric distribution of factors in a wide variety polarized cells. In this review, we discuss the mechanisms that control the localization and activity of PAR-1/MARK kinases, including their antagonistic interactions with the anterior PAR proteins. We focus on the role PAR-1 plays in the asymmetric division of the Caenorhabditis elegans zygote, in the establishment of the anterior/posterior axis in the Drosophila oocyte and in the control of microtubule dynamics in mammalian neurons. In addition to conserved aspects of PAR-1 biology, we highlight the unique ways in which PAR-1 acts in these distinct cell types to orchestrate their polarization. Finally, we review the connections between disruptions in PAR-1/MARK function and Alzheimer's disease and cancer.
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[
Worm Breeder's Gazette,
1994]
DNA fingerprinting in C. elegans - an approach Mark Beneckea, Jorg T. Epplenb and Einhard Schierenberga a Zoologisches Institut der Univeritat, 50923 Koln, Germany b Ruhr-Universitat, Ab1. fur Molekulare Humangenetik, 44780 Bochum, Germany
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[
Sci Rep,
2016]
Adapters bind motor proteins to cargoes and therefore play essential roles in Kinesin-1 mediated intracellular transport. The regulatory mechanisms governing adapter functions and the spectrum of cargoes recognized by individual adapters remain poorly defined. Here, we show that cargoes transported by the Kinesin-1 adapter FEZ1 are enriched for presynaptic components and identify that specific phosphorylation of FEZ1 at its serine 58 regulatory site is mediated by microtubule affinity-regulating kinases (MARK/PAR-1). Loss of MARK/PAR-1 impairs axonal transport, with adapter and cargo abnormally co-aggregating in neuronal cell bodies and axons. Presynaptic specializations are markedly reduced and distorted in FEZ1 and MARK/PAR-1 mutants. Strikingly, abnormal co-aggregates of unphosphorylated FEZ1, Kinesin-1 and its putative cargoes are present in brains of transgenic mice modelling aspects of Alzheimer's disease, a neurodegenerative disorder exhibiting impaired axonal transport and altered MARK activity. Our findings suggest that perturbed FEZ1-mediated synaptic delivery of proteins arising from abnormal signalling potentially contributes to the process of neurodegeneration.
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[
Mol Cell,
2013]
In this issue of Molecular Cell, Castellano-Pozo etal. (2013) describe a connection between R loop structures and histone 3 S10 phosphorylation (H3S10P), a mark of chromatin compaction. Their results constitute asignificant advance in our understanding of the role of R loops in genomic instability.
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[
Curr Biol,
2010]
The small GTPases Rab5 and Rab7 mark temporally distinct but sequentially connected stages in phagosome maturation, but the mechanism underlying the transition between these stages has been unclear. Recent studies in Caenorhabditis elegans have now uncovered a new protein complex that connects Rab5 to Rab7.
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[
Trends Genet,
2001]
Four recent papers mark a major shift in functional genomic analysis for multicellular organisms. RNA-mediated interference was applied to inactivate individual genes systematically on a genomic scale. These studies subjected a third of the genes in the genome of Caenorhabditis elegans to reverse genetic analysis.
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[
Worm Breeder's Gazette,
1990]
The CGC Bibliography has been translated into a couple of programs other than dBase by various worm people. David Barker and Andy Fire both have sent HyperCard versions to the CGC and Mark Blaxter has sent us a version in FileMaker 2.0. None of these is perfectly up-to-date, so you'll have to be somewhat familiar with the programs to add new references. The data files are available free from the CGC; to get yours, just send a blank 3.5' diskette to Mark Edgley at the CGC with a request letter. In addition, Lew Jacobson has translated the bibliography into a DOS program called Memory Mate and he is willing to distribute the data file to anyone who sends him a blank 5.25' 360 Kb diskette. Memory Mate can be operated as a TSR and called up with a hotkey from the middle of a word processor or other program. Addresses for Mark and Lew can be found in the Subscriber Directory Update in this issue.