Miyeko Mana et al. (2006) Development & Evolution Meeting "A localizome project of C. elegans embryogenesis: a beginning"

Kristin Gunsalus, Miyeko Mana, Fabio Piano, Tony Tao

[Development & Evolution Meeting, 2006]

Phenotypic, proteomic, genetic interaction and expression correlation maps have produced a wealth of molecular information about the early C. elegans embryo. Although powerful, these data offer a static network of global interaction. An important next step is to add a dynamic dimension. The early C. elegans embryo provides an ideal system in which to explore the dynamics of molecular networks by time-lapse imaging of protein localization. In order to accomplish this goal we are working on the following steps: a. develop a cloning and transformation pipeline to express fluorescently tagged proteins in the early embryo; b. set up a data collection system to capture localization dynamics at high spatial and temporal resolution; c. analyze the image data in an unbiased and systematic way to digitally represent the localization patterns and d. archive and distribute all the raw data and strains. We will describe the current state of our project.

Naoki Hisamoto et al. (2002) Japanese Worm Meeting "Survey of components of JNK/p38 MAP kinase cascades in C. elegans"

Naoki Hisamoto, Kunihiro Matsumoto, Kenji Nishide

[Japanese Worm Meeting, 2002]

The c-Jun N-terminal kinase (JNK) and p38 MAP kinase (MAPK) pathways are involved in various stress responses and apoptosis. In mammal, JNK is activated by MKK7 and MKK4 MAPK kinases (MAPKKs) and p38 is activated by MKK3, MKK4 and MKK6 MAPKKs. These members of the MAPKK superfamily are activated by members of MAPKKK superfamily such as MEKK, TAK1, MLK, TAO and ASK1. Although these pathways have been implicated in the response to stresses and inflammation, the physiological roles of the JNK and p38 pathways in the normal development and function of the organism is less well understood. In C. elegans, several mutants corresponding to the components of JNK/p38 cascades have been isolated: jnk-1 (JNK homolog), jkk-1 (MKK7 homolog), sek-1 (MAPKK), mek-1 (MAPKK), mom-4 (TAK1 homolog) and nsy-1 (ASK1 homolog). The C.elegans genome project shows that there are at least seven members of the MAPKKK superfamily, five members of the MAPKK superfamily and seven members of the MAPK superfamily in C.elegans. To investigate the roles of these components and the relationships among them, we isolated several deletion mutants of these genes. Analysis of these mutants will be discussed.

Duerr JS et al. (1995) International C. elegans Meeting "ANATOMY OF CHOLINERGIC NEURONS IN NORMAL AND MUTANT C.ELEGANS AND IN ASCARIS SUUM"

Spaulding D, Johnson CD, Rand JB, Duerr JS

[International C. elegans Meeting, 1995]

Special thanks to Tony Stretton and Judy Donmoyer for assistance with Ascaris We are studying the anatomy of the cholinergic nervous system using two proteins as markers, choline acetyltransferase (ChAT), the acetylcholine synthetic enzyme, and UNC-17, a synaptic vesicle acetylcholine transporter. In C. elegans, antibodies to UNC-17 appear to label synaptic regions, while antibodies to ChAT are enriched in synaptic regions, but also faintly label somas. ChAT and UNC-17 are found in many putative excitatory motor neurons in C. elegans, including the ventral cord motor neurons DA, DB, AS, VA, and VB, as well as VC. Interestingly, the ventral and dorsal sublateral cords show regular punctate staining with anti-UNC-17, suggesting that they have cholinergic synaptic regions. Motor neurons and interneurons in the pharynx are also labeled. Few, if any, purely sensory neurons contain significant levels of these cholinergic proteins. In A.suum, anti-UNC-17 or ChAT also show punctate labeling in a subset of neurons. Labeled neurons include motor neurons that are known from biochemical and physiological studies to be cholinergic. We have begun to examine the level and distribution of these proteins in mutants of C. elegans and find differences in ric-7 (courtesy of Erik Jorgensen), unc-18, unc-33, unc-44, unc-58, unc-75, and unc-104. Supported by grants from OCAST and NIH.

Carrie Cowan et al. (2006) European Worm Meeting "The Establishment of Cortical Polarity in One-Cell Embryos: A Genomics Approach"

Carrie Cowan, Tony Hyman

[European Worm Meeting, 2006]

Carrie Cowan and Tony Hyman. The establishment of the first axis of polarity in the C. elegans embryo is indicated by the formation of two cortical domains corresponding to the anterior and posterior of the embryo. These two domains are marked by different complements of PAR proteins: PAR-3 and PAR-6 localize to the anterior cortex; PAR-1 and PAR-2 localize to the posterior cortex. It remains unknown how the PAR proteins recognize the anterior and posterior cortices. However, a coincident establishment of contractile polarity, mediated by the asymmetric distribution of an acto-myosin contractile network, indicates that cortical activity provides upstream cues for PAR protein segregation. We are interested in understanding how cortical polarity is established in one-cell embryos. We have assessed the effect of approximately 150 genes on embryonic polarity using RNAi-mediated depletion and time-lapse imaging of GFP-PAR-2. The subset of genes analyzed exhibited contractility and/or polarity phenotypes in genome-wide RNAi-based phenotypic screens. We have analyzed our data for 36 individual phenotypes ranging from ruffling and pseudocleavage to centrosome rotation and spindle displacement, allowing us to group the genes into 18 phenotypic categories. These phenotypic categories reveal several important aspects of polarity establishment, including thresholds of contractility during polarization. For instance, reducing cortical contractility prevents ruffling but not PAR-2 polarity, while eliminating cortical contractility prevents both ruffling and PAR-2 polarity. We present a generalized model (and the genes required) for polarity establishment in the one-cell embryo: 1) centrosome activation, 2) acto-myosin contraction, 3) centrosome migration to the cortex, 4) initiation of posterior domain formation, 5) posteriorization, 6) spreading of the posterior cortical domain, 7) pseudocleavage, and 8) regulation of posterior domain size.

Spiga, Fabio et al. (2010) C. elegans: Development and Gene Expression, EMBL, Heidelberg, Germany "The TAO kinase KIN-18 regulates contractility and establishment of polarity in the embryo."

Spiga, Fabio, Gotta, Monica, Prouteau, Manoel

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

Establishment and maintenance of cell polarity are fundamental processes for many aspects of cell and developmental biology, including asymmetric cell division. In the C. elegans embryo the first axis of polarity, the anterior-posterior axis, is determined by the sperm entry site. Shortly after meiosis, the cortex is uniformly contractile and the conserved PAR-3/PAR-6/PKC-3 complex (PAR-3 complex) is localised all around the cortex. Entry of the sperm triggers posterior smoothening and anterior-directed cortical acto-myosin flows, which are regulated by the small G protein RHO-1. Asymmetric contractions lead to the formation of an anterior domain containing the PAR-3 complex, while PAR-2 occupies the posterior cortex. Although contractility is necessary for the localization of PAR proteins, anterior P AR proteins exert a feedback regulation on contractility. Despite the central role of cortical contractility and of the PAR-3 complex in cell polarity, little is known on how they regulate each other. Here we describe the role of KIN-18, a member of the TAO/Ste20-like kinase family, in the regulation of cortical contractions and cell polarisation. kin-18(RNAi) embryos exhibit increased and prolonged cortical contractions, producing a more persistent pseudocleavage. This hyper-contractility results in a defect in the position of the boundary between the anterior and posterior PAR domains during polarity establishment. We find that KIN-18 binds to the small G protein RHO-1 and acts by modulating its activity in a PAR-3 regulated fashion. Our data reveal that KIN-18 provides a novel link between the cytoskeleton remodelling and the PAR polarity machineries and show that KIN-18 is essential to precisely regulate the establishment of cell polarity.