Musset-Bilal F et al. (1994) Worm Breeder's Gazette "Characterization of the C. elegans Basement Membrane-Associated Protein SPARC"

Musset-Bilal F, Schwarzbauer JE, Ryan CS

[Worm Breeder's Gazette, 1994]

Characterization of the C. elegans Basement Membrane- Associated Frederique Musset-Bilal, Carol S. Ryan, and Jean E. Schwarzbauer Department of Molecular Biology, Princeton University, Princeton NJ 08544

Samuel LIEGEOIS et al. (2003) International Worm Meeting "Genetic evidence suggesting that the V0 domain of V-ATPase is required for exocytosis independently of its role as a proton pump"

Michel Labouesse, Agnes JANOSHAZI, Samuel LIEGEOIS, Jean-Marie GARNIER

[International Worm Meeting, 2003]

Current models of exocytosis are based on genetic data in yeast and biochemical data in neurons. According to the SNARE hypothesis, the final sequence of events in exocytosis would involve the pairing of SNARE proteins on the vesicle and target membranes, followed by a complete fusion of the two membranes. Whether the SNARE hypothesis is valid for all cargo proteins during exocytosis in epithelial cells is unclear, particularly for apical sorting. We previously showed that the gene che-14 is required for secretion in the epidermis. We performed a genetic screen to uncover additional mutations with a che-14-like phenotype and cloned two of them, rdy-1 and rdy-2 (rdy, Rod-like larval lethality and DYe-filling defect). rdy-1 encodes an a subunit of V-ATPase, a proton pump present in all cells. The V-ATPase comprises two domains, a cytoplasmic domain responsible for ATP hydrolysis called V1, and a transmembrane domain responsible for proton transport called V0 (the a subunit belongs to the latter). RDY-1 is localized at the apical membrane of excretory, support and epidermal cells. As shown by mosaic experiments, rdy-1 activity in the excretory cell is essential for survival, while its activity in the epidermis is required for secretion. RDY-1 may be required for secretion of specific but not all proteins. Recent biochemical analysis in yeast indicates that the V0 domain might have a function in membrane fusion. To find out if the rdy-1 mutant phenotypes are associated with a membrane fusion function or with a proton pump activity, we performed site-directed mutagenesis in order to separate both activities. We altered conserved residues on a transgene and introduced them in a rdy-1 null animal to assay survival (as a probable measure of proton pump activity) and cuticle secretion. Of 18 mutations tested so far, E830Q allows almost normal secretion but makes the animal sick and vacuolated, while L786S impairs secretion but does not seem to affect the excretory system. These results support the notion that the V-ATPase might act not only as a proton pump, but also as part of a fusion pore during apical sorting in epithelial cells. rdy-2 encodes a tetra-spanning membrane protein without any homology in databases. RDY-2 is also localized at the apical membrane of some epithelial cells, and preliminary evidence using FRET analysis indicates that it interacts with RDY-1. Moreover, the first protein suspected to be required for apical sorting in vertebrates (VIP/MAL) is also a tetraspanin.

Alexandre Benedetto et al. (2005) International Worm Meeting "The v-ATPase is required for exocytosis at the apical membrane in epithelia, independently of its role as a proton pump"

Jean-Marie GARNIER, Alexandre Benedetto, Michel Labouesse, Samuel LIEGEOIS

[International Worm Meeting, 2005]

The vacuolar H+-ATPase is considered as the main proton pump responsible for osmoregulation in metazoan excretory systems, creating a H+ gradient used as a motor force for most solute exchanges. It is also crucial for pH regulation in all eukaryotic organelles. Mutations in its subunits lead to numerous diseases (for example a subunit disruption in osteoclasts is responsible for osteopetrosis) and prevent tumoral cells to further metastasize. In neurons the v-ATPase-generated H+ gradient is used to fill secretory vesicles with neurotransmitters, and is thus essential for their secretion. The v-ATPase comprises two sectors, V1 is cytoplasmic and responsible for ATP hydrolysis, V0 is transmembraneous and involved in H+ transport. Biochemical experiments and cell culture studies support the notion that V0 proteolipids mediate membrane fusion independently from V1. These observations show that the v-ATPase might play a dual role in secretion: either through its pumping activity, or via V0-facilitated membrane fusion. Our laboratory is analyzing secretion at the apical membrane in C. elegans epithelia. Compared to basolateral trafficking, apical secretion remains poorly understood, and our work provides new insights in this field. We previously showed that rdy-1 (a.k.a. vha-5) encodes the a subunit of the V0 sector that is expressed in hyp7, the excretory, rectal, vulval and support cells associated with sensory neurons. rdy-1 mutants display secretion and pH-regulation defects causing L2 lethality. To determine the role of the a subunit in apical secretion, we attempted to separate its two putative activities: proton pumping performed by the V1+V0 complex and membrane fusion due to V0 alone. We generated 66 site-directed mutations on a functional rdy-1 transgene and introduced mutated constructs in a rdy-1 null animal. We completed this approach by knocking-down V1 and V0 subunits by RNAi. We then assayed excretory canal formation (as a criterion for pumping), and alae formation as well as secretion of apically targeted proteins (as a criterion for apical exocytosis). We obtained mutations that specifically affect the excretory canal (W190A, R191A) and other that mostly affect secretion (L786S, E830Q, V844F). Besides, knocking-down V1 subunits by RNAi also leads to osmoregulation defects, without impairing secretion. Conversely, mutations such as L786S, E830Q and V844F have abnormal or absent alae and accumulate apical or secreted proteins in the hyp7 (RDY-1, CHE-14, WRT-2, WRT-8). We conclude that V1-associated pumping activity is not required for proper secretion to the apical membrane in epithelial cells, and propose that the V0 sector alone is a key player in apical secretion.

Martinelli SD et al. (1995) International C. elegans Meeting "ACEDB"

Durbin RM, Martinelli SD

[International C. elegans Meeting, 1995]

We hope to provide a demonstration of the current state of the ACeDB worm database on Unix workstations, and if possible Apple Macintosh, throughout the poster sessions. This will be based on the new version 4 release of the acedb software (Jean Thierry-Mieg, Richard Durbin and numerous others), which contains many new features for greater efficiency, more flexible printing, and display of new features.

Hinton SD (2019) Biochim Biophys Acta Mol Cell Res "The role of pseudophosphatases as signaling regulators."

Hinton SD

[Biochim Biophys Acta Mol Cell Res, 2019]

Pseudophosphatases are atypical members of the protein tyrosine phosphatase superfamily. Mutations within their catalytic signature motif render them catalytically inactive. Despite this lack of catalytic function, pseudophosphatases have been implicated in various diseases such as Charcot Marie-Tooth disorder, cancer, metabolic disorder, and obesity. Moreover, they have roles in various signaling networks such as spermatogenesis, apoptosis, stress response, tumorigenesis, and neurite differentiation. This review highlights the roles of pseudophosphatases as essential regulators in signaling cascades, providing insight into the function of these catalytically inactive enzymes.

Kelley S (2000) Brief Bioinform "Getting started with Acedb."

Kelley S

[Brief Bioinform, 2000]

Acedb is one of the more venerable pieces of Genomics software. Acedb was originally created in 1992 by Richard Durbin and Jean Thierry-Mieg to manage the data from the Caenorhabditis elegans mapping project and subsequently the C. elegans sequencing project. From beginnings as a C. elegans-specific tool, it has been continuously developed into a flexible suite of data management, display and scripting tools providing facilities for managing and annotation mapping information and DNA and peptide sequences.This paper gives a basic overview of the Acedb suite, and step-by-step guidance on how to download and install Acedb. It is intended to take an Acedb novice to stage where they can begin to experiment and explore the facilities that are available.

(2021) Development "The people behind the papers - Clement Dubois, Shivam Gupta, Andrew Mugler and Marie-Anne Felix."

[Development, 2021]

Cell migration needs to be precisely regulated during development so that cells stop in the right position. A new paper in Development investigates the robustness of neuroblast migration in the <i>C. elegans</i> larva in the face of both genetic and environmental variation. To hear more about the story, we met the paper's four authors: Clement Dubois and Shivam Gupta, and their respective supervisors Andrew Mugler (currently Assistant Professor at the Department of Physics and Astronomy at the University of Pittsburgh, where his lab recently moved from Purdue University) and Marie-Anne Felix (Principal Investigator at Institut de Biologie de l'Ecole Normale Superieure in Paris and Research Director at CNRS).

Dhananjay, Samiksha et al. (2022) MicroPubl Biol

Dhananjay, Samiksha, Neumann, Brent, Chandhok, Gursimran

[MicroPubl Biol, 2022]

Mitochondria are energy-converting organelles that shift between fusion and fission states in order to perform a variety of essential functions. Disruption of these dynamics is detrimental to cellular health and is associated with a range of human diseases. Mitofusin 2 is an essential large GTPase protein that orchestrates fusion of outer mitochondria membranes, and mutations in the encoding gene are causative for Charcot-Marie-Tooth disease. In order to gain further insights into the function of this crucial protein, we have performed large-scale yeast two-hybrid screening to identify interactors of the orthologous protein in Caenorhabditis elegans (FZO-1) . From this screening, we identified 12 novel interactors of FZO-1/mitofusin 2 that, based on their known functions, are strong candidates for further study.

Edgley ML et al. (1995) International C. elegans Meeting "CYBERWORMS"

Collins D, Bryer J, Baillie DL, Edgley ML, Rose AM

[International C. elegans Meeting, 1995]

We are engaged in two projects to improve the alignment of the genetic and physical maps for C. elegans. The Genetic Toolkit project uses PCR to tie the endpoints of balanced deficiencies to the physical map; and the CGAT project uses microinjection of cloned cosmids from the worm sequencing project to generate transgenic arrays that are used to rescue mapped, balanced lethal mutations. Both projects are generating large amounts of genetic map information, which is being integrated into the CGC map (compiled by Jonathan Hodgkin) using the ACEDB program (Richard Durbin and Jean Thierry-Mieg). We also have established a World-Wide Web hypertext server to provide access to information generated by these projects. This Genetic Toolkit web site (http: genetic balancers and transgenic strains available from our laboratories. For stable transgenic strains, listed cosmids are linked to modified GenBank entries describing the sequence. This work is funded by the NIH National Center for Research Resources (NCRR) and the Canadian Genome Analysis and Technology Program (CGAT).

Consortium Wormbase et al. (2000) West Coast Worm Meeting "From ACeDB to a more complete and usable database"

Consortium Wormbase, Foltz NL, Schwarz EM, Sternberg PW

[West Coast Worm Meeting, 2000]

We briefly describe the current status and plans for WormBase, initially an extension of the existing ACeDB database with a new user interface. The WormBase consortium includes the team that developed ACeDB (Richard Durbin and colleagues at the Sanger Centre; Jean Thierry-Mieg and colleagues at Montpellier); Lincoln Stein and colleagues at Cold Spring Harbor, who developed the current web interface for WormBase; and John Spieth and colleagues at the Genome Sequencing Center at Washington University, who along with the Sanger Centre team, continue to annotate the genomic sequence. The Caltech group will curate new data including cell function in development, behavior and physiology, gene expression at a cellular level, and gene interactions. Data will be extracted from the literature, as well as by community submission. We look forward to providing the C. elegans and broader research community easy access to vast quantities of high quality data on C. elegans. Also, we welcome your suggestions and criticism at any time. WormBase can be accessed at www.wormbase.org.