Tax FE et al. (1994) Worm Breeder's Gazette "Some Notes on Mapping Contigs Using Molecular Analysis of Deficiences"

Thomas JH, Tax FE

[Worm Breeder's Gazette, 1994]

Some notes on mapping contigs using molecular analysis of deficiencies Frans Tax and Jim Thomas, Dept. of Genetics, University of Washington, Seattle WA 98195

Broverman SA et al. (1992) Worm Breeder's Gazette "Transformation Rescue of him-8"

Meneely PM, Broverman SA

[Worm Breeder's Gazette, 1992]

1) him-8, him-5 and him-1 asymmetrically affect recombination (pairing?) of the X chromosome. - 2) Transformation rescue of him-8. Sherryl Broverman and Philip Meneely, FHCRC, 1124 Columbia St., Seattle WA 98104, (206) 667-4523i FAX 206 667 4737

Baldwin AT et al. (2016) J Cell Sci "Unique and redundant -catenin regulatory roles of two Dishevelled paralogs during C. elegans asymmetric cell division."

Phillips BT, Clemons AM, Baldwin AT

[J Cell Sci, 2016]

The Wnt/-catenin signaling pathway is utilized across metazoans. However, the mechanism of signal transduction, especially dissociation of the -catenin destruction complex by Dishevelled proteins, remains controversial. Here we describe the function of the Dishevelled paralogs DSH-2 and MIG-5 in the Wnt/-catenin Asymmetry (WA) pathway in Caenorhabditis elegans, where WA drives asymmetric cell divisions throughout development. We find that DSH-2 and MIG-5 redundantly regulate cell fate in hypodermal seam cells. Similarly, both DSH-2 and MIG-5 are required for negative regulation of SYS-1/-catenin localization, but MIG-5 has a stronger effect on polarity of SYS-1 localization. We show that MIG-5 controls cortical APR-1/APC localization. DSH-2 and MIG-5 both regulate the localization of WRM-1/-catenin, acting together as negative regulators of WRM-1 nuclear localization. Finally, we demonstrate that overexpression of DSH-2 or MIG-5 in seam cells leads to stabilization of SYS-1 in the anterior seam daughter, solidifying the Dishevelleds as positive regulators of SYS-1/-catenin. Overall, we have further defined the role of Dishevelled in the WA signaling pathway, and demonstrated that DSH-2 and MIG-5 regulate cell fate, -catenin nuclear levels and polarity of -catenin regulation.

Akhoon BA et al. (2018) Exp Gerontol "Withanolide A extends the lifespan in human EGFR-driven cancerous Caenorhabditis elegans."

Rathor L, Pandey R, Akhoon BA

[Exp Gerontol, 2018]

The conserved EGFR pathway is linked with multiple cancers in humans including breast, ovarian, and lung carcinoma. Withanolide A, one of the major withanolidal active compounds isolated from the Withania somnifera, extends lifespan and ameliorates stress resistance in wild-type C. elegans by targeting the Insulin/IGF-1 signaling pathway. Up-regulation of IGF1 can transactivate EGFR which inturn reduces longevity and promotes tumor development in an organism. We examined the effects of Withanolide A on the lifespan of a human EGFR-driven C. elegans transgenic model exhibiting the multivulva (Muv) phenotype. The results showed that WA extends the lifespan of both wild human EGFR-driven C. elegans model (human wild-type tyrosine kinase) as well as models bearing single (L858R), and double mutations (T790M-L858R). The lifespan extension observed in these transgenic strains was 20.35, 24.21 and 21.27%, respectively. Moreover, the reduced fat levels were noticed in both wild-type N2 worms and transgenic strains. These observations support the heathspan promoting effect of WA as lipid-rich diet has been reported to promote tumor development. In view of the fact that most of the well known FDA approved drugs such as gefitinib fail to inhibit the EGFR-associated cancers because of these mutations, the present findings show the potential of Withanolide A as a foreseen future nutraceutical to improve the average survival of cancer patients.

Higazi TB et al. (2011) Am J Trop Med Hyg "Polymerase chain reaction pool screening used to compare prevalence of infective black flies in two onchocerciasis foci in northern Sudan."

Hassan HK, Richards F, Mackenzie CD, Mohamed HA, Higazi TB, Unnasch TR, Deran TC, Aziz N, ElMubark WA, Mohamed WA, Zarroug IM, Katabarwa M

[Am J Trop Med Hyg, 2011]

Onchocerciasis remains an important debilitating disease in many areas of Africa, including Sudan. The status of infection transmission in 2007 was assessed in the vectors of two disease foci in Sudan: Abu Hamed in northern Sudan, which has received at least 10 years of annual treatment and Galabat focus in eastern Sudan, where only minor, largely undocumented treatment activity has occurred. Assessment of more than 30,000 black flies for Onchocerca volvulus infectious stage L3 larvae by using an O-150 polymerase chain reaction protocol showed that black fly infectivity rates were 0.84 (95% confidence interval = 0.0497-1.88) per 10,000 flies for Abu Hamed and 6.9 (95% confidence interval = 1.1-16.4) infective flies per 10,000 for Galabat. These results provide entomologic evidence for suppressed Onchocerca volvulus transmission in the Abu Hamed focus and a moderate transmission rate of the parasite in the Galabat focus.

Darby C et al. (1993) Worm Breeder's Gazette "A Call for Bacteria and for Soil Samples"

Darby C, Thomas JH, Manoil C

[Worm Breeder's Gazette, 1993]

We are interested in using genetics to analyze simultaneously both organisms in a two-species interaction. Therefore, we are searching for either a symbiotic or pathogenic relationship between C. elegans and a bacterium. A commensalism, in which bacteria persist within the worm but produce no obvious effects, would also qualify, but would be technically difficult to analyze. We ask your help in providing two things: Bacteria. Contaminants growing on worm plates are the most obvious source. If you have a contaminant that appears to kill worms or make them sick, or which worms obviously avoid, please send an agar stab or even the original plate. It is not necessary for you to identify the bacterium or perform any experiments, but please provide a brief description of how the organism was found and what its effects appeared to be. Soil: We are screening soil samples for bacteria that produce the kinds of effects described above. Any soil in which bacteria-eating nematodes occur is a candidate, but the best results are likely to be from rich, moist, temperate soils. It is not necessary for you to confirm the presence of nematodes. A 50 ml capped plastic tube should be ample. Please supply the location and date and a general description of the habitat (e.g. "Olympic National Forest, Hoh River valley, 9-1-93, river bank in temperate rain forest"). Please send samples to: Creg Darby Dept. of Genetics, SK-50 University of Washington Seattle, WA 98195 Comments and suggestions are also welcome: email: cdarby@u.washington.edu phone: (206) 543-9446 fax: (206) 543-0754

Ahmed M. Mohamed et al. (2007) International Worm Meeting "The Adaptor Molecule NCK-1 is a Component of the VAB-1 Eph RTK Signaling Pathway."

Jeffrey Boudreau, Ian D. Chin-Sang, Ahmed M. Mohamed

[International Worm Meeting, 2007]

VAB-1 is a C. elegans Eph receptor tyrosine kinase (RTK), and is involved in various aspects of neuronal and epidermal development. Null mutations in vab-1 result in abnormal neuroblast movements, defective ventral epidermal enclosure and various axon guidance defects [1-5]. We created animals carrying a hyper-active VAB-1 tyrosine kinase in the touch neurons by targeting the intracellular portion of VAB-1 to the plasma membrane via N-terminal myristoylation (MYR-VAB-1). The MYR-VAB-1 protein resulted in a penetrant, premature termination defect in the PLM axons that are independent of the EFN-1 ligand [3]. Using a candidate gene approach we identified nck-1 as a suppressor of MYR-VAB-1. nck-1 encodes a protein with three SH3 domains and a single SH2 domain that is highly similar to the Drosophila DOCK, and mammalian NCK2 protein. Characterization of the NCK-1 expression pattern indicates that, like VAB-1, NCK-1 is expressed in the neuroblasts of embryos, the nerve ring, ventral nerve cord and head neurons. Unlike VAB-1, NCK-1 is also expressed in epidermal cells. Analysis of nck-1 mutants revealed the presence of PLM axon defects similar to vab-1 mutants, where PLM axons overextended beyond their target region. Furthermore, NCK-1 seems to be required for proper amphid neuronal cell positioning and axon guidance. Western blots using anti-NCK-1 antibodies revealed two isoforms of NCK-1. The larger NCK-1 isoform was identified as a binding partner of VAB-1 in a yeast two hybrid screen. In a vab-1 null background the larger NCK-1 isoform levels are reduced suggesting that the larger NCK-1 isoform is less stable in the absence of VAB-1. Protein deletion analysis demonstrates that NCK-1 physically associates with VAB-1 through its SH2 domain in a VAB-1 tyrosine kinase dependent manner. Our results suggest that NCK-1 is involved in neuronal development, and is a component of the VAB-1 signaling pathway. We are currently investigating the role of NCK-1 in axon guidance, and its interaction with VAB-1. [1] Boulin et al., Current Biology 2006 16:1871-83. [2] George et al.,, Cell 1998 92:633-43. [3] Mohamed and Chin-Sang, Developmental Biology 2006 290:164-76. [4] Mork et al., Developmental Biology 2003 260:158-75. [5] Zallen et al. Development 1999 126:3679-92.

Clari Valansi et al. (2003) International Worm Meeting "EFF-1A and EFF-1B transmembrane proteins are required for cell fusion and are differentially expressed in pharynx, seam cells and hypodermis."

Benjamin Podbilewicz, Clari Valansi

[International Worm Meeting, 2003]

Cell fusion is an abundant process during development in C. elegans. Previous work led to the isolation of eff-1, a gene essential for cell fusion. EFF-1 predicted structure shows five putative domains: a signal peptide, a phospholipase A2 site, a fusion peptide, a trans membrane domain and a cytoplasmic tail (1). eff-1 encodes at least four isoforms, two predicted type-I membrane proteins (EFF-1A and EFF-1B) and two secreted proteins (EFF-1C and EFF-1D). The transcriptional eff-1::GFP fusion construct was shown to be expressed in hypodermal cells committed to fusion and absent from hypodermal cells that escape fusion (2). In order to study the functions of the different isoforms, we decided to analyze their expression pattern by producing polyclonal antibodies against two different peptides. The first antibody raised against the N-terminus of EFF-1, recognizes the extracellular domain, which is shared by EFF-1A (74.4 kDa), EFF-1B (67.3 kDa), EFF-1C (15 kDa) and EFF-1D (27 kDa) isoforms. The second antibody raised against the C-terminus, recognizes a specific domain within the cytoplasmic tail, existing only in the EFF-1A isoform. Our western blot and immunoprecipitation results imply that the antibody against the N-terminus (extracellular domain) recognizes three proteins of 73 / 68, 41 and 29 kDa, while the antibody against the C-terminus only identifies the high molecular weight proteins. Immunofluorescence of worms using the two antibodies showed that: 1) the antibody recognizing all isoforms strongly stains the pharynx throughout development and weakly stains embryonic hypodermis; whereas 2) the antibody specific for EFF-1A recognizes the seam cells and the hypodermis at the L2 and the L3 stages. These results are consistent with results of an EFF-1A::GFP expression construct that was obtained in our laboratory (3). We are currently expressing EFF-1B in bacteria and mammalian cells; we will show western blots testing the antibodies in bacteria, mammalian cells, N2 and different eff-1 alleles. 1.Mohler WA. et al. Dev Cell 2002. 2.Shemer G. and Podbilewicz B. Genes Dev 2002. 3.Assaf-Reizel N. and Podbilewicz B. (2003) 14th International C. elegans Meeting

Zanetti, Michael R. et al. (2011) International Worm Meeting "Suppressors of daf-18 L1 arrest phenotype and investigating a role for DAF-18 protein phosphatase activity."

Zanetti, Michael R., Chin-Sang, Ian D., Hand, Peter E.

[International Worm Meeting, 2011]

The C. elegans Eph receptor tyrosine kinase (RTK), VAB-1, has been identified as an important player in signaling pathways controlling lifespan, axon guidance and morphogenetic movements. As such, it is important to understand how the individual components of this signaling cascade are functioning. Mutations in one of the downstream targets, daf-18(ok480), results in defective dauer formation and defective L1 arrest. DAF-18 is the homolog of the human tumor suppressor PTEN and is a negative regulator of an insulin/IGF-like pathway for longevity and dauer larva formation. In order to identify genes that function with daf-18 in L1 arrest we used two approaches to look for suppressors of the daf-18 L1 arrest phenotype. First, we used a candidate gene approach using RNAi knockdowns, which identified eleven suppressors. For suppressor genes that have mutants available, we will create double mutants with daf-18(ok480) to confirm suppression of the daf-18 L1 arrest phenotype. Secondly, we used an unbiased EMS mutagenesis screen, which identified twelve potential suppressors. These suppressors are being mapped to chromosome locations and non-complementation tests will be carried out to determine if they are novel suppressors. Previous work in our lab showed that VAB-1 and DAF-18/PTEN are substrates for each other and mutually inhibit each other [1]. We also have shown vab-1 mutants can partially suppress daf-18(ok480) L1 arrest. Human EphA3, a VAB-1 homolog, is frequently mutated in lung cancers, in particular a missense mutation in the tyrosine kinase domain EphA3(K761N), and predicted to have oncogenic properties [2]. To test gain-of-function properties in vivo we created C. elegans transgenic lines with human EphA3(K761N) and the analogous mutation in VAB-1(K859N). VAB-1(K859N) transgenic worms have a phenotype similar to hyperactive VAB-1 gain-of-function mutations where PLM axons show premature termination [3]. Based on the antagonistic interaction between VAB-1 and DAF-18 we predict that DAF-18 will dephosphorylate VAB-1(K859N) and rescue the premature termination defects. We will also test whether this regulation is conserved in human EphA3 and PTEN. We will report our findings at the meeting. [1] Brisbin et al. 2009, Dev. Cell 17, 459-469 [2] Ding et al. 2008, Nature 455, 1069-1075 [3] Mohamed and Chin-Sang. 2006, Dev. Biol 290, 164-176.

Jeffrey R Boudreau et al. (2003) International Worm Meeting "The VAB-1 Eph Receptor Tyrosine Kinase Interacts with the C. elegans Tumor Suppressor DAF-18/PTEN and May Regulate PI(3,4,5)P3 Signaling During Neuronal Development."

Jeffrey R Boudreau, Ian D Chin-Sang

[International Worm Meeting, 2003]

The C. elegans VAB-1 Eph RTK and its ephrin ligand, EFN-1 function in neurons for proper neuronal and epidermal morphogenesis. To identify downstream components of the Ephrin signaling we used the tyrosine kinase domain of VAB-1 as bait in a yeast two-hybrid screen. We isolated 4 independent cDNA clones of the daf-18 gene. The daf-18 gene encodes a homolog of the human tumor suppressor PTEN. PTEN is a dual-specificity protein phosphatase that can dephosphorylate protein substrates such as FAK and the adapter protein Shc but its physiological substrate appears to be Phosphatidylinositol 3,4,5-trisphosphate (PIP3), a lipid signaling molecule. PTEN antagonizes phosphatidylinositol-3 kinase (PI3K) the enzyme that is required to produce PIP3. The role of PTEN is evolutionarily conserved as Caenorhabditis elegans DAF-18/PTEN antagonizes the PI3 Kinase, AGE-1, in regulating metabolism, and life span. However, the DAF-18/PTEN in C. elegans probably does not act exclusively in the aging and dauer pathways, as this gene is pleiotropic and other phenotypes including defects in head and vulva morphogenesis and embryo lethality are observed. Phosphatidylinositol lipids, such as PI(4,5)P2 and PI(3,4,5)P3 mediate diverse intracellular signaling pathways to regulate the actin cytoskeleton during cell growth and movement. The VAB-1 kinase binds to the C- terminal portion and not the phosphatase region of DAF-18. Although very little is known about how PTEN is regulated, it has been proposed that the C-terminus regulates the PTEN activity possibly by phosphorylation. The VAB-1 interaction appears to be kinase dependent as a missense mutation (G912E), which disrupts kinase activity, abolished the interaction in the yeast two-hybrid assay. To understand how VAB-1 might act with DAF-18 we used a constitutively active VAB-1 kinase in the touch neurons that causes severe axon defects (See Mohamed and Chin-Sang, this meeting). We find that both age-1(mg44) and daf-2 (e1375) can partially suppress these axon defects. Our hypothesis is that VAB-1 has a negative influence on DAF-18, which leads to increased PIP3 levels, and this is compensated for in the age-1 and daf-2 mutants, which reduce PIP3 levels. Our finding that PTEN/DAF-18 physically interacts with the VAB-1 kinase may provide a model in which these two enzymes are necessary to set the PIP3 or PIP2 levels required for proper morphogenesis in C. elegans.