Michael E Hurwitz et al. (2005) International Worm Meeting "A Screen for Genes Synthetically Lethal with lin-35 Rb"

Michael E Hurwitz, Bob Horvitz

[International Worm Meeting, 2005]

The class B synMuv gene lin-35 encodes a C. elegans protein similar to Rb, a tumor suppressor inactivated in many human solid tumors. lin-35 mutants may provide a model for mammalian cells harboring mutant Rb genes. Since lin-35 mutations are not lethal, we can screen for genes with functions required for viability in lin-35mutants but not in wild-type animals to identify potential targets for cancer therapy. Pharmacologic inactivation of such target proteins could cause the specific deaths of Rb-deficient cells. We have screened the LGI RNAi library described by Fraser et al. (2000) for genes that are essential specifically in lin-35(n745) animals. n745 causes an early stop codon in lin-35 and is probably a null allele. We have compared the phenotypes following RNAi of lin-35(n745) animals to the published RNAi phenotypes of the wild-type N2 strain. Of the 2445 dsRNAs we have screened, 2128 did not cause abnormalities in either lin-35(n745) or N2 worms. 212 dsRNAs caused similar abnormalities in lin-35(n745) and N2 worms. 105 dsRNAs resulted in abnormalities in lin-35(n745) but not in N2 worms. These 105 dsRNAs were tested in the RNAi-hypersensitive strain rrf-3(pk1426) to assess whether a more complete knockdown of gene function by the dsRNAs might generate more severe defects. 74 caused severe growth defects in rrf-3(pk1426) worms, suggesting that the effects seen in lin-35(n745) worms might be related to increased sensitivity to RNAi of lin-35 worms. To determine whether the effects of the remaining 31 dsRNAs with apparent synthetic lethality on lin-35(n745) animals are strain-specific and possibly allele-specific, we tested these dsRNAs on four other presumptive lin-35 null mutants (n2242, n2239, n2996, n373). In these strains, only three of the remaining dsRNAs caused lethality. These three genes may act in molecular pathways parallel to Rb for an essential cellular function. Currently, we are obtaining deletion alleles of lin-35 and these other genes. It is possible that the abnormal RNAi phenotypes seen in lin-35 mutants but not in N2 animals are caused by non-specific additive effects of two harmful mutations. For example, RNAi of some genes may affect one cell type and the lin-35 mutation another cell type, so that together these two distinct defects result in severely affected animals. Since our goal is to identify pathways that are redundant within single cells, it is crucial to identify RNAs that cause cell-autonomous synthetic lethality. To find such genes, we are developing an assay to determine the effect of inactivation of lin-35 in combination with inactivation of any of the genes identified in our screen in a single tissue.

Hurwitz, Michael E. et al. (2011) International Worm Meeting "sli-1 Cbl Opposes the Engulfment of Apoptotic Cells."

Hurwitz, Michael E., Horvitz, Bob, Anderson, Courtney

[International Worm Meeting, 2011]

In C. elegans, apoptotic cells are engulfed by neighboring cells. Three parallel and partially redundant core pathways comprising at least nine genes (ced-1, ced-6, ced-7, dyn-1 and ced-2, ced-5, ced-10, ced-12 and abi-1) function in this process. Loss-of-function (lf) mutations in these genes cause the persistence of unengulfed cell corpses. CED-7 is an ABC-type transporter that appears to signal through CED-1, a transmembrane receptor and CED-6, an adapter protein predicted to bind the phosphotyrosines of CED-1. CED-6 activates the dynamin, DYN-1, which regulates membrane dynamics. CED-2 is an adapter that contains an SH2 and two SH3 domains and activates the CED-5/CED-12 heterodimeric guanine nucleotide exchange factor (GEF). The CED-5/CED-12 GEF, in turn, activates CED-10, a small Rac GTPase. ABI-1 is a cytoskeletal regulator found in several protein complexes. One protein that interacts with Abi proteins in mammals is the c-Cbl oncoprotein, an adaptor and E3 ubiquitin ligase. In C. elegans, the c-Cbl homolog SLI-1 inhibits LET-23 EGFR/LET-60 Ras signaling by downregulating LET-23 through ubiquitination. We tested whether mutations in sli-1 modify engulfment pathway gene loss of function defects. Loss of sli-1(lf) suppressed the engulfment defects of strong loss-of-function mutations in the ced-1/6/7 pathway but not of the ced-10 Rac pathway, suggesting that sli-1 opposes these pathways but not by acting on the ced-1/6/7 pathway. Instead, sli-1 might act upstream of the ced-10 Rac pathway or in parallel to both pathways. To further investigate the role of sli-1, we determined the effects of sli-1(lf) on another phenotype associated with engulfment mutants. The ced-10 Rac pathway genes are required for normal distal tip cell (DTC) migration and, therefore, normal gonad morphology. By contrast to its effects in engulfment, sli-1(lf) suppressed the DTC migration defects of strong loss-of-function alleles of all the ced-10 Rac pathway genes, suggesting that sli-1 acts in parallel to that pathway. To determine whether SLI-1 acts through the LET-23/LET-60 pathway to inhibit engulfment we assessed the effect of a let-60(gf) mutation on engulfment. let-60(gf) did not cause an engulfment defect and did not enhance or suppress the engulfment defects of genes from either the ced-1/6/7 pathway or the ced-10 Rac pathway. We propose that sli-1 suppresses a genetic pathway parallel to ced-10 Rac for apoptotic cell engulfment and DTC migration independent of its known function to inhibit the LET-23 EGFR/LET-60 Ras pathway. We are currently testing whether sli-1 suppression of engulfment and DTC migration happens in abi-1 mutant animals to assess whether sli-1 acts through abi-1.

Colonnello A et al. (2020) Neurotox Res "Correction to: Comparing the Neuroprotective Effects of Caffeic Acid in Rat Cortical Slices and Caenorhabditis elegans: Involvement of Nrf2 and SKN-1 Signaling Pathways."

Tunez I, Rubio-Lopez LC, Aguilera-Portillo G, Silva-Palacios A, Evaristo-Priego Y, Santamaria A, Galvan-Arzate S, Cortez-Nunez S, Aschner M, Rangel-Lopez E, Chen P, Colonnello A, Robles-Banuelos B, Garcia-Contreras R

[Neurotox Res, 2020]

One of the authors has incorrect family name spelling in the original article_. Michael Ashner should read as Michael Aschner.

Hengartner MO et al. (1992) Worm Breeder's Gazette "unc-50, a Gene Required for Acetylcholine Receptor Function, Encodes an Unfamiliar Protein"

Horvitz HR, Tsung N, Hengartner MO, Lewis JA

[Worm Breeder's Gazette, 1992]

unc-50, a gene requiired for acetylcholine receptor function, encodes an unfamiliar protein Michael Hengartner, Nancy Tsung, Jim Lewist, and Bob Horvitz HHMI, Dept. Biology, MIT, Cambridge, MA 02139, USA. t Division of Life Sciences, U. of Texas at San Antonio, San Antonio, IX 78249

BC Prasad et al. (1999) International C. elegans Meeting "The unc-3 locus encodes an O/E transcription factor, CeO/E, which is required for axonal guidance and chemosensory function."

BC Prasad, RR Reed

[International C. elegans Meeting, 1999]

The O/E family of rHLH transcription factors has been implicated in neurogenesis, axonal pathfinding, muscle formation and B cell maturation. The murine O/E proteins are expressed transiently in the developing CNS and PNS during times of axonogenesis and are expressed in olfactory neurons throughout development where they may regulate components of the olfactory signaling cascade. We have identified a C. elegans O/E homologue (CeO/E) that is encoded by the unc-3 locus. Unc-3 mutants display an uncoordinated phenotype attributed to a severe defasiculation and miswiring of the ventral cord (VNC). Using a GFP fusion to the unc-3 promoter and specific antibodies, we observed that CeO/E is expressed transiently in the excitatory VNC motor neurons and throughout development in a pair of chemosensory neurons (ASI amphid neurons). Several observations suggest that the protein may have distinct roles in the two cell types. Although the axonal and dendritic projections of the ASI appear to be normal when labeled with DiI, unc-3 mutants enter the dauer pathway under inappropriate conditions. Although CeO/E possesses highly conserved domains associated with DNA binding in the mammalian homologue, identification of DNA binding sites for the C. elegans protein has not been achieved. We have shown that CeO/E protein can homodimerize in vitro, although the DNA binding specificity of CeO/E is distinct from the mammalian O/E proteins. Several unc-3 target genes ( daf-7, unc-17/cha-1, unc-4 ) have been identified by other laboratories and each contains a mammalian binding site in the promoter region. Remarkably, we have been unable to demonstrate CeO/E binding at these sites. We are generating chimeras of O/E-1 and CeO/E to understand the DNA binding specificity of the CeO/E protein and utilizing a yeast-2-hybrid screen with both O/E-1 and CeO/E to identify interacting proteins in C. elegans . These studies should reveal the mechanism of CeO/E transcriptional activation and target gene regulation.

Holway AH et al. (2007) J Cell Biol "Checkpoint silencing during the DNA damage response in Caenorhabditis elegans embryos"

Kim SH, Michael WM, La Volpe A, Holway AH

[J Cell Biol, 2007]

After publication of this manuscript, the authors noticed that two images (Fig. 4 B, top left and middle) had been duplicated from a previous publication (Holway, A.H., C. Hung, and W.M. Michael. 2005. Genetics. 169:14511460). Here we provide new images corresponding to these control experiments. The authors apologize for this oversight.

Shaulov Y et al. (2018) PLoS Pathog "Escherichia coli mediated resistance of Entamoeba histolytica to oxidative stress is triggered by oxaloacetate."

Shaulov Y, Lamm AT, Ankri S, Trebicz-Geffen M, Weiss-Cerem L, Hisaeda H, Methling K, Lalk M, Nagaraja S, Shimokawa C

[PLoS Pathog, 2018]

Amebiasis, a global intestinal parasitic disease, is due to Entamoeba histolytica. This parasite, which feeds on bacteria in the large intestine of its human host, can trigger a strong inflammatory response upon invasion of the colonic mucosa. Whereas information about the mechanisms which are used by the parasite to cope with oxidative and nitrosative stresses during infection is available, knowledge about the contribution of bacteria to these mechanisms is lacking. In a recent study, we demonstrated that enteropathogenic Escherichia coli O55 protects E. histolytica against oxidative stress. Resin-assisted capture (RAC) of oxidized (OX) proteins coupled to mass spectrometry (OX-RAC) was used to investigate the oxidation status of cysteine residues in proteins present in E. histolytica trophozoites incubated with live or heat-killed E. coli O55 and then exposed to H2O2-mediated oxidative stress. We found that the redox proteome of E. histolytica exposed to heat-killed E. coli O55 is enriched with proteins involved in redox homeostasis, lipid metabolism, small molecule metabolism, carbohydrate derivative metabolism, and organonitrogen compound biosynthesis. In contrast, we found that proteins associated with redox homeostasis were the only OX-proteins that were enriched in E. histolytica trophozoites which were incubated with live E. coli O55. These data indicate that E. coli has a profound impact on the redox proteome of E. histolytica. Unexpectedly, some E. coli proteins were also co-identified with E. histolytica proteins by OX-RAC. We demonstrated that one of these proteins, E. coli malate dehydrogenase (EcMDH) and its product, oxaloacetate, are key elements of E. coli-mediated resistance of E. histolytica to oxidative stress and that oxaloacetate helps the parasite survive in the large intestine. We also provide evidence that the protective effect of oxaloacetate against oxidative stress extends to Caenorhabditis elegans.

Ye H et al. (2008) Neurobiol Learn Mem "Trace administration of vitamin E can retrieve and prevent UV-irradiation- and metal exposure-induced memory deficits in nematode Caenorhabditis elegans."

Ye B, Ye H, Wang D

[Neurobiol Learn Mem, 2008]

Vitamin E (alpha-tocopherol), a lipid-soluble anti-oxidant, prevents the uncontrolled propagation of lipid peroxidation by free radicals. Nevertheless, there is weak or no evidence of a protective effect of previous vitamin E intake on cognitive function in humans. In the present study, we explored the thermosensation model to investigate the possible effects of vitamin E administration on memory behaviors in Caenorhabditis elegans. Administration of 100 and 200mug/mL of vitamin E had no significant effects on the memory for different time intervals, whereas relatively high concentration (400mug/mL) of vitamin E exposure shortened the extinction period of the association paradigm (food at 20 degrees C). Following the UV-irradiation, post-treatment with 200mug/mL of vitamin E not only retrieved the UV-irradiation-induced memory deficits, but also enhanced the memory functions in UV-irradiating animals. Post-treatment with trace vitamin E could also ameliorate the memory deficits in metal (Al or Pb) exposed worms. In addition, pre-treatment with 200mug/mL of vitamin E could effectively prevent the occurrence of memory deficits induced by metal exposure and UV-irradiation. Therefore, the close association may exist between trace dietary vitamin E intake and memory behaviors, and a specific response mechanism may be activated after the administration of vitamin E in stress-exposed animals. Moreover, treatment with 200mug/mL of vitamin E could restore the memory deficits formed in the ncs-1 mutant worms, suggesting that exogenous treatment with trace vitamin E can largely mimic the function of NCS-1 in regulating the memory for thermosensation.

Saiki R et al. (2008) Aging Cell "Altered bacterial metabolism, not coenzyme Q content, is responsible for the lifespan extension in Caenorhabditis elegans fed an Escherichia coli diet lacking coenzyme Q."

Saiki R, Clarke CF, Furukawa S, Larsen PL, Bixler T, Lunceford AL, Lee W, Dang P

[Aging Cell, 2008]

Coenzyme Qn is a fully substituted benzoquinone containing a polyisoprene tail of distinct numbers (n) of isoprene groups. C. elegans fed E. coli devoid of Q(8) have a significant life span extension when compared to C. elegans fed a standard "Q-replete"E. coli diet. Here we examine possible mechanisms for the life span extension caused by the Q-less E. coli diet. A bioassay for Q uptake shows that a water-soluble formulation of Q(10) is effectively taken up by both clk-1 mutant and wild-type nematodes, but does not reverse life span extension mediated by the Q-less E. coli diet, indicating that life span extension is not due to the absence of dietary Q per se. The enhanced longevity mediated by the Q-less E. coli diet cannot be attributed to dietary restriction, different Qn isoforms, reduced pathogenesis or slowed growth of the Q-less E. coli, and in fact requires E. coli viability. Q-less E. coli have defects in respiratory metabolism. C. elegans fed Q-replete E. coli mutants with similarly impaired respiratory metabolism due to defects in complex V also show a pronounced life span extension, although not as dramatic as those fed the respiratory deficient Q-less E. coli diet. The data suggest that feeding respiratory incompetent E. coli, whether Q-less or Q-replete, produces a robust life extension in wild-type C. elegans. We believe the fermentation-based metabolism of the E. coli diet is an important parameter of C. elegans longevity.

Zhou MH et al. (1996) Worm Breeder's Gazette "Liquid Culture of the Worms with a Fermentor"

Zhou MH, Yang H, Walthall WW

[Worm Breeder's Gazette, 1996]

The yield of C. elegans and their food (E.coli) are relatively low in the large liquid cultures growth (Koelle et al.,1994) and the whole growth process is tedious. Here we report the successful growth of C.elegans with a fermentor. The following protocol is based on the protocol by Michael Koelle and Tory Herman (1994) from Internet.