Wong, Caleb K.H. et al. (2009) International Worm Meeting "Interaction of BMP facilitator, crm-1, with a downstream target lon-1."

Chow, King L., Wong, Caleb K.H., Mok, May G.Y.

[International Worm Meeting, 2009]

CRM-1 encoded by the worm homolog of vertebrate Crim1, has multiple cysteine-rich (CR) domains with potential bone morphogenetic proteins (BMP) binding activity. crm-1 mutant displays small body phenotype with decreased ploidy in the hypodermis, a feature opposite to lon mutants. We have previously suggested that crm-1 controls body size via modulation of dbl-1/sma activity in the hypodermis and thus endoreduplication, and it functions non-autonomously in facilitating dbl-1 signaling event. Interestingly, data from the C. elegans Interactome Network showed that CRM-1 interacts with LON-1 in yeast. Would this LON-1 transmembrane protein regulated by BMP signal be able to act extracellularly to antagonize BMP signaling as a feedback circuit to sustain its own effect of blocking endoreduplication? As an alternative, CRM-1 may indirectly facilitate BMP signaling via association of LON-1 and blocks the lon-1 repression on endoreduplication. To test these hypotheses, characterization of crm-1 and its interaction with lon-1 was initiated. Isoforms of crm-1 were tested for their ability to interact with lon-1 product. Body length rescue experiments by crm-1 cDNA isoforms were also conducted to identify functional domains of the CRM-1 protein. At this moment, only the intracellular function of lon-1 has been documented for repressing endoreduplication in the hypodermis. We will examine the extracellular function of LON-1 by ectopically expressing lon-1 cDNA in tissues outside the hypodermis. Mutant rescue activities are monitored in these ectopic expression studies to evaluate lon-1''s potential non-autonomous and autonomous function. Furthermore, the double mutant of crm-1 and lon-1 has been constructed, the body length and hypodermal ploidy will be examined to elucidate their genetic relationship. The data from these experiments and the implication will be discussed in this poster. (The study is supported by Research Grants Council, Hong Kong.).

Agnes K.Y. Hui et al. (2008) "Stress and mis-assembled collagens cause C. elegans male tail anomaly."

King L. Chow, Caleb K.H. WONG, Agnes K.Y. HUI

[2008]

"Stress-induced physiological responses are used as strategies to adapt variable environmental conditions. However, the stress responses can alter normal developmental process if the stress is applied during specific developmental time window. C. elegans male tail was used as a model to test the impact of various stresses on development. Various stress inducers such as heat and tunicamycin were applied during late L4 stage. We found that both heat shock and tunicamycin cause male tail anomaly similar to what we observed in ram mutants. As both heat shock and tunicamycin can trigger ER stress and thus unfolded protein response (UPR), we would like to test whether expression of unfolded protein in male tail can result in male tail morphological defect. Three male tail specific collagens ram-1, ram-2 and ram-4 were used for this analysis because 1) they are expressed in the male tail hypodermis at L4 stage; 2) Dominant negative and loss of function genetic mutants of these genes are available. 3) Collagens were assembled at the ER before secreting to the extracellular matrix. We reason that missing one or two chains of the pro-collagen could leave the unassembled collagen chains being recognized as unfolded protein. We monitor whether wild type ram-4 collagen was trapped in ER of ram-2 mutant by immunostaining HA-tagged ram-4 collagen in transgenic worms expressing hypodermal ER marker. Stronger immunostaining signal in ER was observed in ram-2 mutant in contrast to wild type animal suggesting collagen assembly process is defective in ram mutant. Further analysis of ram collagen in different ram mutants will be performed and the results will be discussed. (The research is funded by Research Grants Council, Hong Kong.)"

Wendy Wong et al. (2006) European Worm Meeting "The Integrative Interaction Map for Caenorhabditis elegans"

Wendy Wong, Andrew Fraser

[European Worm Meeting, 2006]

. Wendy Wong and Andrew Fraser. C. elegans is a popular model system for the systematic analysis. However, relatively little is known for its networks of genetic interactions. We hereby construct an integrative interaction map for C. elegans by integrating diverse functional genomics Datasets. We will also present some of the novel tools in querying the interaction map for answering powerful biological hypotheses.

Ewbank JJ et al. (1997) International C. elegans Meeting "Expression and activity of clk-1"

Ewbank JJ, Lemieux JY, Felkai S, Labbe J-C, Hekimi S

[International C. elegans Meeting, 1997]

Mutations in clk-1 affect biological timing and extend longevity (Wong et al., Genetics, 139: 1247, (1995); Lakowski and Hekimi, Science, 272: 1010, (1996)). clk-1 encodes a protein of 187 amino acids that is highly conserved from yeast to mammals. The yeast homologue is the metabolic regulator Cat5p/Coq7p. clk-1 complements the phenotype of cat5/coq7 mull mutants, demonstrating that clk-1 and its yeast homolgue have a common biochemical function (Ewbank et al., Science, 275: 982, (1997)). We have now found that clk-1 is expressed at high levels in all somatic cells throughout development. Furthermore, under heat shock conditions the transcription of clk-1 and/or the stability of its message requires the activity of CLK-1. We are currently using Northern analysis, reporter constructs and antisera to study clk-1 expression further. We are particularly interested in the effects of temperature on clk-1 expression and determining the subcellular localization of CLK-1. We are also analyzing the phenotypic effects of everexpressing wild type or mutant versions of CLK-1 and investigating the molecular correlates of the maternal effect displayed by clk-1 (Wong et al., (1995)).

Lakowski B et al. (1996) East Coast Worm Meeting "The Clock genes interact to determine lifespan by a novel mechanism"

Lakowski B, Hekimi S

[East Coast Worm Meeting, 1996]

Mutations in the genes clk-1, clk-2, clk-3 and gro-1 are all maternally rescued and lead to a very similar phenotype: a pleiotropic alteration of developmental and behavioral timing (Hekimi et al. 1995, Wong et al. 1995). Mutations in the best characterized of these genes, clk-1, lengthen early embryonic cell cycles, embryonic and post-embryonic development, the periods rhythmic behaviors, such as defecation, and ultimately total lifespan (Wong et al. 1995). We have determined that mutations in all four of these genes increase both mean and maximum lifespan. To examine how these genes interact genetically, we made seven double mutant strains containing combinations of Clock mutations. Although there is a range of severity, all double mutants take longer to develop than mutants in the individual constituent genes. Most double mutants also have much longer mean and maximum lifespans than the individual Clock mutants. The strongest double mutant combination, clk-3(qm38); clk-1(qm30), takes four times as long to develop and lives three times as long as N2. Life span extension in clk mutants and gro-1 is not dependent on the activity of daf-16, which is essential for the life extension seen in age-1, daf-2 and daf-23 mutants, suggesting that the Clock genes and the dauer genes act in different pathways. To examine the effect of carrying a life extending mutation from two different pathways, we made a daf-2(e1370) clk-1(e2519) double mutant. daf-2 clk-1 double mutants live longer than either daf-2 or clk-1 and can live up to five times as long as N2. This is the greatest increase in lifespan recorded for any organism by any means. By other criterion, lifespan extension in the Clock mutants also appears to be mechanistically distinct from that of the dauer mutants. For example, when dauer mutants are raised at the permissive temperature, adult lifespan is lengthened without any increase in the length of development. However, in Clock mutants, increased adult lifespan is accompanied by changes in developmental and behavioral timing. In particular, mean adult lifespan and the length of post-embryonic development are strongly correlated among all clk single and double mutants. These observations lead us to suggest that Clock mutants have a slower "rate of living" than the wild type, which may lead to longer lifespan, possibly by slowing the production of reactive byproducts of metabolism. Hekimi et al. Genetics 141, 1351 (1995). Wong et al. Genetics 139, 1247 (1995).

Lemieux JY et al. (1997) International C. elegans Meeting "TOWARDS THE MOLECULAR CHARACTERIZATION OF THE BIOLOGICAL TIMING GENE gro-1"

Lakowski B, Barnes TM, Lemieux JY, Hekimi S

[International C. elegans Meeting, 1997]

In a screen for C. elegans viable maternal-effect mutants, a number of novel phenotypic classes were defined (1). One of these classes is defined by the three clk genes, mutations in which result in a mean lengthening of development and life span, and an increase in the periods of rhythmic behaviors such as defecation, swimming and pumping (1). We have found that a fourth gene, gro-1, also shows the Clk phenotype (2,3). gro-1 was first recovered by Jonathan Hodgkin as a spontaneous segregant from a wild strain (personal communication). Of the four genes, only clk-1 has been thoroughly studied; a comprehensive phenotypic analysis was carried out by Wong et. al. (2) and the molecular cloning by Ewbank et. al. (4). The molecular characterization of each of the Clk genes will aid in understanding the Clk phenotype and ultimately the processes in which these genes are involved. We have mapped gro-1 to the interval between dpy-17 and clk-1 on LGIII. Injection of individual cosmids spanning this region identified three overlapping cosmids which were capable of rescue. Using the available genomic sequence, we tested individual predicted genes from the common rescuing region. This led to the identification of a single gene necessary and sufficient for rescue. The subsequent sequencing of the gene from the gro-1 mutant strain identified the e2400 lesion, confirming the gene assignment. Studies are now underway to determine the expression pattern of GRO-1. Details will be presented at the meeting. 1. Hekimi, S. et. al. Genetics 141: 1351-1364. 2. Wong, A. et. al. Genetics 139: 1247-1259. 3. Lakowski, B. and S. Hekimi. Science. 272: 1010-1013. 4. Ewbank, J. J. et. al. Science 275: 980-983.

Murakami S et al. (1996) West Coast Worm Meeting "LIFE-EXTENSION PATHWAY IN C.ELEGANS"

Johnson TE, Dames SA, Murakami S, Tedesco PM, Duhon SA

[West Coast Worm Meeting, 1996]

Life extension is conferred either by mutants extending adult life (Age), including age-1, daf-2, daf-23, daf-28, spe-26, and an allele of clk-1. Another mode of life extension is conferred by mutants delaying developmental stage (Clk), including clk-1, clk-2, clk-3, gro-1(Wong et al., 1995; Lakowski and Hekimi, 1996). Three groups have identified a life-extension pathway formed by some Daf mutations (Dorman et al., 1995; Larsen et al., 1995; Murakami and Johnson, 1996). We found that the pathway is not limited to Daf mutations but includes other types of the Age mutants (Murakami and Johnson, 1996). Interestingly, the pathway confers UV resistance, suggesting that UV resistance is a good indicator of Age. It is also consistent with the hypothesis that stress resistance is a rate limiting factor determining longevity. We are also testing whether the Age-pathway can confer other types of stress such as H2O2 and heat.

H Pamjav et al. (1999) International C. elegans Meeting "Phylogenetic analysis of entomopathogenic nematode / bacterium symbiotic complexes by using PhastSystem PAGE PCR-RFLP of rDNA spacer sequences"

D Triga, A Fodor, H Pamjav, Z Buzas, E Szallas

[International C. elegans Meeting, 1999]

Entomopathogenic nematode (EPN)species belonging to the Steinernema and Heterorhabditis genera are closely related to C. elegans (Blaxter et al., 1998). They form an intimate symbiotic complex with bacteria Xenorhabdus spp. and Photorhabdus spp., respectively, which are closely related to E. coli . The symbiosis is developmentally regulated: there is a direct cell-cell contact between the gut cell of the (infective) dauerlarva and the primary phase variant of the symbiotic bacteria. The symbiosis is taxon specific. Individual Xenorhabdus species used as symbionts by each Steinernema species and individual strains of Photorhabdus luminescens used as symbionts by each Heterorhabditis strains belonging to different species. The phylogenetic relations between species and strains have been analysed by RFLP of the internally transcribed spacer region of the respective rRNA operon. The PCR-amplified DNAs were digested by different restriction enzymes and the restriction patterns obtained by PhastSystem PAGE were compared. On the basis of the comparative analysis of the ITS1 - ITS2 patterns nematodes could be identified at species level. Xenorhabdus species could also be identified on the basis of the comparative analysis of the spacer region of the 16S - 23S rRNA operon. P. luminescens strains, belonging to different subclusters (Szallas et al., 1997) could be separated on the basis of the RFLP pattern of the spacer region of the 16S - 23S rRNA operon. Adams, B.J. (1998). Species conception and the evolutionary paradigm in modern nematology. J. nematol. 30, 1-21. Adams, B.J., Burnell, A.M. @ Powers,T.O. (1998). A phylogenetic analysis of Heterorhabditis based on ITS 1 DNA sequence data. J. nematol. 30, 22-39. Blaxter,R. et al.(1998).Molecular evolutionary framework for Phylum nematoda. Nature 392,71-75. Szallas E., Koch, H., Fodor,A.,Szentirmai, A., Nealson,K.H. Stackebrandt, E.(1997).Phylogenetic evidence for taxonomic heterogeneity of P.luminescens. IJSB 47, 402-407.

Wong, Garry et al. (2019) International Worm Meeting "miRToolsGallery: a microRNA bioinformatics resources database portal."

Wang, Changliang, Knott, K. Emily, Heikkinen, Liisa, Yang, Yang, Chen, Liang, Wong, Garry

[International Worm Meeting, 2019]

microRNAs (miRNAs) are a class of small noncoding RNA derived from primary miRNA transcripts and processed into precursors and eventually to mature miRNAs 21-22 nucleotides in length. miRNAs were originally discovered in C. elegans and are now known to be expressed in animals, plants, and viruses. miRNAs exert their actions as important regulators of cell proliferation, growth, and development via RNA silencing mechanisms. Bioinformatics resources for miRNA study have been developed, but identifying a correct tool for a specific application is tedious and laborious. In order to facilitate this process, we developed a database portal named miRToolsGallery by manual curation of approximately 1000 tools and resources. With miRToolsGallery, a user can easily locate the appropriate tool for an investigation by implementing a searchable and rankable query function. Tools are ranked by different criteria from PageRank algorithm to most recent publication date to number of publications. The portal contains tools covering a broad range of organisms including C. elegans. The tools also encompass an expansive range of applications from target identification to orthology search. Moreover, the tool is a rapid way to locate recent literature within the miRNA bioinformatics field and to identify current trends. One such trend identified with the tool was the stability and frequent use of some very old computational tools, despite the rapid and enormous advancement in knowledge of miRNA function. The resource is available at www.mirtoolsgallery.org. This research was supported in part by the Research grant of Garry Wong MYRG2016-00101-FHS.

Felkai S et al. (1998) East Coast Worm Meeting "Control of Respiration, Behavior and Aging by clk-1"

Hekimi S, Felkai S, Lemieux JY, Labbe J-C, Brown GG, Ewbank JJ

[East Coast Worm Meeting, 1998]

Mutations in the clk-1 gene of the nematode Caenorhabditis elegans affect the timing of a variety of developmental and physiological processes and increase life span (Wong et al., Genetics 139, 1247 (1995)). clk-1 encodes a protein with high similarity to yeast Coq7p as well as to predicted mammalian proteins (Ewbank et al., Science 275, 980 (1997)). Coq7p is required for respiration as it is necessary for the biosynthesis of the lipid co-factor coenzyme Q (CoQ) (also called ubiquinone). Indeed, no or very little CoQ is found in these mutants (Marbois and Clarke, J. Biol. Chem. 271, 2995 (1996)). We show that in worms CLK-1 is fully active when fused to Green Fluorescent Protein (GFP) and is found in mitochondria. We show that mitochondrial respiration is only mildly depressed in clk-1 mutants (~85% of the wild type). Furthermore, an excess of exogenous CoQ cannot restore normal respiration, suggesting that a deficiency in CoQ is not the main cause of the clk-1 phenotype. A transgene overexpressing CLK-1 activity in wild-type worms can increase respiration beyond the wild type level, accelerate behavioral rates (the defecation cycle) during aging and shorten life span, indicating that clk-1 controls these processes. These observations (that decreased respiration in clk-1 mutants results in a longer life and that increased respiration in worms overexpressing CLK-1 results in a shorter life) suggest that decreased mitochondrial respiration at older ages, which is observed in all animals, might be the result of an active process promoting longer life.