Conroy, Brian et al. (2013) International Worm Meeting "The neuronal basis of food choice behavior in Caenorhabditis elegans."

Morabe, Maria, Chambers, Melissa, Conroy, Brian, Macfarlane, Rachel, Haynes, Lillian, Glater, Elizabeth

[International Worm Meeting, 2013]

Caenorhabditis elegans uses chemosensation to distinguish among various species of bacteria, their major food source (Ha et al., 2010; Shtonda and Avery, 2006). Although the neurons required for the detection of specific food-odors have been well-defined (Bargmann, 2006), less is known about the sensory circuits underlying the discrimination among the mixtures of odors released by bacteria. We plan to examine the neural machinery underlying bacterial preference among a diverse set of bacterial species. Does bacterial choice use one common neuronal mechanism or a diversity of mechanisms depending on the bacteria? Do some bacterial choices involve a single sensory neuron and others involve multiple sensory neurons? To address these questions, we are testing the food preferences of C. elegans for bacteria found in their natural habitats (kindly provided by Marie-Anne Felix, Institut Jacques Monod, Paris, France). We have found that C. elegans strongly prefers the odors of Providencia sp., Alcaligenes sp., and Flavobacteria sp., to Escherichia coli HB101, a commonly used food source for C. elegans. We have identified that the olfactory neuron AWC is necessary for this preference. We intend to test whether other amphid sensory neurons are also necessary for bacterial preference. In the future we will extend our analysis to other bacterial species to determine the diversity of the underlying neuronal mechanisms. Bargmann, C.I. (2006). http://www.wormbook.org. Ha, H.I., Hendricks, M., Shen, Y., Gabel, C.V., Fang-Yen, C., Qin, Y., Colon-Ramos, D., Shen, K., Samuel, A.D., and Zhang, Y. (2010). Neuron 68, 1173-1186. Shtonda, B.B., and Avery, L. (2006). J Exp Biol 209, 89-102.

Katz, Menachem et al. (2009) International Worm Meeting "CEP-sheath glia are required for normal locomotion in C. elegans."

Katz, Menachem, Lu, Yun, Shaham, Shai

[International Worm Meeting, 2009]

C. elegans glial cells share morphological, functional and genetic resemblance to their vertebrate counterparts. However, in contrast to vertebrate glia, C. elegans glia are not essential for neuronal survival, offering a unique opportunity to study the involvement of glia in the development and function of the nervous system in vivo. C. elegans CEP sheath glia (CEPsh) are bipolar cells that ensheath the dendrites of CEP neurons, and also envelope the nerve ring (the animal''s brain) extending processes that abut synapses. During development, these cells are involved in regulating axon guidance, dendrite extension, formation of the nerve ring (Yoshimura et al., 2008), and synaptogenesis (Colon-Ramos et al., 2007). However, post-developmental roles for CEPsh glia have not been studied. Here we demonstrate that animals in which the CEPsh glia were genetically ablated during the first larva stage display various motor defects including reduced speed of locomotion and intermittent periods of extended paralysis. Ablated animals also display high levels of small-angle turning, resulting in a spiral pattern of movement that restricts their movement to small areas on the plate. EM analyses of CEPsh glia-ablated animals reveal disorganization of the nerve ring, and swelling of neurons in the ventral ganglion. Thus CEPsh glia may play key homeostatic roles in the neuropil. To understand the molecular basis of CEPsh glia function we have begun to identify genes expressed in these cells using an mRNA-tagging method, with the aim of examining the roles of enriched genes in C. elegans locomotory behavior.

Magalska A et al. (2000) European Worm Meeting "The worm is not enough - the construction of cDNA and gDNA libraries from Ancylostoma ceylanicum"

Kapulkin V, Ciesielski A, Wedrychowicz HZ, Magalska A

[European Worm Meeting, 2000]

We are interested in usage of C. elegans as system for expression of hookworm genes. Hookworms (Ancylostomatideae ) are economically and medically important blood feeding nematodes which cause intestinal infections in humans and domestic animals. It has been reported recently that above 1 billion of human beings are currently infected with these worms. The application of C. elegans as a heterologous host for hookworm genes is limited mainly by the lack of hookworm genomic data. For this purpose we have constructed cDNA and gDNA libraries from the Ancylostoma ceylanicum -important human/animal parasite related to C. elegans. The cDNA library is based on PCR mediated amplification of spliced leader of parasite cDNA reported previously ( Martin SA, Thompson FJ, Devaney E. Mol. Biochem. Parasitol. 1995 Mar;70(1-2):241-5.). PCR amplified cDNA has been gel fractionated and cloned. The most abundant library is available now from adult mixed sex A. ceylanicum (above 6000 clones), smaller library was constructed from infective L3 larvae. The large inserts gDNA libraries were constructed by ligation of PFGE fractionated Ancylostoma genomic DNA into the pBACe3.6 (provided by Pieter deJong), both digested with EcoRI. Ligation mixture was precipitated with ethanol in the presence of yeast tRNA prior electroporation into DH10B host. Both cDNA and gDNA libraries were arrayed in 96 well format, and stored at -70C. Randomly picked clones were sequenced from both ends, blastn and blastx searches were performed. For cDNA clones our sequencing results show clear similarities to C. elegans genes : gene F38E11.2 hsp-12.6; similar to Human Alpha crystallin B chain, gene C15C7.6; similar to syntaxin-6, gene T20F5.2; similar to the S25B family of peptidases. For BAC clones, end sequencing was also performed ( in collaboration with M. obocka, Institute of Biochemistry and Biophysics ) , however no significant similarities were found (yet!).

Garriga G et al. (2000) West Coast Worm Meeting "Role of the Caenorhabditis elegans homologs of cdk5 and p35 in migration and axon outgrowth"

Harbaugh TE, Garriga G

[West Coast Worm Meeting, 2000]

While cdk5 was originally identified in vertebrates based on its homology to the cell cycle regulated cdc2, further study has demonstrated a function not in the cell cycle but instead in post-mitotic neurons. Mouse knockouts of cdk5 and its activator, p35, result in defects in the pattern of neuronal migration in the cortex (1,2). Studies in cultured rat neurons demonstrated that the cdk5/p35 kinase activity can promote neurite outgrowth (3). The C. elegans homolog of cdk5 is 74% identical to mouse, and the p35 homolog contains a 159 AA region that is 54% identical. Translational GFP reporter constructs for cdk5 and p35 are expressed in the cytoplasm of most neurons beginning around the two-fold stage of embryogenesis and continuing into adulthood. Simultaneous overexpression of both cdk5 and p35 produces a strongly uncoordinated phenotype, with defects in fasciculation and axon pathfinding. Along with the expression pattern, these phenotypes suggest a role for cdk5 and p35 in neuronal development. The mig-18(k140) mutation was identified based on defects in distal tip cell migration (4) and may represent a Ce-cdk5 allele. Extrachromosomal arrays containing the Ce-cdk5 genomic region can partially rescue the distal tip cell migration defect of mig-18(k140). Furthermore, sequencing of the Ce-cdk5 genomic region revealed a point mutation predicted to change a conserved histidine into a tyrosine. To confirm that k140 is a Ce-cdk5 allele, we plan to isolate additional mig-18 mutants and determine whether they contain Ce-cdk5 molecular lesions. Also, we will screen a deletion library for alleles of Ce-cdk5 and its activator Ce-p35. (1) Gilmore EC, Ohshima T, Goffinet AM, Kulkarni AB, Herrup K, J Neurosci, 18:6370-7 (1998) (2) Chae T, Kwon YT, Bronson R, Dikkes P, Li E, Tsai LH, Neuron, 18:29-42 (1997) (3) Nikolic M, Dudek H, Kwon YT, Ramos YF, Tsai LH, Genes Dev, 10:816-25 (1996) (4) Nishiwaki K, Genetics, 152:985-97 (1999)

Carolina Ibanez-Ventoso et al. (2007) International Worm Meeting "MicroRNAs in Aging and Healthspan."

Carolina Ibanez-Ventoso, Monica Driscoll, Mehul Vora

[International Worm Meeting, 2007]

Over 200 mutations that affect C. elegans longevity have been identified. To date, much of the research on C. elegans aging has focused on factors that extend lifespan with less emphasis on their effects on healthspan (the period of mid life vigor that precedes decline). Our lab has been interested in identifying genes that extend healthspan, conferring longer periods of youthful behavior. We have identified genetic and pharmacological manipulations that can extend healthspan with the use of new aging biomarkers and assays we''ve developed: decline of muscle integrity (physical markers and automated computer vision swimming analysis), age-associated accumulation of autofluorescent cellular material (lipofuscin and age pigments), and micoRNA (miRNA) abundance. miRNAs are small molecules prevalent in metazoans that regulate cellular expression by binding to partially homologus sites on gene transcripts to down-regulate translation. Our interest in these broad conserved regulators has been influenced by the suggestion that they might function in biology to limit stochastic biological processes—lost of robustness and increase in stochastic events is associated with aging. To address whether miRNAs could influence the aging process, we conducted the first genome wide-analysis of how miRNAs change with age in any organism. This work showed that about half of C. elegans miRNAs change significantly during adult lifespan and could thus contribute to healthspan [1]. Indeed the miRNA lin-4 influences lifespan and the expression of healthspan indicators like lipofuscin [2], work we have confirmed. Our preliminary work supports that several other miRNAs also impact lifespan and the quality of aging. We are exploiting the powerful experimental advantages of the nematode C. elegans to decipher roles of miRNAs in nematode healthspan and to test whether engineered expression of these small molecules can confer anti-aging therapies in C. elegans. Since many miRNAs are conserved in expression pattern and function, we anticipate that our exploratory work in this area may suggest specific working hypotheses for mammalian aging, perhaps influencing design of novel means of clinical intervention. 1.Ibanez-Ventoso et al. (2006) Modulated microRNA expression during adult lifespan in C. elegans. Aging Cell 5: 235-246. 2.Boehm M, Slack FJ (2005) A developmental timing microRNA and its target regulate life span in C. elegans. Science 310: 1954-1957.

Peedikayil Kurien, Sonu et al. (2021) International Worm Meeting "Integration of neuronal connectivity, activity and synaptic plasticity drives sexually dimorphic learning in C. elegans"

Oren-Suissa, Meital, Peedikayil Kurien, Sonu

[International Worm Meeting, 2021]

Sexually dimorphic traits stemming from distinct processing of environmental cues bring about the holistic survival of the species and are mostly driven by sexual selection. However, the underlying mechanisms driving sex-dependent complex decisions remain largely unknown. Previous studies have shown that C. elegans hermaphrodites are able to shift their preference from attraction to aversiveness to a specific pathogenic bacterium Pseudomonas aeruginosa (PA14), after certain hours of exposure (Zhang et al, 2005). Using a modified version of this learned avoidance behavior, we show that C. elegans makes sexually dimorphic choices based on short past experiences. The hermaphrodites learn efficiently, as was previously shown. However, males do not learn to avoid PA14, even after training, thus retaining their initial preference. Using a blind testing paradigm, we show that larval stages of both sexes are unable to shift the preference, suggesting that maturation of nervous system is necessary for short term experience-based plasticity. By performing fast killing assays, we show that the lethality kinetics toward PA14 toxins is similar in the sexes, thus ruling out dimorphic susceptibility. To understand tissue specificity in developing a learned response, tissue specific sex-reversal experiments were performed and the importance of the nervous system was unraveled. In the same vein, we show that short term exposure does not involve signaling from the gut. Previous work has shown the importance of the interneuron AIY within several learning circuits (Ha et al, 2010; Jin et al, 2016). By analyzing AIY synaptic activity using fluorescently labelled RAB-3, we show that, upon short-term exposure, there is a decrease in synaptic activity, specifically, in an axonal region that forms connections with the interneuron RIA (Colon-Ramos et al, 2007). While this trend was prevalent in both the sexes, close examination of Ca2+ fluxes in the same region, using calcium indicators, reveal dimorphic responses to the odor of PA14. These results suggest that downstream encoding of short-term learning is sex-dependent and thus, its neuronal connectivity. To summarize, we present a model in which dimorphic processing of environmental cues, in a sex-specific manner, evokes diverse behavioral phenotypes that help shape the fitness of the species.

TE Harbaugh et al. (1999) International C. elegans Meeting "Role of the Caenorhabditis elegans homologs of cdk5 and p35"

TE Harbaugh, G Garriga

[International C. elegans Meeting, 1999]

While cdk5 was originally identified in vertebrates based on its homology to the cell cycle regulated cdc2, further study has demonstrated a function not in the cell cycle but instead in post-mitotic neurons. Mouse knockouts of cdk5 and its activator, p35, result in defects in the pattern of neuronal migration in the cortex (1,2). Studies in cultured rat neurons demonstrated that the cdk5/p35 kinase activity can promote neurite outgrowth (3). At this point, however, little is known about the upstream activators and downstream targets of the cdk5/p35 kinase. The Caenorhabditis elegans homolog of cdk5 is 74% identical to mouse, and the p35 homolog contains a 159 aa region that is 54% identical. Translational GFP reporter constructs for cdk5 and p35 are expressed in the cytoplasm of most neurons beginning around the two-fold stage of embryogenesis and continuing into adulthood. To produce a loss of function phenotype, we injected dsRNA for either cdk5, p35, or both into N2 worms. In each case, most of the progeny had no gross visible phenotype, although a few Dpy Unc worms were produced. To identify loss of function mutations in cdk5 and p35, we will also screen a deletion library by PCR. To generate an overexpression phenotype, DNA containing the cdk5 or p35 genomic region was injected into N2 worms to produce extrachromosomal arrays. Worms carrying a cdk5 array (40 microg/ml) have no visible phenotype, and worms carrying a p35 array (40 microg/ml) are occasionally slightly uncoordinated (Unc). Worms carrying an array with both cdk5 and p35, however, are strongly Unc. These worms have defects in fasciculation and axon pathfinding. Along with the expression pattern, this strongly suggests a role for cdk5 and p35 in neuronal development. We will screen for suppressors of this Unc phenotype to identify other genes that are acting in the cdk5/p35 pathway. (1) Gilmore EC, Ohshima T, Goffinet AM, Kulkarni AB, Herrup K, J Neurosci, 18:6370-7 (1998) (2) Chae T, Kwon YT, Bronson R, Dikkes P, Li E, Tsai LH, Neuron, 18:29-42 (1997) (3) Nikolic M, Dudek H, Kwon YT, Ramos YF, Tsai LH, Genes Dev, 10:816-25 (1996)