ayyadevara, srinivas et al. (2019) International Worm Meeting "A novel microtubule-binding drug attenuates and reverses protein aggregation in animal models of Alzheimer's disease."

Kakraba, Samuel, Alla, Ramani, Penthala, Narsimha Reddy, Griffin, Sue, Ganne, Akshatha, Barger, Steven, Ayyadevara, Srinivas, Liu , Ling, Shmookler Reis, Robert, Balasubramaniam, Meenakshisundaram, Bommagani, Shoban Babu, Crooks, Peter

[International Worm Meeting, 2019]

Age-progressive neurodegenerative pathologies, including Alzheimer's disease, are distinguished and diagnosed by disease-specific components of intra- or extra-cellular aggregates. Increasing evidence suggests that neuroinflammation promotes protein aggregation, and is involved in the etiology of neurological diseases. We synthesized and tested analogues of the naturally occurring tubulin-binding compound, combretastatin A-4. One such analogue, PNR502, markedly reduced the quantity of Alzheimer-associated amyloid aggregates in the BRI-A?1-42 mouse model of Alzheimer's disease, while blunting the ability of the pro-inflammatory cytokine IL-1? to raise levels of amyloid plaque and its protein precursors in a neuronal cell-culture model. In transgenic C. elegans strains that express human A?1-42 in muscle or neurons, PNR502 rescued A?-induced disruption of motility (3.8-fold, p<0.0001) or chemotaxis (1.8-fold, p<0.05), respectively. Moreover, in C. elegans with neuronal expression of A?1-42, a single day of PNR502 exposure reverses the chemotaxis deficit by 54% (p<0.01), actually exceeding the protection from longer exposure. Moreover, continuous PNR502 treatment also extends nematode lifespan 23% (p?0.001). Given that PNR502 can slow, prevent, or reverse Alzheimer-like protein aggregation in human-cell-culture and animal models, and that its principal predicted and observed binding targets are proteins previously implicated in Alzheimer's, we propose that PNR502 has therapeutic potential to inhibit cerebral A?1-42 aggregation and prevent or reverse neurodegeneration.

Yemini, E.I. et al. (2017) International Worm Meeting "Whole-Brain Imaging with Single-Neuron Identity in C. elegans."

Venkatachalam, V., Yemini, E.I., Hobert, O., Samuel, A.D.

[International Worm Meeting, 2017]

Stereotypy of the C. elegans nervous system affords single-neuron registration across animals, and consequently, robust statistics for neurobehavioral coding and transcriptomics. Fast methods of whole-brain imaging in worm exist, but unfortunately determining the identity of neurons within these volumes remains a bottleneck - requiring a long, difficult, ad hoc process. We have developed a landmarked strain for whole-brain neural identification, NeuroPAL (a Neuronal Polychromatic Atlas of Landmarks). Each neuron is assigned an invariant fluorophore barcode such that color and position specify unambiguous neural identity via the unique 3-tuple (color, position, ganglion). The GFP channel is preserved for reporters of neural activity (GCaMP) and transcriptomics (GFP). Our landmark strain employs 5 fluorophores. To visualize this strain we have used a new microscope, developed by the Samuel lab, with the capability of imaging 9 unique fluorescence channels generated by 4 excitation lines and 4 emission bands. This microscope acquires whole-brain volumes, of 4 fluorophores, simultaneously, at 10Hz. Together, these two innovations permit fast whole-brain imaging, with single-neuron identity and neuronal registration, across an animal populace.

Minniti AN et al. (1995) International C. elegans Meeting "THE HERMAPHRODITE-INITIATED SPERMIOGENESIS PATHWAY."

Minniti AN, Nance JF, Sadler C, Ward S

[International C. elegans Meeting, 1995]

During spermiogenesis, round nonmotile spermatids are rapidly transformed into asymmetrical crawling spermatozoa. In addition to spe-8 and spe-12, we found two new genes, spe-27 and spe-29, that when mutated, disrupt spermiogenesis in an identical manner: mutant hermaphrodites are self-sterile, while mutant males are fertile. Sperm from both sexes activate abnormally in vitro, suggesting that these gene products are expressed in both sperm, but only hermaphrodites require them for activation. The hermaphrodite's spermatids, however, can be activated by mating. This phenotype has been explained by a model that has two distinct pathways of activation, one for males and one for hermaphrodites (Shakes and Ward, 1989). The four genes are needed only for the hermaphrodite pathway. spe-27 and spe-12 have been cloned. Their mRNA is found exclusively in the germline during spermatogenesis. Their predicted protein sequences show no similarities to known proteins. We are currently cloning spe-29. To further understand how these genes act during spermiogenesis we are determining the subcellular localization of their gene products. In addition, suppressor analysis is under way (see abstract by Paul Muhlrad and Samuel Ward) to look for other genes in this pathway and for interactions between these genes.

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.

Adam Brown et al. (2003) International Worm Meeting "Genetic analysis of AFD thermosensory neuron function in C. elegans."

Adam Brown, Piali Sengupta

[International Worm Meeting, 2003]

Thermosensation plays an important role in the development, metabolism, and behavior of animals. C. elegans can be used to genetically dissect thermosensation, as they demonstrate distinct behaviors in response to temperature gradients. Genetic and laser ablation experiments have identified a circuit of neurons responsible for this behavior, including the primary temperature sensing neuron type, AFD. However, little is known about how AFD mediates its functions, as well as how worms form the "memory" of their cultivation temperature that modulates this behavior. Previously, a screen for mutations that alter the expression of a fusion gene gcy-8::gfp expressed specifically in the AFD neuron pair was performed in our lab to investigate AFD development and function. This screen led to the identification of the Otx/Otd-like homeobox transcription factor gene ttx-1, which is necessary and sufficient to specify AFD fate (Satterlee et al, 2001), and the cmk-1 CaMKI ortholog (see abstract by Satterlee and Sengupta). We are currently continuing this screen to identify further components of the AFD developmental and sensory pathways. We have screened the progeny of over 25,000 haploid genomes and identified a number of mutants. It is expected that a subset of these mutants may define additional genes required for both the function and development of the AFD neurons. Additionally, we also plan to carry out thermosensory behavioral screens to identify mutants with defects in distinct aspects of thermosensation. Finally, in collaboration with Will Ryu and Aravi Samuel (Rowland Institute, Harvard University), we have initiated behavioral analyses of both new and existing signaling mutants in an attempt to further dissect the neuronal pathways and molecules required for thermosensory behaviors.

Urquiza, S. et al. (2019) International Worm Meeting "Natural microbiome of Chilean C. elegans isolates and their relationship with neuroprotection."

Calixto, A., Urrutia, A., Urquiza, S.

[International Worm Meeting, 2019]

In nature Caenorhabditis elegans eats bacteria from rotting fruits and vegetation. The natural microbiome of C. elegans influences growth, lifespan and stress (Dirksen et al., 2016. Zhang et al,. 2017. Samuel et al., 2017). We are interested in understanding how the relationship between C. elegans and its intestinal bacteria influences behavior and phenotype. We showed that specific metabolites from laboratory bacteria affect the rate at which mechanosensory neurons (MN) degenerate in a genetic model of neuronal necrosis (Urrutia et al., under review). The bacterial enzyme involved in MN neuroprotection is the glutamate decarboxylase enzyme (GAD) and the protective metabolite, its product g-amino butyric acid (GABA). In this work we studied the impact of a natural microbiome in neurodegeneration of the MN. The natural microbiome we studied was isolated from wild C. elegans obtained from the Universidad Mayor Campus in Huechuraba, Chile. This microbiome was composed of three different culturable bacteria identified by means of ribosomal RNA 16S marker sequencing. Isolate 1 has 100% identify with Brucellaceae bacterium bfzh12, Pseudochrobactrum sp. AR-333 and Pseudochrobactrum kiredjianiae. Isolate 2 has 100% identity with Stenotrophomonas humi and Stenotrophomonas sp. Isolate 3 has 100% identity with Bacillus pumilus and Bacillus altitudinis. GAD enzymatic activity provided information about the bacteria neuroprotective potential because all isolates displayed higher levels than E. coli OP50 (10%, 15% and 40% respectively). Functional morphological categories of the MN were significantly higher in animals feeding natural isolates than on E. coli OP50, suggesting a correlation with GABA production. Ongoing and future work include dissecting the contribution in abundance of each bacterium to the final intestinal microbiome as well as the metabolite production of individual bacteria and in conjunction.

Nikhil Bhatla et al. (2008) C.elegans Neuronal Development Meeting "Reduction of Movement by flp-11, which Encodes FMRFamide-related Peptides"

Bob Horvitz, Nikhil Bhatla, Niels Ringstad

[C.elegans Neuronal Development Meeting, 2008]

FMRFamide-related peptides (FaRPs) make up a large and diverse family of C. elegans neuropeptides. To learn how these peptides control C. elegans behavior, we overexpressed genes predicted to encode FaRPs (the flp genes) and characterized the resulting behavioral defects. Overexpression of flp-11 from an integrated transgene greatly reduces locomotion compared to the wild type. We quantified this reduction in movement using a wormtracker (developed by Damon Clark, Aravi Samuel, and Dan Omura) and found that the average speeds of flp-11 transgenic strains are less than 50% of the wild-type speed. We also quantified locomotion by body bend counts: flp-11 overexpressors execute 70% fewer body bends per unit time than the wild type. The overexpressors lay motionless more often and, when they are moving, move more slowly than normal. To identify genes involved in the flp-11 overexpression phenotype, we screened EMS-mutagenized flp-11 overexpressors for suppressors of the locomotion defect. An F2 non-clonal screen of 20,000 haploid genomes yielded 1 suppressor, and an F1 clonal screen of 2,500 haploid genomes isolated 2 suppressors. These suppressors all exhibit average speeds similar to the wild-type speed. We are currently mapping these 3 suppressors. We are also planning to measure the speed of a flp-11 deletion mutant. To identify neural circuits that use flp-11 to control behavior, we have constructed a flp-11::GFP translational fusion reporter gene. Transgenic worms show GFP expression in a single cell near the posterior bulb of the pharynx. We plan to characterize the functions of flp-11-expressing cells by laser-ablation experiments. We are also searching for flp-11 receptors through our suppressor screens and by testing candidate genes. Identification of flp-11 receptors will help us identify the sites of action of flp-11 neuropeptides.

Kissoyan, K.A.B. et al. (2019) International Worm Meeting "Natural C. elegans Microbiota Protects against Infection via Production of a Cyclic Lipopeptide of the Viscosin Group."

Dierking, K., Kissoyan, K.A.B.

[International Worm Meeting, 2019]

C. elegans is associated in nature with a species-rich, distinct microbiota, which was characterized only recently [1]. Our understanding of C. elegans microbiota function is thus still in its infancy. Here, we identify natural C. elegans microbiota isolates of the Pseudomonas fluorescens subgroup that increase C. elegans resistance to pathogen infection. We show that different Pseudomonas isolates provide paramount protection from infection with the natural C. elegans pathogen Bacillus thuringiensis through distinct mechanisms [2] . The P. lurida isolates MYb11 and MYb12 (members of the P. fluorescens subgroup) protect C. elegans against B. thuringiensis infection by directly inhibiting growth of the pathogen both in vitro and in vivo. Using genomic and biochemical approaches, we demonstrate that MYb11 and MYb12 produce massetolide E, a cyclic lipopeptide biosurfactant of the viscosin group, which is active against pathogenic B. thuringiensis. In contrast to MYb11 and MYb12, P. fluorescens MYb115-mediated protection involves increased resistance without inhibition of pathogen growth and most likely depends on indirect, host-mediated mechanisms. We are currently investigating the molecular basis of P. fluorescens MYb115-mediated protection using a multi-omics approach to identify C. elegans candidate genes involved in microbiota-mediated protection. Moreover, we are further exploring the antagonistic interactions between C. elegans microbiota and pathogens. This work provides new insight into the functional significance of the C. elegans natural microbiota and expands our knowledge of immune-protective mechanisms. 1. Zhang, F., Berg, M., Dierking, K., Felix, M.A., Shapira, M., Samuel, B.S., and Schulenburg, H. (2017). Caenorhabditis elegans as a model for microbiome research. Front. Microbiol. 8:485. 2. Kissoyan, K.A.B., Drechsler, M., Stange, E.-L., Zimmermann, J., Kaleta, C., Bode, H.B., and Dierking, K. (2019). Natural C. elegans Microbiota Protects against Infection via Production of a Cyclic Lipopeptide of the Viscosin Group. Curr. Biol. 29.

Vidal-Gadea A G et al. (2010) Neuronal Development, Synaptic Function and Behavior, Madison, WI "Biogenic amines mediate transition between crawling and swimming in C. elegans"

Brauner M, Gottschalk A, Erbguth K, Kressin L, Vidal-Gadea A G, Topper S, Young L, Pierce-Shimomura J T

[Neuronal Development, Synaptic Function and Behavior, Madison, WI, 2010]

A wide variety of animals must quickly adjust their pattern of locomotion to successfully navigate through different environmental niches. Selection and execution of the appropriate locomotory pattern is therefore paramount to survival. Although C. elegans is capable of performing many adaptive behaviors, it has been controversial whether forward crawling and swimming represent distinct gait-like forms of locomotion or the modulation of a single form of locomotion [1-3]. Biogenic amines have been shown to mediate the transition between gait-like forms of locomotion across taxa as diverse as sea slugs, leeches, lampreys and humans. We previously reported that C. elegans crawls and swims with distinct kinematics and different patterns of muscle activity [2]. We now combine quantitative behavioral analysis, optogenetic tools and neuronal ablation to show that C. elegans uses biogenic amines to switch between crawling and swimming in a gait-like manner. As in other invertebrates, we find that serotonin mediates the smooth transition from crawling to swimming in C. elegans. Serotonin is further required to inhibit motor behaviors (e.g. foraging and pharyngeal pumping) during swimming that normally only accompany crawling. Mirroring the role of dopamine in other invertebrates, C. elegans uses dopamine to successfully initiate and maintain crawling when emerging from liquid. Over 600 million years of separate evolution notwithstanding, the highly conserved role played by biogenic amines such as dopamine and serotonin across taxa attests to how vital their function is to adaptive strategies for locomotion. Korta J, Clark DA, Gabel CV, Mahadevan L, Samuel AD. J. Exp. Bio. 2007 210:2383-9.Pierce-Shimomura JT, Chen BL, Mun JJ, Ho R, Sarkis R, McIntire SL. PNAS. 2008 105:20982-7.Berri S, Boyle JH, Tassieri M, Hope IA, Cohen N. HSFP J. 2009 3:186-93.

Narayan, Anusha et al. (2009) International Worm Meeting "Transfer at a thermosensory synapse in C. elegans."

Laurent, Gilles, Sternberg, Paul, Narayan, Anusha

[International Worm Meeting, 2009]

C.elegans is an attractive system for neural circuit analysis. To analyze the functional dynamics of circuits that control behavior, it is necessary to understand the underlying synaptic transformations. Thermotaxis is an established behavior in C. elegans[1-2]. We attempt to characterize the transfer function at a prominent synapse within the thermotactic circuit: between AFD, the primary thermosensory neuron[3], and AIY, its principal post-synaptic partner. We drive expression of Channelrhodopsin-2(chR2), a light-activated cation channel [4], solely in AFD, using a cell-specific promoter, and use whole-cell patch-clamp recording techniques to measure the light-evoked synaptic response at AIY. We are able to reliably activate the presynaptic cell AFD, evoking depolarizing potentials of up to 40 mV and inward currents of up to 15 pA. The postsynaptic response at AIY is small: less than 5 mV, with inward currents of less than 1 pA, and is graded and tonic, lasting the duration of the stimulus. The response reverses around 0 mV and seems to be frequency independent, showing no obvious short-term facilitation or depression. Our results further validate the use of chR2 to stimulate neural activity, and indicate that this synapse has low gain, and transmits information from AFD to AIY with short latencies and high fidelity. It will be interesting to see how AIY integrates this information with other incoming streams, and to examine processing downstream in the thermotactic circuit. 1.Mori, I., H. Sasakura, and A. Kuhara, Worm thermotaxis: a model system for analyzing thermosensation and neural plasticity. Curr Opin Neurobiol, 2007. 17(6): p. 712-9. 2.Ryu, W.S. and A.D. Samuel, Thermotaxis in Caenorhabditis elegans analyzed by measuring responses to defined Thermal stimuli. J Neurosci, 2002. 22(13): p. 5727-33. 3.Clark, D.A., et al., The AFD sensory neurons encode multiple functions underlying thermotactic behavior in Caenorhabditis elegans. J Neurosci, 2006. 26(28): p. 7444-51. 4.Boyden, E.S., et al., Millisecond-timescale, genetically targeted optical control of neural activity. Nat Neurosci, 2005. 8(9): p. 1263-8.