Aoki, Yuki et al. (2021) International Worm Meeting "Behavioral and Ca2+ imaging analysis of odor and temperature sensory integration in C. elegans"

Aoki, Yuki, Takeishi, Asuka

[International Worm Meeting, 2021]

Most of living organisms are exposed to multiple environmental stimuli at the same time in nature. When animals sense external stimuli of heterogeneous sources, they weigh and integrate the information in their nervous system to respond appropriately. It is essential for animals to make behavior choice for survival, however, the neural and molecular mechanism of multisensory integration is still largely unknown. C. elegans exhibits behavioral responses against various stimuli such as temperature and odors. Previous studies have revealed detail neural networks and signaling pathways that are required to produce behavior in response to each stimulus. To understand how the nervous system process multiple information, we are investigating the behavior and neural response of worms that are exposed to both temperature and odor stimuli at the same time. To examine the behavior choice of worms, we evaluated the direction of worm migration on the agar plate with thermal gradient and an odor. We also set up a microscope with auto-tracking stage that enable us to observe neural responses and locomotion of free-moving worms simultaneously with stimuli of temperature and an odor. We are currently analyzing the neural activities of sensory neurons and inter neurons in order to identify the neurons and the mechanisms that drive worm behavior. These results are expected to allow us to understand the detail neural mechanism of multi-information integration in a complicated environment.

Zugasti, Olivier et al. (2013) International Worm Meeting "Triggering antifungal innate immunity."

Schroeder, Frank, Bose, Neelanjan, Squiban, Barbara, Zugasti, Olivier, Pujol, Nathalie, Belougne, Jerome, Ewbank, Jonathan, Kurz, Leo

[International Worm Meeting, 2013]

Infection of worms by the fungus D. coniospora leads to the up-regulation of genes such as nlp-29, one of a cluster of six paralogous nlp antimicrobial peptide genes, via a PMK-1/p38 MAP kinase pathway. The worm genome, however, lacks orthologs of the receptors known to be important for pathogen recognition in other species. How D. coniospora triggers an immune response is an open question. For epidermal defenses, the p38 signaling cassette acts downstream of the Ga protein GPA-12, implicating a G-protein coupled receptor (GPCR) in the immune response. By RNAi, we knocked down individually three-quarters of the worm's >1,500 GPCR genes. Quantification with the COPAS Biosort of the effect of each RNAi clone on the expression of a nlp-29p::gfp reporter following infection allowed us to identify dcar-1 (dihydrocaffeic acid receptor) as a candidate innate immune receptor gene. dcar-1 mutants exhibit an almost complete block of nlp genes induction following infection and a heightened susceptibility to infection compared to wild-type worms. We found that dcar-1 is expressed in the major epidermal syncytium, hyp7, site of expression of the infection-inducible nlp genes. Restoring dcar-1 expression in the epidermis rescued nlp gene expression and the mutant's resistance to infection. DCAR-1 has been previously shown to have a high affinity for dihydrocaffeic acid (DHCA) a derivative of DOPA. We found that unlike DOPA, direct addition of DHCA or oxidized DOPA to worms triggers the expression of nlp genes but could not demonstrate the production in vivo of DHCA. This suggests that DHCA might not be the natural ligand of DCAR-1. Our current hypothesis is that a derivative of DOPA is rapidly produced upon infection and triggers an immune response via DCAR-1. We are currently applying genetic and biochemical techniques to identify the putative DCAR-1 ligand. Thanks to Christophe Melon, and Reina Aoki and Yoshio Goshima for reagents.

Hiromi Moriguchi et al. (2005) International Worm Meeting "A search for new components of the RNAi pathway in C. elegans"

Hiroaki Tabara, Kazuma Aoki, Hiromi Moriguchi, Katsuya Okawa

[International Worm Meeting, 2005]

RNAi is a form of sequence-specific gene silencing induced by the introduction of double-stranded RNA (dsRNA). The ability of dsRNA to induce silencing was first discovered in C. elegans, and similar responses have been observed in a variety of eukaryotes. Although genetic and biochemical studies in several model organisms have revealed a rough mechanism of the RNAi response, it is far from a complete understanding. With several approaches, we are searching for new components required for RNAi. In C. elegans, genetic factors required for RNAi have been revealed by screens for RNAi deficient (rde or other names) mutants. In original screens, rde-1 and sid-1 mutations on LG V were recovered very frequently, and therefore it was not easy to collect new rde mutations reflecting new loci. In order to avoid this problem, we performed a genetic screen utilizing nT1 which is a balancer chromosome of LG V (EAWM 2004, ab112). The presence of nT1 prevents from recovering recessive mutaions on the balanced chromosomal region. Among total 51 mutants isolated from this and similar screens, we are currently focusing and analyzing mutants whose RNAi responses are severely deficient. The mutant stocks include rde-3 (one allele), mut-16 (one allele), rrf-1 (two alleles), rde-4 (10 alleles) as well as several new mutants possibly reflecting new components in the RNAi pathway. In addition, we are trying to identify proteins interacting with known RNAi factors (such as RDE-1) to find candidates for new RNAi factors (see also Aoki et al., this meeting). For this purpose, we generated several transgenic worms expressing the known RNAi factors attached with an epitope tag. Utilizing the epitope tag, we next immuno-purified the RNAi factor complex and analyzed the protein components of the complex with mass spectrometry. From these experiments, a protein possessing DExD/H helcase domain was identified. We isolated a deletion mutant lacking the helicase gene (with the kind help of Nishiwaki and Kuroki) to investigate its function. This mutant (fj52) is sterile showing a defect in oogenesis. Currently, we are analyzing the detailed phenotype of fj52.