Slice LW et al. (1989) International C. elegans Meeting "isolation and characterization of cadmium-induced metallothionein-like protein and cDNA from C elegans."

Slice LW, Freedman JH, Rubin CS

[International C. elegans Meeting, 1989]

Freedman JH et al. (1991) International C. elegans Meeting "MOLECULAR CHARACTERIZATION OF HEAVY METAL STRESS IN C. ELEGANS"

Linder B, Fire A, Slice LW, Dixon DK, Freedman JH, Rubin CS

[International C. elegans Meeting, 1991]

Caenorhabditis elegans provides a model system for investigating developmental and molecular mechanisms underlying adaptations to environmental stresses. We examined C. elegans adapted for growth in toxic concentrations (> 0.2mM) of Cd. Physiological responses to this persistent stress were characterized by studying the properties and expression of metallothioneins (MTs) and a group of stress-induced proteins (SIPs) that accumulate in Cd-treated nematodes. C elegans showed elevated expression of two iso-MTs, MT-l (7.9 kDa) and MT-2 (6. 4 kDa), that are 65% identical, but have no significant overall sequence homology with the conserved MTs from other species. The gene encoding MT-2 (designated mtl-2) comprises 330 bp and contains a single intron. The DNA flanking the 5'-end of mtl-2 has putative consensus sequences for a cAMP response element (CRE), oct-l, cjun and a metal regulatory element (MRE). The mtl-l gene is organized similarly; it is 440 bp and contains S'-CRE and oct-l but surprisingly, lacks an MRE. The mtl-2 promoter /enhancer was fused with the , Bgalactosidase (,B-gal) reporter gene and introduced into stable transgenic worms. High-level expression of ,B-gal was selectively induced in intestinal cells by Cd. This suggests that a single cell type might confer the initial line of defense against heavy metal toxicity. Three SIP cDNAs were isolated by differentially screening a cDNA library corresponding to mRNA templates from Cd-treated C. elegans. SIP-l encodes a novel low Mr 'stress protein' related to C. elegans heat shock proteins; SIP-2 encodes a putative secreted proteolytic aspartyl proteinase-like enzyme; SIP-3 encodes the catalytic (C) subunit of protein kinase A. The activity, mass and mRNA for C are increased 10-20-fold in Cd-adapted C. elegans. These results suggest that resistance to Cd toxicity in C. elegans is a complex process that entails the coordinated regulation of the expression of numerous genes. The atypical elevation in C and the CRE elements upstream from mtl-l and mtl-2 raises the possibility that the persistent activation of PKA is essential for a coordinated, adaptive transcriptional response to Cd.

Laura Herndon et al. (2008) C.elegans Aging, Stress, Pathogenesis, and Heterochrony Meeting "What's New on WormAtlas and WormImage?"

Zeynep Altun, Tylon Stephney, Carolyn Norris, Laura Herndon, David Hall, Chris Crocker

[C.elegans Aging, Stress, Pathogenesis, and Heterochrony Meeting, 2008]

We are updating the WormAtlas website to give it a fresh look. This is the first major revamping of WormAtlas since its launch in 2002. These changes will start from the front page and then be implemented throughout the Handbook and many other portions of the website. Here we will explain the principles of the new organization, and show you how to find your favorite features. We hope you will find the site simpler to navigate and we expect it will be more intuitive for beginners. Inside the WormAtlas website, there will be several major changes. First will be an improved adult hermaphrodite handbook; it will include several completely revised chapters and a new one covering the nervous system. Second will be the launch of a handbook for anatomy of the worm embryo. Third will be the addition of more data to our slice-by-slice worm viewer, Slidable Worm. Lastly, we will be adding many new Neuron pages for the male nervous system in order to highlight new synaptic patterns emerging from the Wired Worm project conducted together with Scott Emmons. The WormImage website, which houses thousands of unpublished electron micrographs and related data, is expanding steadily. It now presents much more data from mutant animals, particularly for genes affecting the nervous system. We continue to rely heavily on MRC datasets, but we are also adding more from the Riddle and Hall lab files, among others. We encourage more laboratories to share their own best archival TEM and SEM images so that this resource can continue to grow and serve the C. elegans community. We are very grateful to many labs that have already contributed ideas, advice and experimental results that are featured on these websites. This work is supported by NIH RR12596.

Ebbing, A.L.P. et al. (2017) International Worm Meeting "Serial-section RNA sequencing: a novel method for genome-wide gene expression pattern analysis in C. elegans."

Ebbing, A.L.P., Korswagen, H.C., van Oudenaarden, A., Betist, M.C., Vertesy, A.

[International Worm Meeting, 2017]

To advance our understanding of development and physiology, it is important to gain detailed insight into the spatial and temporal expression patterns of all genes. Such a comprehensive overview of gene expression patterns will identify cell type and organ specific genes. It will also provide a framework for studying how gene expression patterns change upon perturbation. RNA sequencing (RNA-seq) is ideally suited for genome-wide analysis of gene expression. However, RNA-seq by itself does not provide information on where genes are expressed in the organism. A solution to this problem is to include spatial information by sectioning the organism along one of the body axes and to perform RNA-seq on each of the separate sections. We have developed a serial-section RNA-seq (serial-seq) method for C. elegans. In brief, we cryo-section a young adult animal from the head to the tail into 20 micrometer thick slices, resulting in a total of 40 to 50 slices. Each slice is then sequenced using the sensitive CEL-seq method and the sequencing data is aligned to create an anteroposterior gene expression map. Using this approach, an average of 8794 genes (43% of coding genes) (n=8) can be detected along the length of the animal. These include genes that are expressed in major tissues such as the germline, but also genes that are known to be expressed in only single neuron pairs in the head, demonstrating the high sensitivity of our approach. Furthermore, we found that these expression patterns are highly reproducible among independently sectioned animals. What makes serial-seq particularly powerful in C. elegans is its relatively simple and invariant anatomy. Because each slice contains only a limited number of cells, we can predict which cells are in the different slices and use this information to identify cell type specific genes. Furthermore, we have developed an algorithm that enables us to search for genes with specific expression patterns. Using known marker genes as a starting point, we have identified genes that are specifically expressed in the germline and sperm, in organ structures such as the pharynx, the intestine, the vulva, the spermatheca and the male reproductive tract, and in neurons of the nerve ring and tail ganglia. The ability to identify cell type specific genes is an important advantage over RNA-seq of isolated cells, which does not distinguish between specific and more generally expressed genes. In summary, we have generated high-resolution anteroposterior gene expression maps of C. elegans, including maps of hermaphrodites, males and germline deficient mutants. In addition to providing an expression pattern resource, serial-seq is a powerful method for studying gene expression pattern changes at the whole-genome level.

Gaylord, Olivia et al. (2021) International Worm Meeting "RNA helicase RHA-1 safeguards thermosensitive sperm fertility by promoting small RNA-mediated mRNA clearance"

Lee, Heng-Chi, Brown, Jordan, Gaylord, Olivia, Wu, Wei-Sheng

[International Worm Meeting, 2021]

Male fertility and sperm development are thermosensitive processes whereby germline exposure to high temperature reduces viable sperm. Small RNAs and Argonaute proteins play critical roles in male fertility, particularly at high temperature conditions. In C. elegans, several small RNA pathways function in spermatogenic germlines and target germline expressed mRNAs. Argonaute CSR-1 targets many spermatogenesis genes and recent evidence highlights a dual role for CSR-1 in mRNA protection and mRNA clearance, the second requiring CSR-1 catalytic activity to slice mRNA targets. Whether and how these distinct functions of CSR-1 promote male fertility remains largely unkown. Intriguingly, a germline enriched helicase RNA helicase A (RHA-1) is also required for male thermosensitive fertility. Here we show that, in rha-1 mutants, CSR-1 small RNAs exhibit a slight decrease in overall abundance. The CSR-1 small RNAs are preferentially depleted from the coding sequence rather than the 3' UTR of target mRNAs, similar to the reported CSR-1 small RNA defects found in CSR-1 slicer mutant. When analyzing the mRNA levels of CSR-1 target genes, rha-1 mutants show increased CSR-1 mRNA targets, including spermatogenesis genes. Taken together, our results suggest RHA-1 facilitates CSR-1 slicing of mRNA transcripts to promote mRNA clearance. Our study further highlights the critical role of small RNA-mediated mRNA clearance in sperm fertility.

Curran, Kevin et al. (2013) International Worm Meeting "Chemosensing a predator: Pristionchus pacificus and C. elegans."

Srinivasan, Jagan, Tong, Ada, Chalasani, Sreekanth, Joens, Matthew, Curran, Kevin, Fitzpatrick, James

[International Worm Meeting, 2013]

A current challenge in neuroscience is to bridge the connections between genes, neurons, neural circuits, and behavior in a single animal model. We are approaching this convergence by exploring the mechanisms governing a social interaction between a predator and its prey. A starving predator, Pristionchus pacificus, an omnivorous nematode, will attack and kill its prey, Caeonrhabditis elegans, when the two species share an agar plate. Certain Pristionchus strains exhibit carnivorous mouth morphology allowing them to slice open C. elegans cuticular exoskeleton and consume the underlying tissue. We have employed a novel imaging technique, Helium Ion Microscopy (HIM), to image the mouth cavities and teeth of several Pristionchus isolates. The tooth morphology revealed via HIM imaging correlates with observed feeding behaviors of the Pristionchus strains. We also find that, upon exposure to Pristionchus, C. elegans exhibit an increase in avoidance behavior. Moreover, we have identified ASI, ASJ and ASH as the specific chemosensory neurons that drive this avoidance behavior. We next performed mutant analysis coupled with cell specific rescue experiments and concluded that ocr-2, a TRPV channel, is required in ASH, while tax-4, a cGMP channel, is required in ASI and ASJ, in order to properly execute C. elegans predator avoidance behavior. Finally, this behavior is modulated by anti-anxiety drugs, suggesting that predator avoidance may serve as a model for complex human behaviors, such as fear and anxiety.

Jonathan M Rhine et al. (2003) International Worm Meeting "Cloning pat-12, an atypical Pat gene that may be necessary for proper muscle attachment"

Benjamin D Williams, A Craig Mackinnon, Jonathan M Rhine

[International Worm Meeting, 2003]

Accurate and effective muscle function requires the transmission of forces via specific attachment complexes. In our lab, we study C. elegans mutants in which these attachments in the body wall muscles have been compromised in some way. These mutants together form a class whose mutants display share a common embryonic lethality termed the Pat (Paralyzed and Arrested at Two-Fold) phenotype. One such Pat mutant, pat-12, was mapped to linkage group III between unc-45 and daf-7. Further work suggested that the predicted gene T17H7.4 might correspond to pat-12. The predicted gene T17H7.4 encodes a complex involving as many as 11 splice isoforms and 25 exons and encompasses over 20 kB of genetic space. Previous published data by Gonzy et al. as well as our own data showed that T17H7.4 showed an embryonic lethal RNAi phenotype (100% and 96.2% respectively). Moreover, this lethal phenotype was the same as the previously described PAT phenotype. Rescue experiments using the cosmids containing the genetic region surrounding and containing T17H7.4 were able to rescue the Pat phenotype in pat-12(st430) mutant individuals. In addition, we were able to sequence a mutation in the pat-12(st430) line in the genetic region corresponding to T17H7.4. This G to A mutation leads to the loss of a splice acceptor site shared by 5 of the predicted slice isoforms. Preliminary staining using an antibody raised against PAT-12 stained circumferential rings overlying the body wall muscles.

Herrera, R. Antonio et al. (2009) International Worm Meeting "Laser-Capture Microdissection and Microarray Profiling of C. elegans Male Tail Tip Morphogenesis."

Fitch, David H. A., S, Sindhoora, Herrera, R. Antonio

[International Worm Meeting, 2009]

Regulation of C. elegans male tail tip morphogenesis involves the input of several distinct molecular pathways. The heterochronic [1], Wnt signaling [2], Hox patterning [4] and sex determination [3] pathways each contribute to the proper development of the male tail tip syncytium. Mutations in certain genes result in unretracted adult male tail tips. Our lab and others have confirmed the intersection between these molecular pathways by examining the expression of transgenic reporters in various mutant backgrounds [3]. Our current understanding of the genetic network is limited, as many of these interactions are indirect. To identify candidate genes that are involved in tail tip retraction, we are performing a microarray analysis of gene expression in tail tips isolated from synchronized males and hermaphrodites prior to male L4 tail tip morphogenesis. We are using laser-capture microdissection to obtain tail tips, from which total RNA is isolated for hybridization onto Affymetrix C. elegans GeneChips. The tail tip lends itself particularly well to analysis of post-embryonic gene expression in specific somatic cells because the four tail tip cells can be removed by a single slice perpendicular to the anterior-posterior axis. Work is in progress to identify genes that are differentially expressed and/or change in expression profile from late L3 to middle L4. To confirm their role in morphogenesis, we will examine tail tip phenotypes in RNAi knockdowns or mutants of these genes. [1] Del Rio-Albrechtsen et al. 2006, Dev. Biol. 297:74. [2] Zhao et al. 2002, Development 129:1497. [3] Mason et al. 2008, Development 135:2373. [4] see MD Nelson Abstract.

Curran, Kevin et al. (2011) International Worm Meeting "Dissecting predator/prey interactions in C. elegans."

Chalasani, Sreekanth, Hong, Ray, Curran, Kevin

[International Worm Meeting, 2011]

A current challenge in neuroscience is to bridge the connections between genes, neurons, neural circuits, and behavior in a single animal model. We are exploring the mechanisms governing a social interaction between a predator and its prey. The predator, Pristionchus pacificus, is omnivorous in the wild yet will attack its prey, Caenorhabditis elegans, when the two species share an agar plate. Certain Pristionchus strains exhibit a carnivorous mouth morphology allowing them to slice open C. elegans cuticular exoskeleton and consume the underlying tissue. In turn, upon sensing this predatorial threat, C. elegans exhibits avoidance behavior, in the form of omega bends and reversals, so as to evade capture and consumption. We have developed assays to assess the degree of C. elegans avoidance behavior in response to a lethal predator. Initially, we qualitatively tested 8 Pristionchus strains for their propensity to attack C. elegans on a shared agar plate and observed a range of aggression. Pristionchus (Bolivia) proved to be the most aggressive Pristionchus strain while Pristionchus (Pasadena) demonstrated the least aggressive activity. Correspondingly, Pristionchus (Bolivia) also provoked a robust C. elegans avoidance response with our quantitative avoidance test, the single drop assay. Conditioned media pulled from a pellet of underfed Pristionchus (Bolivia) elicits robust avoidance, while supernatant from underfed Pristionchus (Pasadena) causes only moderate avoidance. Interestingly, C. elegans uses many of the classic neurotransmitter pathways found in vertebrate models (for ex. acetycholine, serotonin, dopamine) to execute cellular communication regarding behavior. Preliminary results reveal that C. elegans pre-treated with Fluoxetine, commonly known as Prozac, demonstrate a reduction in predator avoidance behavior, suggesting involvement of the serotonin pathway in predator avoidance behavior.

Santella, Anthony et al. (2013) International Worm Meeting "Towards the Complete Embryonic Cell Lineage."

Bao, Zhirong, Santella, Anthony, Yu, Zidong, Shroff, Hari, Wu, Yicong, Du, Zhuo

[International Worm Meeting, 2013]

The invariant lineage has been a cornerstone of C. elegans biology. John Sulston's initial lineage 30 years ago used multiple embryos to assemble the invariant pattern. Complete continuous lineaging has never been performed for a single animal. While image analysis software has facilitated lineage creation during early embryogenesis, embryo movement at later stages has hampered the analysis of later development. We have made progress towards lineage tracing during this developmental period through a combination of innovative computational and imaging methods. Our efforts on image analysis are focused on the challenge of reliably following small and crowded nuclei over long periods of time in under sampled images. Our nuclear detection method uses per slice segmentation and a learned shape model to robustly detect and segment nuclei in crowded configurations. The detection results are merged into a cell lineage using multiple linking steps, which select from a set of possible causes and actions, including cell movements, divisions, or detection errors. This is based on probabilistic models of nuclear appearance and local spatial configuration. These computational improvements are bolstered by a qualitative improvement in image quality through the dual-view inverted Selective Plane Illumination Microscope (diSPIM), which captures isotropically sampled volumes at speeds that largely eliminate motion artifacts even through the final stage of embryogenesis. We anticipate that our combined effort will allow not just lineage tracing, but that the detailed dynamics of development (including cell positions and expression patterns) can be followed at high spatiotemporal resolution to build a quantitative model of development. A long term application is the production of WormGUIDES, a 4D atlas tracking both cell positions and neuronal outgrowth.