Filina, O. et al. (2019) International Worm Meeting "Stress discrimination by body-wide, stochastic DAF-16/FoxO nuclear translocation pulses."

Olmedo, M., van Zon, J.S., Filina, O., Sanchez-Romero, M.A.

[International Worm Meeting, 2019]

The transcription factor DAF-16/FoxO is the primary output of the insulin/IGF-1 pathway in C. elegans, which induces stress-response genes upon a broad range of external stresses. How a single pathway can mount a response that is tailored to different types of stress is not understood. Upon stress, DAF-16 translocates from the cytoplasm to the nucleus to induce gene expression. So far, it is assumed that the level of nuclear DAF-16 does not change if stress conditions remain constant. Surprisingly, when we visualized DAF-16 in individual larvae under constant stress, we instead observed stochastic changes in nuclear translocation, with DAF-16 moving between the nucleus and cytoplasm in ~1hr pulses. Moreover, these pulses exhibited striking body-wide synchronization, with cells from different tissues, e.g. the intestine, hypodermis and neurons, switching within 10 minutes of one another, suggesting the presence of a long-range synchronization signal. We used automated image analysis to quantify DAF-16 pulse dynamics in the intestine, under different stresses that caused developmental arrest in L1 larvae. Whereas, for all stresses the fraction of time that DAF-16 was nuclear increased with stress magnitude, different stress types generated clearly distinguishable DAF-16 pulse dynamics: starvation resulted in pulses that resembled stochastic oscillations, while osmotic stress gave rise to random pulses whose duration increased with salt concentration. Finally, for temperature stress we still observed pulses but with a baseline that increased with temperature. Overall, this suggests that changes in the dynamics of DAF-16 nuclear translocation pulses allow cells to determine both magnitude and type of stress. An important question forming the current focus of our research is how DAF-16 pulse dynamics impacts down-stream expression of its target genes and developmental output. Finally, we also observed pulsatile dynamics of the DAF-16 homolog FoxO in mammalian cells at low nutrient levels that activate insulin signaling. Overall, this indicates that DAF-16/FoxO translocation pulses are a general feature of insulin signaling that might be crucial for its ability to differentiate between and mount the correct response to many different types of stress.

Croydon-Sugarman, Melanie et al. (2009) International Worm Meeting "Geographic and genetic variation in fecundity under temperature stress in C. briggsae."

Prasad, Anisha, Croydon-Sugarman, Melanie, Cutter, Asher D.

[International Worm Meeting, 2009]

In C. briggsae, patterns of genetic diversity among strains from across the globe correlate perfectly with the geographic origin of the natural isolates, corresponding to clades of worms from temperate regions, the tropical circles of latitude, and near the equator (Cutter et al. 2006; Dolgin et al. 2007). Ecologically, these geographic regions differ dramatically in temperature regime, begging the question of whether heritable phenotypic differences might also conform to the geographic partitioning of variation in a potentially adaptive manner. An association between the temperature at which a particular isolate is optimally fecund and the temperature of the isolate''s clade of origin could indicate local adaptation and provide insight into C. briggsae ecology and evolution. To address this issue, we tested the thermal tolerance, as quantified by self-fecundity, of 10 wild-isolate strains originating from the three latitudinal regions when the strains were subjected to extreme high and low temperatures. Our results demonstrate a decline to zero progeny production at 32 deg C that was exhibited by worms from all three regions, indicating an upper fertile limit between 30 deg C and 32 deg C for C. briggsae as a species. However, at 30 deg C we observed a significant 4-fold difference in lifetime fecundity for strains from the Tropic circles of latitude clade compared to those of both the temperate and equatorial clades, suggesting a tolerance of the tropical isolates to higher temperatures. Ongoing work explores fecundity at low temperatures (12 deg C - 16 deg C) to test for heritable differences among strains at cooler temperatures. Cutter, A.D., M.A. Felix, A. Barriere & D. Charlesworth. 2006. Patterns of nucleotide polymorphism distinguish temperate and tropical wild isolates of Caenorhabditis briggsae. Genetics. 173: 2021-2031. Dolgin, E.S., M.A. Felix & A.D. Cutter. 2008. Hakuna nematoda: genetic and phenotypic diversity in African isolates of Caenorhabditis elegans and C. briggsae. Heredity. 100: 304-315.

Stroeher VL et al. (1991) International C. elegans Meeting "POSSIBLE LINEAGE-SPECIFIC ACTIVATORS OF THE C. elegans ges-1 GENE."

Stroeher VL, McGhee JD

[International C. elegans Meeting, 1991]

The ges-1 gene of Caenorhabditis elegans encodes a nonspecific carboxylesterase that is expressed exclusively in the intestinal lineage. Deletion and transformation analyses show that this restricted expression is due to lineage specific activators and repressors that bind to upstream promoter sequences (see abstract by E. J. Aamodt, M.A. Chung and J.D.M.). As a first step to clone these regulatory factors, we are identifying their binding sites using gel retardation assays (bandshifts) and DNasel footprinting experiments. At least seven regions of protein interaction have been found in the sequences 935 to 1309 basepairs (bp) upstream of the translation start site. One of these sites, which may bind a putative 'gut activator' molecule is being examined more closely. A doublestranded, 30 bp oligomer (30mer) representing this region has been synthesized. In gel retardation assays with crude nuclear extract from FUdR-blocked embryos, this 30mer gives a single major shifted species, which is effectively competed with unlabelled double-strand 30mer. DNA-protein binding reactions have been exposed to UV irradiation to crosslink the bound protein(s) to the labelled, doublestrand 30mer; a major 80 kd protein is detected. We are now investigating when this protein appears in development.

Wang, Xin et al. (2013) International Worm Meeting "Mutations in C9ORF72/F18A1.6 produce developmental and behavioral defects."

Buttner, Edgar, Wang, Xin

[International Worm Meeting, 2013]

Expanded hexanucleotide repeats in the human C9ORF72 gene cause frontotemporal dementia and amyotrophic lateral sclerosis (Boeve et al., 2012; Renton et al., 2011). The normal and abnormal mechanisms of action of this human disease gene are not fully understood. We are characterizing phenotypes associated with mutations in F18A1.6, the C. elegans homolog of C9ORF72. We found that feeding RNAi of F18A1.6 caused developmental delay, while the insertion/deletion allele ok3062 did not. F18A1.6(RNAi) also reduced locomotor behavior in a thrashing assay, compared to empty-vector control. Surprisingly, ok3062 animals displayed increased thrashing, compared to N2. To better understand the cellular basis of the locomotion phenotypes, we are now conducting pharmacological and neuronal GFP reporter assays. We are also generating transgenic animals to test the ability of C9ORF72 to rescue F18A1.6-associated phenotypes and to test the effects of expressing expanded hexanucleotide repeats in C. elegans. References Boeve, K. B. Boylan, N. R. Graff-Radford, M. DeJesus-Hernandez, D. S. Knopman, O. Pedraza, and P. Vemuri (2012). Characterization of frontotemporal dementia and/or amyotrophic lateral sclerosis associated with the GGGGCC repeat expansion in C9ORF72. Brain 135, 765-783. Renton, E. Majounie, A. Waite, J. Simon-Sanchez, S. Rollinson, J. R. Gibbs, J. C. Schymick, and H. Laaksovirta (2011). A hexanucleotide repeat expansion in C9ORF72 is the cause of chromosome 9p21-linked ALS-FTD. Neuron 72, 257-268.

Honnen, S. et al. (2011) International Worm Meeting "C. elegans vang-1 modulates life span and stress resistance via insulin/IGF-1-like signaling."

Schroder, V., Bossinger, O., Kampkotter, A., Honnen, S., Buchter, C.

[International Worm Meeting, 2011]

The planar cell polarity (PCP) pathway is highly conserved from Drosophila to humans and a PCP-like pathway has recently been described in the nematode Caenorhabditis elegans (1-3). The developmental function of this pathway is to coordinate the orientation of cells or structures within the plane of an epithelium or to organize cell-cell intercalation required for correct morphogenesis (4-5). Here, we describe a novel role of VANG-1, the only C. elegans ortholog of the conserved PCP component Strabismus/Van Gogh. We show that two alleles of vang-1 and depletion of the protein by RNAi cause an increase of mean life span up to 40%. In addition, vang-1 mutants show enhanced resistance to thermal and oxidative stress and decreased lipofuscin accumulation. Life span extension in vang-1 mutants depends on the insulin/IGF-1 like receptor DAF-2 and DAF-16/Foxo transcription factor. This is the first time that a correlation between a key player of the PCP pathway and the modulation of life span and stress resistance has been established. references: 1.Green, J., Inoue, T., and Sternberg, P. (2008). Opposing Wnt pathways orient cell polarity during organogenesis. Cell 134, 646-656. 2.Wu, M., and Herman, M.A. (2006). A novel noncanonical Wnt pathway is involved in the regulation of the asymmetric B cell division in C. elegans. Dev Biol 293, 316-329. 3.Hoffmann, M., Segbert, C., Helbig, G., and Bossinger, O. (2010). Intestinal tube formation in Caenorhabditis elegans requires vang-1 and egl-15 signaling. Dev Biol. 4.Wang, Y., and Nathans, J. (2007). Tissue planar cell polarity in vertebrates: new insights and new questions. Development 134, 647-658. 5.Wu, J., and Mlodzik, M. (2009). A quest for the mechanism regulating global planar cell polarity of tissues. Trends Cell Biol 19, 295-305.

Jin, Eugene et al. (2021) International Worm Meeting "Coordination of neuronal activity and transcriptional programs in motor circuit remodeling"

Jin, Yishi, Jin, Eugene

[International Worm Meeting, 2021]

Neuronal plasticity and circuit stability are fundamental properties of brain development and function. Activity-dependent changes to neuronal connectivity often occur within a defined time window, also known as a developmental plasticity window. How neuronal activity contributes to such precise timing of neural circuit rewiring is a central question in neuroscience. Ultrastructural connectomic studies that began nearly 50 years ago revealed that during the first larval stage, the C. elegans locomotor circuit undergoes dramatic synaptic rewiring known as 'DD synapse remodeling' as postembryonic motor neurons are born to establish the mature motor circuit (White et al., 1978). Live imaging studies subsequently showed that presynaptic terminals in DD motor neurons are progressively removed from the ventral side and new synapses are formed at dorsal locations from mid to late L1 stages (Hallam and Jin, 1998). The precise timing of DD synapse remodeling has been shown to depend on several transcriptional programs and can be modulated by neuronal activity. However, it remains unclear which form of neuronal activity affects the time window of this developmental plasticity, and how neuronal activity is molecularly coupled to transcription regulation. To address this, we are using fluorescently tagged reporters for in vivo detection of the key transcription factors, such as LIN-14 and UNC-30. To precisely determine L1 developmental stages, we use P cell nuclear migration and divisions using Nomarski optics (Sulston and Horvitz, 1977). We have also generated a nuclear calcium sensor to measure activity in DD neurons during synapse remodeling. We will present our detailed findings on the changes in nuclear calcium dynamics before, during and after DD synapse remodeling. References: Hallam, S.J., and Y. Jin. 1998. Nature. 395(6697):78-82. Sulston, J.E., and H.R. Horvitz. 1977. Dev. Biol. 56:110-156. White, J.G., D.G. Albertson, and M.A. Anness. 1978. Nature. 271:764-766.

Song, Myung Chul et al. (2015) International Worm Meeting "C09F5.1, the BRICHOS containing protein functions in stress resistance."

Han, Ching Tack, Song, Kyung Hee, Song, Myung Chul, Kim, Sung Hee

[International Worm Meeting, 2015]

C09F5.1, one of several genes expressed differentially in heat sensitive C. elegans mutants was isolated. To confirm whether C09F5.1 is involved in heat stress response, the expression pattern of C09F5.1 in heat stress condition was analyzed, showing the induction of expression of mRNA and protein by 33degC heat treatment in wild type N2. The induction of C09F5.1 protein by heat treatment delayed in heat shock transcription factor-1(HSF-1) mutants (sy441) compared to N2 implying that the HSF regulates the transcription of C09F5.1 as HSPs. C09F5.1 RNAi-treated worms became more sensitive to heat stress than control worms. These results indicate that C09F5.1 transcribed by HSF-1 may function in heat stress response. To find the putative functional domain of C09F5.1, the computational analysis of C09F5.1 protein was done. The results showed that C09F5.1 consists of an N-terminal region, a type II transmembrane domain and a BRICHOS domain. The various BRICHOS proteins are known related with familial dementia (BRI2), chondrosarcoma (Chondromodulin-1) and interstitial lung disease (SP-C). BRICHOS domain containing proteins function as molecular chaperone(Sanchez-Pulido, Devos et al. 2002). Furthermore recent studies revealed that BRICHOS domain directly interacts with Abeta42 peptide and delays amyloid-beta fibril formation(Willander, Presto et al. 2012, Hermansson, Schultz et al. 2014). When the C09F5.1 was expressed in animal cells (293T), the C09F5.1 protein directly interacted with Abeta42. In order to identify the chaperone activity of C09F5.1 on abnormal protein aggregation, we generated transgenic worms expressing each of the C09F5.1 protein with full length, BRICHOS deletion and N-terminus deletion in Abeta42 expressing worm. The transgenic worms expressing the BRICHOS domain deleted mutant showed the increased survival rate compared to worms expressing Abeta42 solely. These results suggest C09F5.1 BRICHOS containing protein has chaperonin functions under stress but not through the BRICHOS domain only. Currently we are studying the mechanism of protection from amyloid toxicity by the N-terminus of C09F5.1.

Gal, C et al. (2015) International Worm Meeting "Regulation of gene expression by the DREAM complex."

Latorre, I, Appert, A, Chesney, M, Ahringer, J, Gal, C

[International Worm Meeting, 2015]

The conserved eight subunit DREAM (DP, Retinoblastoma [Rb]-like, E2F, and MuvB) complex is a regulator of cell cycle and quiescence genes and plays an important role in development and disease. The C. elegans DREAM members are LIN-35/Rb-like, EFL-1/E2F, DPL-1/DP1, LIN-8, LIN-37, LIN-52, LIN-53, and LIN-54. Recent work has demonstrated that C. elegans direct DREAM targets are de-repressed in the absence of the Rb-like protein LIN-35 and that high gene body levels of the histone variant H2A.Z are linked with target repression1. However, the mechanism that generates gene body H2A.Z enrichment, and the relationship between DREAM and H2A.Z are not currently understood. Furthermore, the functions and regulatory interactions of individual complex members are not clear. For example, LIN-35 has been demonstrated to be a transcriptional repressor by a number of groups, whilst the DP and E2F proteins have previously been reported to act as activators of transcription2. In addition, mutants of individual DREAM complex members do not all have the same phenotype, indicating different functions. Therefore, we are investigating the contribution of DPL-1 (DP) and EFL-1 (E2F) to DREAM target gene repression. We are also carrying out a GFP reporter screen to identify additional components required for DREAM-mediated repression, to elucidate the mechanisms behind DREAM mediated developmental control.1. Latorre, I., Chesney, M.A., Garrigues, J.M., Stempor, P., Appert, A., Francesconi, M., Strome, S., and Ahringer, J. (2015) 'The DREAM complex promotes gene body H2A.Z for target repression', Genes Dev., 29, 495-5002. Chi, W & Reinke, V. (2006) 'Promotion of oogenesis and embryogenesis in the C. elegans gonad by EFL-1/DPL-1 (E2F) does not require LIN-35 (pRB)', Development, 133, 3147-57.

Honnen, Sebastian et al. (2010) C. elegans: Development and Gene Expression, EMBL, Heidelberg, Germany "vang-1 modulates life span and stress resistance via insulin/IGF-1-like signaling"

Bossinger, Olaf, Kampkotter, Andreas, Schroder, Verena, Honnen, Sebastian, Buchter, Christian

[C. elegans: Development and Gene Expression, EMBL, Heidelberg, Germany, 2010]

The planar cell polarity (PCP) pathway is highly conserved from Drosophila to humans and a PCP-like pathway has recently been described in C. elegans [1-3]. The developmental function of this pathway is to mediate the coordinated orientation of cells or structures within the plane of an epithelium or to regulate the organization of cell-cell intercalation that is required for correct morphogenesis [4, 5]. Here, we describe a novel role of VANG-1, the only ortholog of Strabismus/Van Gogh in C. elegans. We show that two alleles of vang-1 and depletion of the protein by RNAi cause an increase of mean life span up to 40%. In addition, vang-1 shows enhanced resistance to thermal-, oxidative-stress and decreased lipofuscin accumulation. Life span extension in vang-1 depends on the insulin/IGF-1 like receptor DAF-2 and DAF-16/Foxo transcription factor. In addition to the modulation of life span, we also observed an extension of reproductive span but a decreased number of progeny, suggesting that VANG-1 links these crucial processes during nematode development. 1.Green, J., Inoue, T., and Sternberg, P. (2008). Opposing Wnt pathways orient cell polarity during organogenesis. Cell 134, 646-656. 2.Wu, M., and Herman, M.A. (2006). A novel noncanonical Wnt pathway is involved in the regulation of the asymmetric B cell division in C. elegans. Dev Biol 293, 316-329. 3.Hoffmann, M., Segbert, C., Helbig, G., and Bossinger, O. (2009). Intestinal tube formation in Caenorhabditis elegans requires vang-1 and egl-15 signaling. Dev Biol. 4.Wang, Y., and Nathans, J. (2007). Tissue/planar cell polarity in vertebrates: new insights and new questions. Development 134, 647-658. 5.Wu, J., and Mlodzik, M. (2009). A quest for the mechanism regulating global planar cell polarity of tissues. Trends Cell Biol 19, 295-305.

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.