Nadine Saul et al. (2008) European Worm Meeting "Analyzing quercetin mediated longevity in C. elegans"

Christian EW Steinberg, Kerstin Pietsch, Ralph Menzel, Nadine Saul

[European Worm Meeting, 2008]

Discovering and understanding longevity pathways are one of the major. intents in Caenorhabditis elegans research. Compound induced longevity as. well as life prolonging phenomena like calorie restriction or dietary. deprivation are well described but the background mechanisms are still. controversially discussed or even unknown. We focused our investigations on. quercetin, a naturally occurred yellow flavonoid, synthesized in different. plants as a phytoalexin. 200 microM quercetin increased nematode''s mean. lifespan up to 26%, apparently without any obvious, negative side effect.. The aim of this study is to determine possible influences on different. fitness parameters and to uncover the pathways and genes, which are. involved in this life extending process. Indeed the resistance for thermal. stress is enhanced, whereas growth and reproduction are not influenced. To. determine the mode of action, different C. elegans aging mutants were. incubated with quercetin. The most surprising effect is the significant. lifespan extension of the short-lived daf-16(mgDf50) null mutant during. quercetin exposure. DAF-16, a forkhead transcription factor, is considered. as key mediator achieving longevity in C. elegans due to the induction of. e.g. antioxidant and chaperone genes. The observed life extension reveals. that DAF-16 is not part of quercetin mediated longevity. Furthermore,. sirtuin SIR-2.1, a putative histone deacetylase and recently discussed as. elicitor for calorie restriction, is probably not involved in quercetin. induced longevity, because sir-2.1(ok434) mutants show a slight lifespan. benefit with quercetin only in some experiments. We suggest a supporting. role for this sirtuin, which can be partly replaced by other proteins.. There is however evidence that suggest AGE-1 and UNC-43 to be involved in. quercetin action, because lifespan assays show no or only a slight increase. in lifespan after quercetin exposure. First results of quantitative real-. time RT-PCR assays indicate a quercetin mediated induction of akt-2. AKT-2. encodes a serine/threonine kinase, supposed to act primarily to antagonize. DAF-16. These data also imply a DAF-16 independent longevity pathway,. whereas the reason for induction of akt-2 gene expression remains to. elucidate.

Saul, Josh et al. (2019) International Worm Meeting "Genetic Control of the Maintenance of the AIA Cell Identity."

Hirose, Takashi, Horvitz, H. R., Saul, Josh

[International Worm Meeting, 2019]

As development progresses in multicellular organisms, the developmental potential of an individual cell becomes increasingly restricted until a final differentiated cellular identity is adopted. Research in recent years has indicated that a cell's differentiated identity can be unstable. The roles of factors that maintain the integrity of a cell's differentiated identity are poorly understood. We are characterizing a C. elegans gene, ctbp-1, that might act in the process of the maintenance of cell identity. ctbp-1 encodes the worm homolog of the C-terminal Binding Protein (CtBP) family of proteins, shown in mice and Drosophila to act as transcriptional corepressors during development. We discovered that ctbp-1 adult worms misexpress an M4 neuron-specific reporter in the AIAs. Strikingly, ctbp-1 L1 worms do not show this AIA misexpression. We hypothesize that ctbp-1 worms properly establish AIA identity but fail to maintain AIA identity as the animal ages and, for example, misexpress genes normally specific to other cell types. Consistent with this hypothesis, we have observed that ctbp-1 mutants similarly show age-dependent defects in AIA cellular morphology (they generate an ectopic neurite) and AIA function (they fail in an AIA-dependent chemotactic behavior), again suggesting a failure in the maintenance of AIA cellular identity. To understand how ctbp-1 acts to regulate AIA cellular identity, we performed a genetic screen looking for suppression of the AIA gene misexpression seen in ctbp-1 animals. We have identified two homeodomain transcription factors, unc-39 and ttx-3, each of which when lost results in suppression of the AIA misexpression phenotype of ctbp-1 mutants. We are now examining the nature of the interactions between ctbp-1 and these transcription factors, e.g., by seeking shared transcriptional targets, with the goal of understanding the consequences of their loss on the regulation of gene expression and the maintenance of AIA cellular identity. We hope that an understanding of the mechanisms that maintain cellular identity in C. elegans will offer insights into how the failure of the maintenance of cell identity can lead to human diseases, e.g., neurodegenerative diseases such as Alzheimer's, Parkinson's and ALS, which might in part involve a loss of neuronal identity.

Saul, Josh et al. (2017) International Worm Meeting "Genetic Control of the Maintenance of the AIA Cell Fate."

Hirose, Takashi, Horvitz, Bob, Saul, Josh

[International Worm Meeting, 2017]

Cell fate can be considered to involve two steps: establishment, in which an undifferentiated cell commits to and expresses a final differentiated fate, and maintenance, in which a differentiated cell preserves the expression of its fate while precluding the expression of other fates. Cell-fate establishment has been well studied and is characterized by changes in gene expression, cell morphology and cellular function. Cell-fate maintenance has received comparatively little attention, and while several examples of defects in cell-fate maintenance exist, the mechanisms by which cell fate is maintained are poorly understood. We have identified a gene, ctbp-1, that appears to be involved in maintaining one or more cell fates in C. elegans. ctbp-1 encodes the only worm member of the C-terminal Binding Protein (CtBP) family of proteins, which in mice and Drosophila have been shown to act as transcriptional corepressors that regulate gene expression during embryonic development1-3. ctbp-1 mutant adult worms show ectopic expression of an M4 neuron-specific reporter in the two embryonically-born AIA neurons, whereas L1 ctbp-1 animals do not display this abnormality. Furthermore, adult but not L1 ctbp-1 animals are defective in an AIA-mediated turning behavior. These observations suggest that AIA gene expression and function are normal in L1 ctbp-1 animals but abnormal in ctbp-1 adults, consistent with the hypothesis that ctbp-1 mutants properly establish the AIA cell fate but fail to maintain this fate. We are further testing this hypothesis and investigating the mechanism by which ctbp-1 might regulate cell-fate maintenance. We performed an EMS mutagenesis screen for suppressors of the M4-specific reporter misexpression phenotype of ctbp-1 mutants to identify genetic interacting partners of ctbp-1 and have identified 21 independent isolates that we are currently mapping. We are also attempting to determine if any cells besides the AIAs are defective in cell-fate maintenance in ctbp-1 mutants. We hope to better understand the genetic control of cell-fate maintenance by determining how ctbp-1 promotes the maintenance of the AIA cell fate. This work might provide insight into diseases, such as cancer, in which perturbation of the maintenance of tumor cell fate might intervene with the oncogenic process. 1. Nibu, Y., Zhang, H. & Levine, M. Interaction of short-range repressors with Drosophila CtBP in the embryo. Science 280, 101-104 (1998). 2. Fang, M. et al. C-terminal-binding protein directly activates and represses Wnt transcriptional targets in Drosophila. EMBO J. 25, 2735-2745 (2006). 3. Hildebrand, J. D. & Soriano, P. Overlapping and unique roles for C-Terminal Binding Protein 1 (CtBP1) and CtBP2 during mouse development. Mol. Cell. Biol. 22, 5296-5307 (2002).

Saul Bert et al. (2007) First Australian C. elegans Meeting "An analysis of CtBP function in C. elegans"

Saul Bert, Hannah R. Nicholas

[First Australian C. elegans Meeting, 2007]

The C-terminal Binding Protein (CtBP) is a well characterised transcriptional co-repressor with homologues found in many diverse organisms. In vertebrates, CtBP has been implicated in a variety of important processes such as apoptosis, with CtBP null organisms exhibiting embryonic lethality. To gain a more complete understanding of this co-repressor''s function, we are investigating CtBP''s function in Caenorhabditis elegans, wherein a CtBP reduction of function strain (ok498) is viable.. in situ hybridisation studies have suggested broad expression of CtBP throughout C. elegans development. We intend to corroborate this finding through the creation of polyclonal CtBP antibodies which will be used to define the expression pattern of CtBP in worms, allowing us to infer potential functions for this protein.. We are particularly interested in a possible role for CtBP in the regulation of apoptosis, and have commenced an investigation using Differential Interference Contrast (DIC) microscopy, together with SYTO(r) 12 staining, to observe variations in germ line apoptosis in live worms. We intend to compare levels of both physiological and gamma radiation induced germ line apoptosis, between wild type (N2) and the ok498 mutant strain of C. elegans. In parallel to this study, we will undertake a non-complementation screen, with the final aim of generating novel CtBP mutants allowing for further functional analysis..

Saul N et al. (2013) Chemosphere "Hormesis and longevity with tannins: free of charge or cost-intensive?"

Steinberg CE, Sturzenbaum SR, Menzel R, Pietsch K, Saul N

[Chemosphere, 2013]

Hormetic lifespan extension is, for obvious reasons, beneficial to an individual. But is this effect really cost-neutral? To answer this question, four tannic polyphenols were tested on the nematode Caenorhabditis elegans. All were able to extend the lifespan, but only some in a hormetic fashion. Additional life trait variables including stress resistance, reproductive behavior, growth, and physical fitness were observed during the exposure to the most life extending concentrations. These traits represent the quality of life and the population fitness, being the most important parameters of a hormetic treatment besides lifespan. Indeed, it emerged that each life-extension is accompanied by a constraining effect in at least one other endpoint, for example growth, mobility, stress resistance, or reproduction. Thus, in this context, longevity could not be considered to be attained for free and therefore it is likely that other hormetic benefits may also incur cost-intensive and unpredictable side-effects.

Kato, Saul et al. (2011) International Worm Meeting "Temporal processing of olfactory information in C. elegans chemosensory neurons."

Cho, Christine, Bargmann, Cornelia, Abbott, L. F., Xu, Yifan, Kato, Saul

[International Worm Meeting, 2011]

Natural behaviors require sensory processing over a wide range of timescales. For example, the nematode Caenorhabditis elegans can track odor concentrations within a second for gradient climbing, escape a noxious stimulus within seconds, and evaluate the quality of its environment over many minutes. All of these behaviors are initiated by specific chemosensory neurons. What mechanisms support chemosensation at different timescales? To address this question we characterized the responses of three C. elegans sensory neurons, AWC, ASH and ASI, to rapidly fluctuating pseudo-random odor sequences. Precise control of input sequences was achieved by switching of laminar liquid streams within a microfluidic chip. All three neuron classes generate highly reliable calcium responses with distinct temporal selectivities. We applied reverse correlation techniques to extract the temporal filtering properties of each neuron. AWC can track odors at a sub-second time scale, consistent with its role in assessing gradient changes during chemotaxis, and the rapid response requires a specific G protein subunit. The ASH neuron acts as a slower integrator of nociceptive signals, including high osmolarity. Surprisingly, ASI also responded to the ASH stimulus, acting as a slow differentiator that subtracts stimulus history over the past few seconds from prior stimulus history. Both AWC and ASH operate in dual regimes with slow (~minute-long) odor responses superimposed on their rapid responses. Our results suggest that C. elegans chemosensory neurons are selective both for chemical identity and for specific temporal signatures.

Saul N et al. (2008) Mech Ageing Dev "Quercetin-mediated longevity in Caenorhabditis elegans: is DAF-16 involved?"

Pietsch K, Saul N, Steinberg CE, Menzel R

[Mech Ageing Dev, 2008]

The polyphenol quercetin has recently been found to extend lifespan and increase stress resistance in the nematode Caenorhabditis elegans. The forkhead transcription factor DAF-16 has previously been linked to these effects. However, by using a daf-16(mgDf50) mutant strain, we show that quercetin exposure leads to increased mean lifespans up to 15%. Furthermore, quercetin-treated daf-16(mgDf50) worms show an enhanced resistance to thermal and oxidative stress. Our data reveal that DAF-16 is not obligatorily required for quercetin-mediated longevity and stress resistance.

Saul N et al. (2011) J Nat Prod "Diversity of polyphenol action in Caenorhabditis elegans: between toxicity and longevity."

Pietsch K, Sturzenbaum SR, Menzel R, Steinberg CE, Saul N

[J Nat Prod, 2011]

The model organism Caenorhabditis elegans was utilized to determine, in vivo, the mode(s) of action of four plant polyphenols, namely, tannic acid (TA), gallic acid (GA), ellagic acid (EA), and catechin (CT). The determination of lifespan, stress resistance, growth, reproduction, eating-related behaviors, antioxidative capacities, and lifespan assays with the mev-1 and the eat-2 mutants as well as in the presence of dead bacteria provided new insights into their action. All four compounds prolonged lifespan, but only TA and CT mediated distinct stress protection. Longevity is unlikely the result of antioxidant capacities but rather due to calorie restriction imitating and hormetic properties in the case of TA and EA or antimicrobial capacities of GA and EA. Furthermore, the prominent "disposable soma theory" is only partly reflected by these polyphenols. In summary, this study underlines the diversity of polyphenolic phytochemicals and their mechanistic background.

Saul N et al. (2022) Biogerontology "Identification of healthspan-promoting genes in Caenorhabditis elegans based on a human GWAS study."

Dhondt I, Temmerman L, Wouters B, Perola M, Saul N, Schmitz-Linneweber C, von Chrzanowski H, Luyten W, Loier T, Kuokkanen M, De Vos WH, Verschuuren C, Zecic A, Braeckman BP

[Biogerontology, 2022]

To find drivers of healthy ageing, a genome-wide association study (GWAS) was performed in healthy and unhealthy older individuals. Healthy individuals were defined as free from cardiovascular disease, stroke, heart failure, major adverse cardiovascular event, diabetes, dementia, cancer, chronic obstructive pulmonary disease (COPD), asthma, rheumatism, Crohn's disease, malabsorption or kidney disease. Six single nucleotide polymorphisms (SNPs) with unknown function associated with ten human genes were identified as candidate healthspan markers. Thirteen homologous or closely related genes were selected in the model organism C. elegans for evaluating healthspan after targeted RNAi-mediated knockdown using pathogen resistance, muscle integrity, chemotaxis index and the activity of known longevity and stress response pathways as healthspan reporters. In addition, lifespan was monitored in the RNAi-treated nematodes. RNAi knockdown of yap-1, wwp-1, paxt-1 and several acdh genes resulted in heterogeneous phenotypes regarding muscle integrity, pathogen resistance, chemotactic behaviour, and lifespan. Based on these observations, we hypothesize that their human homologues WWC2, CDKN2AIP and ACADS may play a role in health maintenance in the elderly.

Kato, Saul et al. (2015) International Worm Meeting "Global brain dynamics generate the holistic motor command pattern in C. elegans."

Kato, Saul, Zimmer, Manuel, Kaplan, Harris, Schrodel, Tina

[International Worm Meeting, 2015]

Due to technical limitations, prior studies in C. elegans have not typically considered distributed coding principles observed in other nervous systems. Instead, identified neurons have been described as dedicated encoders of specific sensory inputs or motor outputs in a context of separable, linear and mostly feed-forward sensory-to-motor pathways. This viewpoint is hard to reconcile with the largely horizontal and recurrent character of the neuronal wiring diagram. Moreover, the circuit elements in these studies overlap, suggesting that processing of various sensory modalities and computation of behavior may be performed by a common system. Here, using a single-cell resolution, brain-wide Ca2+-imaging approach developed by our group, we find that neural population dynamics exhibit a widely shared, low-dimensional component with a cyclical state space trajectory, indicative of a continuous attractor manifold. Next, by calcium imaging in free-moving worms, we find that the activities of key neurons correlate to both high level motor state as well as analog movement parameters such as speed, and that these activities map onto the state space trajectory in a well-ordered manner: trajectory bundles can be mapped to motor command states and decisions between alternate behaviors are readily observable at bundle branch points. Moreover, an analog parameter like speed drive is discernible by the position on the manifold. We argue that this dynamical organization assembles action sequences of discrete motor states and at the same time encodes graded metrics of motor intent. This study establishes, for the first time in any animal, a real-time mapping between neural and behavioral dynamics on a single-trial basis. Using chemical genetics, we find that network dynamics persist when decoupled from output by pre-motor interneuron inhibition. Moreover, manifold topology is robust and provides a framework for a sensory input, which modulates the probability of branch traversal, to produce stimulus-evoked behavioral transitions. Both results indicate that population state structure is stably maintained by intrinsic dynamics. This work shows that many neuron classes participate in a pervasive low-dimensional signal which holds the high-level motor command sequence. In mammals, cortical population dynamics produce functionality in a distributed manner; e.g. in macaque motor cortex, goal parameters are encoded across the neural population and movement is produced by collective dynamics. The character and function of the collective neural dynamics we observe suggests that despite profoundly differing neuroanatomy, the principles of C. elegans and mammalian brain function are far more similar than previously suspected.