Hall, Sarah E. et al. (2009) International Worm Meeting "Environmental programming of gene expression in C. elegans."

Russ, Carsten, Sengupta, Piali, Beverly, Matthew H., Hall, Sarah E., Nusbaum, Chad

[International Worm Meeting, 2009]

Environmental programming describes a phenomenon in which early environmental and developmental experience can affect adult health and behavior through long-lasting changes in gene expression. Increasing evidence suggests that affected genes are ''programmed'' by an environmental stimulus through changes in chromatin states. C. elegans makes an environmentally-regulated developmental decision in early larval stages to continue developing into a reproductive adult, or to enter the alternative dauer stage. When environmental conditions improve, animals exit the dauer stage and continue normal development. Although adults that have passed through or bypassed the dauer stage appear grossly identical, post-dauer adult worms have been reported to retain a cellular memory of their developmental decision. To identify the gene set whose expression may be regulated by a memory of passage through the dauer stage, we compared the transcription profiles of postdauer adult animals and control adult animals that bypassed the dauer stage. We found that the expression of 2,126 genes is predicted to be significantly up or down-regulated in postdauer as compared to control animals. Genes predicted to play roles in reproduction, sensory signal transduction, transcriptional regulation, RNAi, and chromatin remodeling were enriched in the affected gene set. In addition, we quantified altered reproductive and aging phenotypes in postdauer adults compared to controls. To determine whether chromatin remodeling plays a role in maintaining the altered gene expression patterns, we performed chromatin immunoprecipitation followed by Illumina Genome Analyzer sequencing (ChIP-Seq) of genomic regions using antibodies against the common histone modifications H3K4me3, H3K9me3, H3K27me3, and H4panAc. Analysis of the ChIP-Seq data in control and postdauer animals showed a direct correlation of gene expression levels and histone modification enrichments in gene regulatory regions for modifications associated with active transcription. However, we found that H3K9me3 and H3K27me3 modifications were independent of gene expression levels. Furthermore, genes whose expression was altered due to passage through the dauer stage exhibited predictable changes in histone enrichment profiles in postdauer animals. Preliminary evidence suggests that a subset of these gene expression changes are dependent on a chromatin remodeling factor and proteins implicated in RNAi, suggesting a RNAi-dependent transcriptional gene silencing mechanism of environmental programming. We expect that this work will allow us to understand how early environmental experiences affect adult phenotypes, and generate phenotypic diversity in an otherwise isogenic population.

Samuel, Buck S. et al. (2011) International Worm Meeting "WORMFOOD GENOMICS: E. COLI DETERMINANTS OF C. ELEGANS' FITNESS."

Carr, Chris, Rowedder, Holli, Plunkett, Guy, Melo, Justine, Durfee, Tim, Nusbaum, Chad, Glasner, Jeremy, Russ, Carsten, Ruvkun, Gary, Sykes, Sean, Samuel, Buck S., Young, Sarah

[International Worm Meeting, 2011]

Like other metazoans, C. elegans fitness (success) within its microbe-rich habitats depends on a tight balance of energy acquisition and expenditure. Thus, it is also highly tuned to microbial cues that allow it to separate potential food or friend from foe. Accordingly, some microbial signals have been postulated to influence fat storage in parallel to endogenous endocrine cues. Several studies also show that the E. coli-adapted N2-Bristol strain is especially sensitive to 'minor' differences in E. coli strains: faster growth rates, increased progeny delivery rates, and less fat retention is seen when worms consume HB101 compared to OP50. Perhaps due to this fitness benefit, worms also exhibit increased satiety and a behavioral preference for HB101. Thus, we have sought to identify the E. coli gene products that modulate C. elegans fitness. To this end, we have sequenced E. coli genomes routinely used in C. elegans cultivation: HB101 (2 isolates), OP50 (2 isolates) and HT115. Despite little variation among strain isolates, 350 and 412 genes are 'unique' to OP50 and HB101, respectively. Many are organized into clusters, and represent a range of gene functions: e.g., carbohydrate utilization (96), cell wall/LPS modification (42), amino acid metabolism (21), regulation (41), the Cascade system (6) and fatty acid metabolism (5). Phenotype microarrays were also used to confirm the metabolic defects. In order to systematically test the impact of these microbial gene products on C. elegans' fitness, we assembled nearly 200 single gene mutants with defined function in a 'neutral' and consistent genetic background (E. coli K12). We then used a number of assays to test a mutant's impact on N2 growth, broods, body size and fat storage. Our analyses indicate that both genes in core metabolism and transport/biosynthesis of conserved mediators of host interaction-autoinducers, biogenic amines, short-chain fatty acids and LPS-influence N2 fitness. Studies of these small molecules as sensory or nutritive cues to C. elegans directly or via regulation of E. coli metabolism are ongoing. However, results so far indicate that the microbial milieu of signals may be just as important of a determinant of C. elegans' fitness as the nutritional potential for supporting growth of a population within a given habitat.

Walker RJ et al. (2000) European Worm Meeting "Inhibitory actions of a fmrf-amide like peptide and octopamine on the Pharyngeal Muscle are mediated through Goa-1"

Walker RJ, Franks CJ, Holden-Dye L, Rogers C, Burke JF

[European Worm Meeting, 2000]

We are interested in the interplay between classical neurotransmitters and neuropeptides in the control of behaviour. To investigate this we have used electrophysiological recordings from C. elegans pharynx and determined the actions of putative neurotransmitters and neuromodulators on muscle activity. To date, we have only found two agents which inhibit the frequency of pharyngeal action potentials without having any significant effect on basal resting membrane potential, namely octopamine (Franks et al., 1998) and a peptide encoded by the gene flp-3 (SAEPFGTMRFamide: FLP-3). Their action may be described as switching the pharynx off. Neither octopamine nor FLP-3 seem to be present in neurones directly impinging on the pharynx (e.g. Alkema & Horvitz, 1999; Kim & Li, 1999) and therefore their actions are likely to be indirect and involve neurotransmitter release from other neurones. We tested this by comparing the actions of octopamine and FLP-3 between wild-type and a mutant with a severe deficit in synaptic transmission, snb-1 . Pharyngeal action potential frequency was measured by intracellular recordings from the terminal bulb. Activity was stimulated at a constant rate by the addition of 500 nM 5-HT to the perfusate and the spike rate then determined upon addition of either octopamine or FLP-3. Both octopamine and FLP-3 caused a more potent inhibition in snb-1 compared to wild-type. This may be explained if octopamine and FLP-3 exert their inhibitory action by inhibiting release of an excitatory transmitter. Therefore, in a mutant with impaired neurotransmission, such as snb-1, this inhibition of release may be achieved at lower concentrations of the modulator. In the motonervous system it has been shown that the inhibition of release of the excitatory transmitter, ACh, by 5-HT is mediated through the G-protein, GOA-1 (Segalat et al., 1995; Nurrish et al., 1999), and this would also appear to be the case in the pharyngeal nervous system, as both FLP-3 and octopamine are less effective in the mutant goa-1. We are now considering whether the switch that both octopamine and FLP-3 control is the cholinergic pharnygeal motoneurone MC. Alkema & Horvitz, 1999 International Worm Meeting Abstract 148 Franks, Chad, Walker & Holden-Dye 1998 European Worm Meeting Abstract t40 Kim & Li, 1999 Soc Neurosci Abs. 69.1 Nonet, Saifee, Zhou, Rand & Wei, 1998 J.Neurosci. Q8, 70. Nurrish, Segalat & Kaplan, 1999 Neuron 24, 231. Segalat, Elkes & Kaplan, 1995 Science 267, 1648. We are grateful to Laurent Segalat for goa-1(n1134). snb-1(md247) was provided by the Caenorhabditis genetics center, which is funded by the NIH National Center for Research Resources.

Dawes HE et al. (1998) West Coast Worm Meeting "sdc-2 TRIGGERS THE HERMAPHRODITE FATE"

Meyer BJ, Davis TL, Matsubara Lapidus D, Dawes HE

[West Coast Worm Meeting, 1998]

sdc-2 is required for hermaphrodite development, initiating both X chromosome dosage compensation and hermaphrodite sexual differentiation. While XO animals are unaffected by sdc-2 mutations, XX animals that lack sdc-2 fail to implement dosage compensation and die as embryos. The few XX animals that escape this lethality are sick, Dpy, and masculinized. We have found that sdc-2 functions as a sex-specific, zygotic switch that triggers hermaphrodite-specific development during early embryogenesis. sdc-2 is normally expressed exclusively in XX animals, and its ectopic expression in XO embryos is sufficient to trigger the hermaphrodite mode of dosage compensation, resulting in extensive XO-specific lethality. When this lethality is suppressed by a dosage compensation mutation, the rescued XO animals are variably feminized, indicating that ectopic sdc-2 expression in XO embryos also promotes the hermaphrodite mode of sexual differentiation. How does sdc-2 activate the dosage compensation process in hermaphrodites? Sequence analysis revealed little about the mode of SDC-2 action: it is a large, novel protein of about 344kD1. Immunostaining experiments showed that SDC-2 is localized specifically to the X chromosomes of XX embryos, along with other members of the dosage compensation machinery. In fact, SDC-2 may directly facilitate the stable recruitment of other dosage compensation proteins to the X chromosome: in sdc-2 mutants, dosage compensation proteins fail to associate with X even though they are stably produced2. Our combined observations indicate that SDC-2 initiates the sex-specific assembly of dosage compensation proteins on the X chromosome. We next addressed the mechanism by which SDC-2 promotes hermaphrodite sexual differentiation. The role of sdc-2 in sexual differentiation is clearly an important one, since it activates hermaphrodite differentiation by repressing the male-specific gene her-13, and its ectopic expression in XO animals leads to feminization. Our observations suggest that SDC-2 may act directly to repress her-1 expression: immunolocalization experiments have shown that endogenous SDC-2 colocalizes with extrachromosomal arrays bearing her-1 sequences (see abstract by Lieb et al.). We are currently investigating the genetic requirements for the association of SDC-2 with her-1. What are the protein interactions that underlie SDC-2's control of dosage compensation and sex determination? SDC-3 and SDC-2 likely act in partnership to effect both processes, since these two proteins are essential for sex determination and dosage compensation, require each other for stability, and act synergistically to effect the dosage compensation process. We are currently performing biochemical experiments to identify proteins that stably interact with SDC-2 in vivo. This approach may shed light on the mechanism by which SDC-2 recruits dosage compensation proteins to the X chromosome, and may identify protein interactions that are specific to SDC-2's role in sex determination. 1D. Berlin & C. Nusbaum, unpublished 2For example, see Lieb et al., Cell 92:265 3Nusbaum and Meyer, Genetics 122:579