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.

Hall, Sarah E. et al. (2011) International Worm Meeting "A memory of developmental experience regulates small RNA populations."

Hall, Sarah E., Chirn, Gung-wei, Lau, Nelson C., Sengupta, Piali

[International Worm Meeting, 2011]

Early environmental and developmental experiences have a significant impact on adult phenotypes. We have established C. elegans as a model system to investigate the mechanisms by which a memory of early developmental experiences modulates adult behavior and physiology. We previously showed that wildtype adult animals that had transiently passed through the dauer stage (postdauers) exhibit different gene expression patterns, genome-wide chromatin structure, and behaviors when compared to adults that bypassed the dauer stage (controls). To determine whether small RNA (sRNA) populations are also differentially affected upon passage through the dauer stage, we deep-sequenced sRNA libraries cloned from growth-synchronized control and postdauer adults. To investigate the developmental progression of sRNA populations, we also sequenced sRNA libraries cloned from the L3 larval stage as well as animals that spent one day in the dauer stage. We found that while control and postdauer libraries contain comparable overall proportions of multiple sRNA classes, the abundance of individual sRNAs within a class could be markedly different. For instance, some 21U-RNAs and miRNAs exhibited 2 to 30-fold differences in abundance in postdauer animals compared to control adults. Unexpectedly, we observed a depletion of miRNAs in our dauer library as compared to the L3 larval stage, such that miRNAs comprise only 30% of total sRNAs in dauers as compared to 74% in L3 animals. Preliminary results suggest this result is reflective of a reduction in the number of miRNAs present in dauers and not an increase in endo-siRNA production. To determine whether endo-siRNAs play roles in the regulation of gene expression, we first mapped them with respect to predicted coding regions. Remarkably, we found that more than 50% of genes whose expression is significantly altered in both dauer and postdauers as compared to control animals, exhibit a >2-fold difference in siRNA levels mapping to their 5' UTR or coding sequences in postdauer animals. These observations suggest that gene expression changes as a consequence of early developmental history may be regulated via endo-siRNA-mediated alterations of chromatin, or via post-transcriptional regulation of mRNA levels. Together, these results indicate that developmental experience can have a profound long-term effect on sRNA populations in adults, and that sRNA-mediated pathways may play critical mechanistic roles in the regulation of gene expression and contribute to phenotypic diversity as a consequence of early experience.

Hall, Sarah (2013) Worm Breeder's Gazette "New Lab Announcement: Sarah E. Hall"

Hall, Sarah

[Worm Breeder's Gazette, 2013]

Sarah E. Hall et al. (2007) International Worm Meeting "A cellular ‘memory of developmental history in C. elegans."

Kyuhyung Kim, Piali Sengupta, Sarah E. Hall

[International Worm Meeting, 2007]

Experiences during a critical period of early developmental stages have been shown to shape adult behaviors in many species. In C. elegans, exposure to high levels of pheromone during the L1/L2 larval stage may trigger entry into the dauer developmental stage. Animals exit from the dauer stage and resume reproductive growth when conditions become more favorable. Although many tissue types are remodeled, and gene expression patterns are altered in the dauer stage, post-dauer animals appear grossly similar to animals that bypassed the dauer stage. Thus, whether post-dauer animals retain a cellular memory of their developmental history remains unclear. A few previous reports have suggested that cell lineages, morphology and gene expression patterns may be altered in post-dauer animals as compared to animals that bypassed the dauer stage, but this issue has not been studied systematically [Sutherlin, M.E. and S.W. Emmons. 1994. Genetics 138: 675-688; Euling, S. and V. Ambros. 1996. Development 122: 2507-2515]. We have found that expression of an osm-9::gfp fusion gene in adult animals is regulated by developmental history. osm-9 encodes a TRPV channel, and is expressed in multiple cell types, including the ADL and AWA chemosensory neurons in animals that bypassed the dauer stage. Interestingly, osm-9::gfp expression is eliminated in the ADL, but not the AWA neurons in post-dauer animals. This alteration in gene expression does not appear to be regulated merely by starvation, L1 arrest, or by exposure to pheromone in early larval stages, suggesting that passage through the dauer stage is necessary for the observed changes in osm-9::gfp expression. The change in gene expression is observed in post-dauer animals that spent as little as 1 day in the dauer stage. We have reduced the required cis-regulatory sequences required for conferring this regulation to 500 bp and are further refining this region. We are also examining osm-9::gfp expression in animals mutant for daf and chromatin remodeling genes. Furthermore, since OSM-9 is required for sensory transduction, we are testing for differences in ADL-mediated and osm-9-dependent behaviors of animals that have passed through the dauer stage as compared to control animals. To identify additional genes that may be regulated in a similar manner, we are also expression profiling post-dauer adult and control animals. We expect that this work will allow us to understand how early environmental and developmental experiences affect adult phenotypes by identifying the required molecules and pathways for developmental ‘memory.

Ow, Maria C. et al. (2020) MicroPubl Biol

Ow, Maria C., Hall, Sarah E.

[MicroPubl Biol, 2020]

During its first larval stage (L1), the nematode C. elegans makes a critical decision regarding its developmental trajectory based on environmental conditions. Poor conditions such food scarcity, crowding, or high temperatures promote the entry into a stress-resistant, non-aging, diapause stage named dauer where they can survive for months. Upon improved conditions, dauer larvae resume development as postdauer (PD) L4 larvae and continue to reproductive adulthood as PD adults (Cassada and Russell, 1975). If conditions are favorable, L1 larvae proceed through additional larval stages (L2-L4) until reaching reproductive adulthood (control adults or CON) (Sulston and Horvitz, 1977). We have previously shown that PD adults retain a cellular memory of their early life experience that results in stress-specific transcriptome changes that promote alterations in their life history traits (Hall et al. 2010, 2013; Ow et al. 2018). For instance, wild-type CON adults and PD adults that experienced early-life starvation exhibit reproductive differences that are manifested as decreased brood size in PD animals relative to CON adults. Reproduction in self-fertilizing hermaphrodites is sperm-limited; thus, brood size differences can result from varying numbers of sperm produced by individual animals. We showed previously that postdauers animals that experienced early-life starvation exhibit a significant delay in germline proliferation during the period when sperm are produced compared to control animals of comparable developmental stage. This observation suggests that the decrease in postdauers adult brood size is consistent with a reduction in sperm number, which we have referred to as reproductive plasticity (Ow et al. 2018). The standard wild-type N2 strain was first cultivated in the laboratory over five decades ago, resulting in the fixation of random mutations over thousands of generations in laboratory conditions atypical to what are experienced by natural populations (Sterken et al. 2015). We wondered whether the reproductive plasticity observed in the canonical laboratory wild-type N2 strain is an adaptive trait acquired over time as a result of laboratory conditions, or represents a conserved mechanistic response to starvation stress. We assessed CON and PD brood sizes of six natural isolates representing various branches of the C. elegans phylogenetic tree (Andersen et al. 2012). CON L4 larvae were obtained from a continuously growing mixed population of worms cultured on NGM plates at 20oC. To obtain PD L4 larvae, well-fed worms were first starved on NGM plates at 20oC for about one week until dauers were visible, then worms were collected and incubated for 30 minutes in 1% SDS (sodium dodecyl sulfate) at room temperature with gentle rotation. Because dauers have suspended pharyngeal pumping (Cassada and Russell, 1975), they survive the 1% SDS treatment that would otherwise be lethal if the detergent were ingested. Dauers were then washed with M9 buffer to rinse away the SDS and placed onto seeded NGM plates at 20oC to promote dauer exit. Once the dauers had developed into postdauer L4s, brood size assays were performed in parallel with their CON counterparts by randomly singling out L4 larvae onto seeded 35 mm NGM plates at 20oC, transferring daily them to fresh plates until egg laying ceased, and counting the number of surviving progeny. These surviving progenies were counted as young adults (four days after the mothers were transferred to fresh NGM plates). We found that certain wild isolates, (e.g. AF16 in Figure 1), had the proclivity to crawl to the side of an NGM plate assay plate and desiccate. We thus censored the data for any animal that died before the end of the egg-laying period. We found that four out of six natural isolates (AB1, CB4856, ED3040, and TR403) displayed a significant reduction in brood size in PD adults relative to CON adults similar to N2. The strains, JU440 and KR314, did not exhibit reproductive plasticity and had a similar number of progeny between CON and PD adults (Figure 1). Interestingly, when we measured the brood size in CON and PD adults of another nematode species closely related to C. elegans, C. briggsae AF16, it displayed a modest but statistically insignificant decrease in PD brood size compared to CON adults (Figure 1). Is the reproductive plasticity between CON and PD a reflection of different mating strategies among Caenorhabditis strains? The spontaneous male frequency in the N2 strain is a low ~0.1%, and reproduction is usually achieved through hermaphrodite self-fertilization (Chasnov and Chow, 2002). However, we found that one natural isolate, CB4856, which harbors a higher frequency of males in their population than the laboratory N2 strain (Wegewitz et al. 2008), also exhibits reproductive plasticity between CON and PD. This observation suggests that reproductive plasticity between CON and PD adults may not be simply due to changes in male frequency and levels of mating. The distinction between strains exhibiting adult reproductive plasticity and those that do not is not immediately obvious when comparing phylogenetic proximity or strain isolation date (Andersen et al. 2012). Thus, additional experiments would be necessary to determine the genetic loci in N2 Bristol and the wild isolates that modulate this reproductive trait. Taken together, these results suggest that reproductive plasticity observed in C. elegans is a naturally occurring developmental trait rather than an adaptive trait stemming from decades of cultivation in a laboratory.

BC Prasad et al. (1999) International C. elegans Meeting "The unc-3 locus encodes an O/E transcription factor, CeO/E, which is required for axonal guidance and chemosensory function."

BC Prasad, RR Reed

[International C. elegans Meeting, 1999]

The O/E family of rHLH transcription factors has been implicated in neurogenesis, axonal pathfinding, muscle formation and B cell maturation. The murine O/E proteins are expressed transiently in the developing CNS and PNS during times of axonogenesis and are expressed in olfactory neurons throughout development where they may regulate components of the olfactory signaling cascade. We have identified a C. elegans O/E homologue (CeO/E) that is encoded by the unc-3 locus. Unc-3 mutants display an uncoordinated phenotype attributed to a severe defasiculation and miswiring of the ventral cord (VNC). Using a GFP fusion to the unc-3 promoter and specific antibodies, we observed that CeO/E is expressed transiently in the excitatory VNC motor neurons and throughout development in a pair of chemosensory neurons (ASI amphid neurons). Several observations suggest that the protein may have distinct roles in the two cell types. Although the axonal and dendritic projections of the ASI appear to be normal when labeled with DiI, unc-3 mutants enter the dauer pathway under inappropriate conditions. Although CeO/E possesses highly conserved domains associated with DNA binding in the mammalian homologue, identification of DNA binding sites for the C. elegans protein has not been achieved. We have shown that CeO/E protein can homodimerize in vitro, although the DNA binding specificity of CeO/E is distinct from the mammalian O/E proteins. Several unc-3 target genes ( daf-7, unc-17/cha-1, unc-4 ) have been identified by other laboratories and each contains a mammalian binding site in the promoter region. Remarkably, we have been unable to demonstrate CeO/E binding at these sites. We are generating chimeras of O/E-1 and CeO/E to understand the DNA binding specificity of the CeO/E protein and utilizing a yeast-2-hybrid screen with both O/E-1 and CeO/E to identify interacting proteins in C. elegans . These studies should reveal the mechanism of CeO/E transcriptional activation and target gene regulation.

Shaulov Y et al. (2018) PLoS Pathog "Escherichia coli mediated resistance of Entamoeba histolytica to oxidative stress is triggered by oxaloacetate."

Shaulov Y, Lamm AT, Ankri S, Trebicz-Geffen M, Weiss-Cerem L, Hisaeda H, Methling K, Lalk M, Nagaraja S, Shimokawa C

[PLoS Pathog, 2018]

Amebiasis, a global intestinal parasitic disease, is due to Entamoeba histolytica. This parasite, which feeds on bacteria in the large intestine of its human host, can trigger a strong inflammatory response upon invasion of the colonic mucosa. Whereas information about the mechanisms which are used by the parasite to cope with oxidative and nitrosative stresses during infection is available, knowledge about the contribution of bacteria to these mechanisms is lacking. In a recent study, we demonstrated that enteropathogenic Escherichia coli O55 protects E. histolytica against oxidative stress. Resin-assisted capture (RAC) of oxidized (OX) proteins coupled to mass spectrometry (OX-RAC) was used to investigate the oxidation status of cysteine residues in proteins present in E. histolytica trophozoites incubated with live or heat-killed E. coli O55 and then exposed to H2O2-mediated oxidative stress. We found that the redox proteome of E. histolytica exposed to heat-killed E. coli O55 is enriched with proteins involved in redox homeostasis, lipid metabolism, small molecule metabolism, carbohydrate derivative metabolism, and organonitrogen compound biosynthesis. In contrast, we found that proteins associated with redox homeostasis were the only OX-proteins that were enriched in E. histolytica trophozoites which were incubated with live E. coli O55. These data indicate that E. coli has a profound impact on the redox proteome of E. histolytica. Unexpectedly, some E. coli proteins were also co-identified with E. histolytica proteins by OX-RAC. We demonstrated that one of these proteins, E. coli malate dehydrogenase (EcMDH) and its product, oxaloacetate, are key elements of E. coli-mediated resistance of E. histolytica to oxidative stress and that oxaloacetate helps the parasite survive in the large intestine. We also provide evidence that the protective effect of oxaloacetate against oxidative stress extends to Caenorhabditis elegans.

Ye H et al. (2008) Neurobiol Learn Mem "Trace administration of vitamin E can retrieve and prevent UV-irradiation- and metal exposure-induced memory deficits in nematode Caenorhabditis elegans."

Ye B, Ye H, Wang D

[Neurobiol Learn Mem, 2008]

Vitamin E (alpha-tocopherol), a lipid-soluble anti-oxidant, prevents the uncontrolled propagation of lipid peroxidation by free radicals. Nevertheless, there is weak or no evidence of a protective effect of previous vitamin E intake on cognitive function in humans. In the present study, we explored the thermosensation model to investigate the possible effects of vitamin E administration on memory behaviors in Caenorhabditis elegans. Administration of 100 and 200mug/mL of vitamin E had no significant effects on the memory for different time intervals, whereas relatively high concentration (400mug/mL) of vitamin E exposure shortened the extinction period of the association paradigm (food at 20 degrees C). Following the UV-irradiation, post-treatment with 200mug/mL of vitamin E not only retrieved the UV-irradiation-induced memory deficits, but also enhanced the memory functions in UV-irradiating animals. Post-treatment with trace vitamin E could also ameliorate the memory deficits in metal (Al or Pb) exposed worms. In addition, pre-treatment with 200mug/mL of vitamin E could effectively prevent the occurrence of memory deficits induced by metal exposure and UV-irradiation. Therefore, the close association may exist between trace dietary vitamin E intake and memory behaviors, and a specific response mechanism may be activated after the administration of vitamin E in stress-exposed animals. Moreover, treatment with 200mug/mL of vitamin E could restore the memory deficits formed in the ncs-1 mutant worms, suggesting that exogenous treatment with trace vitamin E can largely mimic the function of NCS-1 in regulating the memory for thermosensation.

Saiki R et al. (2008) Aging Cell "Altered bacterial metabolism, not coenzyme Q content, is responsible for the lifespan extension in Caenorhabditis elegans fed an Escherichia coli diet lacking coenzyme Q."

Saiki R, Clarke CF, Furukawa S, Larsen PL, Bixler T, Lunceford AL, Lee W, Dang P

[Aging Cell, 2008]

Coenzyme Qn is a fully substituted benzoquinone containing a polyisoprene tail of distinct numbers (n) of isoprene groups. C. elegans fed E. coli devoid of Q(8) have a significant life span extension when compared to C. elegans fed a standard "Q-replete"E. coli diet. Here we examine possible mechanisms for the life span extension caused by the Q-less E. coli diet. A bioassay for Q uptake shows that a water-soluble formulation of Q(10) is effectively taken up by both clk-1 mutant and wild-type nematodes, but does not reverse life span extension mediated by the Q-less E. coli diet, indicating that life span extension is not due to the absence of dietary Q per se. The enhanced longevity mediated by the Q-less E. coli diet cannot be attributed to dietary restriction, different Qn isoforms, reduced pathogenesis or slowed growth of the Q-less E. coli, and in fact requires E. coli viability. Q-less E. coli have defects in respiratory metabolism. C. elegans fed Q-replete E. coli mutants with similarly impaired respiratory metabolism due to defects in complex V also show a pronounced life span extension, although not as dramatic as those fed the respiratory deficient Q-less E. coli diet. The data suggest that feeding respiratory incompetent E. coli, whether Q-less or Q-replete, produces a robust life extension in wild-type C. elegans. We believe the fermentation-based metabolism of the E. coli diet is an important parameter of C. elegans longevity.

Lee JH et al. (2014) Phytomedicine "Coumarins reduce biofilm formation and the virulence of Escherichia coli O157:H7."

Cho HS, Lee JH, Kim YG, Ryu SY, Lee J, Cho MH

[Phytomedicine, 2014]

E. coli O157:H7 is the most common cause of hemorrhagic colitis, and no effective therapy exists for E. coli O157:H7 infection. Biofilm formation is closely related to E. coli O157:H7 infection and constitutes a mechanism of antimicrobial resistance. Hence, the antibiofilm or antivirulence approach provides an alternative to antibiotic strategies. Coumarin and its derivatives have a broad range of biological effects, and in this study, the antibiofilm activities of nine coumarins were investigated against E. coli O157:H7. Coumarin or umbelliferone at 50g/ml was found to inhibit biofilm E. coli O157:H7 formation by more than 80% without affecting bacterial growth. Transcriptional analysis showed that coumarins repressed curli genes and motility genes in E. coli O157:H7, and these findings were in-line with observed reductions in fimbriae production, swarming motility, and biofilm formation. In addition, esculetin repressed Shiga-like toxin gene stx2 in E. coli O157:H7 and attenuated its virulence in vivo in the nematode Caenorhabditis elegans. These findings show that coumarins have potential use in antivirulence strategies against persistent E. coli O157:H7 infection.