Jiang, Karen et al. (2009) International Worm Meeting "Conserved transcription factor networks link fatty acid and one carbon metabolism."

Rottiers, Veerle, Jiang, Karen, Watts, Jennifer, Hart, Anne, Naar, Anders, Walker, Amy

[International Worm Meeting, 2009]

Human metabolic disorders such as cardiovascular disease, obesity and metabolic syndrome are linked to changes in fatty acid or cholesterol biosynthesis; processes controlled by the sterol regulatory element binding protein (SREBP) family of transcription factors. To examine SREBP function in vivo, and to delineate SREBP transcriptional functions, we have examined SREBP (SBP-1) gene regulatory mechanisms in C. elegans, combined with complementary studies in mammalian cell lines. SREBP activates genes important for cholesterol, fatty acid and phospholipid biosynthetic pathways, in addition to ensuring production of co-factors such as Acetyl-CoA and NADPH. We have identified a novel set of SREBP-responsive genes in C. elegans, the methyl-group producing enzymes of the 1-carbon cycle (1CC). Multiple genes in this pathway, including sams-1(MAT1A) are responsive to SBP-1/SREBP in both C. elegans and mammalian cells. sams-1/MAT1A encodes a s-adenosyl methionine transferase, which provides the methyl donor (SAMe) for most methylation reactions. We hypothesize these genes are important components of SREBP function because methylation is necessary for phospholipid biogenesis, particularly the synthesis of phosphatidylcholine. Interference with the 1CC in C. elegans through RNAi or in sams-1(ok3033) disrupts lipid homeostasis, resulting in the accumulation of fat droplets in the intestine and body cavity. Importantly, this phenotype is highly reminiscent of lipid accumulation in fatty liver disease in humans and similar to the MAT1A KO in mice (Lu et al PNAS, 2001). Furthermore, we have found that phosphatidylcholine levels are low in sams-1(RNAi) animals suggesting that defects in phospholipid metabolism could contribute to the phenotypes of these animals. SInce sbp-1 is necessary for fat storage, it is surprising that a SBP-1 regulated gene causes increases in lipids. We have found that several SBP-1 target genes that also important for lipid metabolism (fatty acid desaturases fat-5, fat-6 and fat-7) are strongly upregulated in sams-1(RNAi) animals. In addition, sbp-1 and the FA desaturases are necessary for lipid accumulation in sams-1(RNAi) animals, suggesting that changes in 1C metabolism could impact key aspects of lipid biosynthesis through SBP-1. Alterations in 1C metabolism and SREBP function are associated with similar diseases, suggesting that co-regulation with lipid homeostasis may be a common impact point in metabolic disorders.

Jiang, Karen et al. (2015) International Worm Meeting "Sensorimotor signal transmission through AIY interneuron in Caenorhabditis elegans during isothermal tracking."

Ling, Linsay, Mendoza, Steve, Sherry, Tim, Nowak, Nathaniel, Arisaka, Katsushi, Jiang, Karen, Haller, Leonard

[International Worm Meeting, 2015]

Perception and navigation through space require accurate translation and transmission of sensory input to motor output. On a linear temperature gradient, Caenorhabditis elegans demonstrate a distinct behavioral phenotype in which they frequently travel along isotherms, maintaining sensitivity within 0.05 degC. This isothermal attractor state is correlated with movement at a constant and maximal velocity. We investigate how AIY, a first layer interneuron and postsynaptic partner to AFD thermosensory neuron, is able to integrate thermal information to return specific well-defined behavioral phenotypes. Prior observations of neural activity in vivo involve partial paralysis or constraint of the worm while stimuli is applied. Other systems circumvent this limitation by re-centering the stage; this generates an external force during stage acceleration introducing another stimulus. We overcome these two primary obstacles through the implementation of a novel automated worm-tracking epi-fluorescent microscope. The three-camera microscope system mounted on a movable XY stage captures dynamic Ca+2 signals in Cameleon-labeled neurons while the nematode navigates unconstrained along the temperature gradient. Implementing this set up, we observed that the greatest temperature difference occurs between the extremes of the head movement while along isotherms which phase lock with fluorescence response in AIY. The steady Ca+2 waveform in AIY suppresses reversals and maintains high speeds to downstream motor circuitry. .

Roussell DL et al. (1994) Parasitol Today "Germ-line determination in Caenorhabditis and Ascaris: will a helicase begin to unravel the mystery?"

Bennett KL, Roussell DL, Gruidl ME

[Parasitol Today, 1994]

How cell lineages are established during development in higher eukaryotes is being addressed by geneticists and by developmental and molecular biologists. In Drosophila melanogaster, a gene corresponding to a germ-line-specific RNA helicase, vasa, has been shown to be a component o f the posteriorly localized germ granules o f the developing embryo. A putative RNA helicase, glh-I r which appears germ-line specific in its expression, has recently been reported from the free-living nematode Caenorhabditis elegans. Parasitologists studying the nematode Ascaris lumbricoides var. suum have found it to be a useful complement to Caenorhabditis. Deborah Roussell, Michael Gruidl and Karen Bennett predict that Ascaris will be valuable in determining the role played by germ-line helicases in development.

Walker, Amy et al. (2010) C. elegans: Development and Gene Expression, EMBL, Heidelberg, Germany "Transcriptional networks link lipid and methyl donor biosynthetic pathways"

Watts, Jennifer, Jiang, Karen, Hart, Anne, Rottiers, Veerle, Norton, David, Walker, Amy, Naar, Anders

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

The highly conserved sterol regulatory element binding protein (SREBP) family of transcription factors regulates cholesterol, fatty acid and phospholipid biosynthesis genes and impact human metabolic disorders. We have examined mechanisms of gene regulation by the SREBP ortholog SBP-1 in C. elegans and identified enzymes in the 1-carbon cycle, a key biochemical circuit for producing methyl groups, as a novel metabolic pathway controlled by SREBP. The SBP-1-regulated gene sams-1 encodes the central enzyme in this metabolic circuit and synthesizes the methyl donor SAMe which is required for most cellular methylation reactions. Interestingly, RNA interference of sams-1 disrupts normal lipid homeostasis, resulting in elevated expression of SBP-1 target genes and fat droplet accumulation throughout the worms. These findings suggest that SBP-1 may respond to changes in some methylation-dependent process when sams-1 activity is decreased. We have also found that levels of the essential membrane phospholipid phosphatidylcholine (PC) are low in sams-1(RNAi) animals, correlating with the importance of methylation for the primary PC biosynthetic pathway. To determine if low PC levels contributed to sams-1 phenotypes, we bypassed methylation-dependent PC synthesis by dietary addition of choline. Choline rescued multiple sams-1 phenotypes, including lipid accumulation and upregulation of SBP-1 target genes. This suggests that, akin to cholesterol inhibition of mammalian SREBPs, SBP-1 may be negatively regulated by PC. Thus, we have discovered a nove l set of SBP-1/SREBP target genes which impact production of methyl donors within the cell. Regulation of the 1-CC by SBP-1/SREBP may allow coordinated production of lipoprotein particles, and opens the possibility that SBP-1/SREBP may affect other methylation-dependent processes as well, such as epigenetic gene regulation.

Mendoza, Steve et al. (2015) International Worm Meeting "A Novel Worm Tracking Calcium Imaging System Utilizing a Mobile Microscope."

Sherry, Tim, Lam, Brian, Kao, Michelle, Nowak, Nate, Mendoza, Steve, Kim, Taejoon, Arisaka, Katsushi, Madruga, Blake, Jiang, Karen

[International Worm Meeting, 2015]

Worm tracking of freely moving worms is essential to study the connection between behavior and neural activity. However, these microscopes are often limited in their ability to track worms under various behavioral simulations. We present a fluorescence worm tracking microscope that has an open geometry and thus can track worms even if the behavioral platform is difficult or impossible to move via a motorized stage. As opposed to other automated worm tracking systems, our microscope is fully mobile-where all the optical components are mounted on top of a motorized xy stage-while the sample stage where C. elegans rests is stationary. This platform allows for ratiometric calcium imaging while also tracking a dark field worm image for behavioral analysis, running at 15 frames per second. The current configuration has three cameras, two for each of the YFP and CFP channels, and a dark field image showing the worm body, under a 10x magnification; the microscope can also be adjusted to image at 20x. Being able to track freely moving worms without moving the sample stage, allows our microscope to perform worm tracking in experimental conditions that similar systems have not been able to achieve. In addition, since the sample is stationary, we also avoid introducing confounding effects on the worms due to stage acceleration. To test this novel hardware, we run a thermotaxis experiment tracking a worm without moving the temperature platform. Our worm was labeled with AIY::CAM and we find a correlation between the AIY activity and the temperature of the head location during isothermal behavior. Our methodology could also apply to other behavioral experiments where an external stimulus would be hard to move via a motorized stage, such as an electrotaxis experiment.

Mondol V et al. (2012) Curr Top Dev Biol "Let's make it happen: the role of let-7 microRNA in development."

Mondol V, Pasquinelli AE

[Curr Top Dev Biol, 2012]

Noncoding RNAs have emerged as an integral part of posttranscriptional gene regulation. Among that class of RNAs are the microRNAs (miRNAs), which posttranscriptionally regulate target mRNAs containing complementary sequences. The broad presence of miRNAs in lower eukaryotes, plants, and mammals highlights their importance throughout evolution. MiRNAs have been shown to regulate many pathways, including development, and disruption of miRNA function can lead to disease (Ivey and Srivastava, 2010; Jiang et al., 2009). Although the first miRNA genes were discovered in the nematode, Caenorhabditis elegans, almost 20 years ago, the field of miRNA research began when they were found in multiple organisms a little over a decade ago (Lagos-Quintana et al., 2001; Lau et al., 2001; Lee and Ambros, 2001; Lee et al., 1993; Pasquinelli et al., 2000; Wightman et al., 1993). Here, we review one of the first characterized miRNAs, let-7, and describe its role in development and the intricacies of its biogenesis and function.

Walker, Amy K. et al. (2009) International Worm Meeting "Conserved SREBP Gene Regulatory Mechanisms Controlling Lipid Homeostasis During Feeding and Fasting."

Naar, Anders, Yang, Fajun, Walker, Amy K., Hart, Anne, Ji, Jun-yuan, Dyson, Nicholas, Rodgers, Joseph, Puigserver, Pere, Watts, Jennifer, Jiang, Karen

[International Worm Meeting, 2009]

Human metabolic disorders such as obesity and metabolic syndrome are linked to changes in fatty acid or cholesterol biosynthesis; processes controlled by the sterol regulatory element binding protein (SREBP) transcription factors. To study SREBP function in vivo, and to delineate SREBP transcriptional functions, we have examined SREBP (SBP-1) gene regulatory mechanisms in C. elegans, combined with complementary mechanistic studies in mammalian cell lines. SREBP activates genes important in cholesterol, fatty acid (FA) and phospholipid biosynthetic (PL) pathways and in production of co-factors such as Ac-CoA and NADPH. We have identified a novel set of SREBP-responsive genes in C. elegans and mammals, the methyl-group producing enzymes of the 1-carbon cycle (1-CC). We hypothesize SREBP regulates these genes because PL biogenesis requires methylation. Interference with the 1-CC in C. elegans disrupts lipid homeostasis, resulting in intestinal lipid accumulation. Importantly, this phenotype resembles lipid accumulation in human fatty liver disease. Co-regulation of the 1CC and lipid homeostasis may be a common impact point in metabolic disease. We have also investigated mechanisms of SREBP inhibition during fasting and found that SIRT1(sir-2.1) limits fat production through direct deacetylation of SREBP, attenuating SREBP-dependent expression of FA biosynthetic genes. Furthermore, we found that C. elegans lacking functional sir-2.1 (SIRT1), or treated with sirtuin inhibitors, retain fats during fasting. SBP-1 targets such as FA desaturase fat-7 are inappropriately expressed during fasting in sir-2.1(lof) animals or after treatment with sirtuin inhibitors. These mechanism are conserved; fasted Drosophila larvae lacking dSir2 (sir-2.1) fail to downregulate dSREBP-dependent genes and retain fat body lipids. SREBP target gene expression increases after RNAi depletion or inhibition of SIRT1 in human cells and after SIRT1 depletion in murine liver during fasting. Thus, SIRT1/SIR-2.1/Sir2 plays an important conserved role in lipid homeostasis by shutting down SREBPs during fasting. These studies highlight the complex relationships between metabolic pathways in humans; delineating common control points will be key for designing treatments.

Dave Markwardt et al. (2001) International C. elegans Meeting "A screen for natural targets of NMD using C. elegans cDNA microarrays"

Philip Anderson, Dave Markwardt

[International C. elegans Meeting, 2001]

mRNA transcripts that contain premature termination codons are degraded in a regulated manner by a specific decay pathway termed mRNA surveillance or nonsense mediated mRNA decay (NMD). mRNA surveillance has been described in all eukaryotes tested including plants, flies, yeast, nematodes, and mammals. Loss-of-function mutations affecting any of seven smg genes eliminates mRNA surveillance in C. elegans . While much progress has been made in defining the cis -acting elements and trans -acting factors necessary for mRNA surveillance, less is know about the mRNAs produced during the normal course of gene expression that serve as substrates of NMD. In order to systematically investigate these natural targets of NMD, I have probed high density microarrays of C. elegans cDNA clones. These arrays, made in Stuart Kim's lab at Stanford University, consist of PCR fragments from 17,871 genes currently annotated in the C. elegans genome (Jiang et al., 2001). We performed nine different hybridizations, each comparing expression profiles of poly(A) + mRNA from smg(-) and smg(+) samples. The microarray screen identified 402 mRNAs whose abundance is 2-fold or greater in smg(-) mutants. Several observations suggest that microarrays are a reliable method for identifying natural targets of mRNA surveillance: 1. Control mRNAs known previously to be natural targets of mRNA surveillance ( rpl-12 and srp-20 ) were consistently elevated on the microarray screen. 2. Both northern blots and quantitative PCR analyses confirm the microarray results. These results indicate that the microarray screen will be useful for the systematic investigation of the C. elegans natural targets of NMD. We are presently analyzing smg(-) affected mRNAs to establish whether they are direct or indirect targets of NMD, and understand why their abundance is increased in smg(-) mutants. We anticipate that analysis of the ~400 candidate genes will identify broad categories of mRNAs produced in wild-type animals yet targeted for degradation by NMD and, possibly, new principles of post-transcriptional regulation of gene expression. Jiang M, Ryu J, Kiraly M, Duke K, Reinke V, Kim SK. Genome-wide analysis of developmental and sex-regulated gene expression profiles in Caenorhabditis elegans . Proc. Nat. Acad. Sci. USA. (2001) 98 (1):218-23.

Mukhopadhyay A et al. (2006) Exp Gerontol "Worming pathways to and from DAF-16/FOXO."

Tissenbaum HA, Oh SW, Mukhopadhyay A

[Exp Gerontol, 2006]

In Caenorhabditis elegans, the insulin/IGF-1 signaling pathway controls many biological processes such as life span, fat storage, dauer diapause, reproduction and stress response . This pathway is comprised of many genes including the insulin/IGF-1 receptor (DAF-2) that signals through a conserved PI 3-kinase/AKT pathway and ultimately down-regulates DAF-16, a forkhead transcription factor (FOXO). DAF-16 also receives input from several other pathways that regulate life span such as the germline and the JNK pathway [Hsin, H., Kenyon, C., 1999. Signals from the reproductive system regulate the lifespan of C. elegans. Nature 399, 362-366; Oh, S.W., Mukhopadhyay, A., Svrzikapa, N., Jiang, F., Davis, R.J., Tissenbaum, H.A., 2005. JNK regulates lifespan in Caenorhabditis elegans by modulating nuclear translocation of forkhead transcription factor/DAF-16. Proc. Natl. Acad. Sci. USA 102, 4494-4499]. Therefore, DAF-16 integrates signals from multiple pathways and regulates its downstream target genes to control diverse processes. Here, we discuss the signals to and from DAF-16, with a focus on life span regulation.

Delattre M et al. (2001) Dev Biol "Development and evolution of a variable left-right asymmetry in nematodes: the handedness of P11/P12 migration."

Felix MA, Delattre M

[Dev Biol, 2001]

In Caenorhabditis elegans, two lateral blast cells called P(11/12)L and P(11/12)R are symmetric left-right homologs at hatching, migrate subsequently in opposite anteroposterior directions during the first larval stage, and adopt two different fates, thus breaking the symmetry between them. Our results show that, unlike most other cell fate decisions in C. elegans, the orientation of P(11/12)L/R migration is highly biased, but not fixed. The handedness of their migration is linked to whole body handedness and is randomized in lin-12/Notch mutants and by ablation of the Y cell. Migration handedness is independent of P11 and P12 fate determination, previously shown to require the LIN-44/Wnt and the LIN-3/EGF pathways (L. I. Jiang and P. W. Sternberg, 1998, Development 125, 2337-2347). We further show that several changes in P(11/12)L/R asymmetry have occurred during nematode evolution: loss of asymmetry or reversals in orientation of migration. Strikingly, for most species studied, handedness of migration is highly biased but not fixed. Thus, whereas the final cell fate pattern of P11/12 is invariant, the developmental route leading to it is subject both to developmental indeterminacy and to evolutionary variations.