Kim, J. et al. (2017) International Worm Meeting "Neural Interactome: Interactive Visualization of a Neuronal System."

Shlizerman, E., Kim, J., Leahy, W.

[International Worm Meeting, 2017]

Both connectivity structure and dynamical biophysical processes determine the functionality of neuronal networks. We therefore develop a real-time framework, called Neural Interactome, to simultaneously visualize and interact with the structure and dynamics of such networks. Neural Interactome is a cross platform framework implemented in Python and is also a Web interface. It combines graph visualization with simulation of neural dynamics, or experimentally recorded multi neural time series, and allows application of stimuli to neurons for examining responses of the network. In addition, Neural Interactome supports structural changes, such as disconnection of neurons from the network (ablation feature), as typically done in experiments. Neural dynamics can be explored on a single neuron level (using a zoom feature), back in time (using a review feature) and recorded (using presets feature). We implement the framework to a model of the nervous system of Caenorhabditis elegans (C. elegans) nematode, and show that it can assist in studying neural response patterns associated with locomotion and other stimuli. In particular, we demonstrate how stimulation and ablation help to identify neurons, which play critical role in dynamics related to experimentally studied touch response circuit, and explore new scenarios that did not undergo extensive experimental studies.

Kim, J. et al. (2017) International Worm Meeting "Evolution of a novel behavior of Auanema freiburgensis, tube-nictation."

Kim, J., Adams, S., Lee, J., Kim, S., Pires-daSilva, A., Tandonnet, S.

[International Worm Meeting, 2017]

Nematodes can be a good model for evolution of novel traits because they have extremely divergent phenotypes. We are interested in the question of how a novel trait emerges and evolves, which is one of the fundamental questions in evolutionary biology. To answer this question, we use a new species of nematodes, A. freiburgensis. This nematode shows nictation behavior, a dispersal behavior, and interestingly, it can use its own cuticle of the previous molt for tail support instead of fungi or any rough surface, which is the case of C. elegans. To understand how changes in the genome affect evolution of this new behavior, we are sequencing its genome and planning to perform forward and reverse genetics.

Kim, J. et al. (2019) International Worm Meeting "Structural and functional basis for the evolution of three condensins in Caenorhabditis elegans."

Morao, A., Ercan, S., Ragipani, B., Kim, J.

[International Worm Meeting, 2019]

Unlike most eukaryotes, which have two condensins for compacting DNA during mitosis/meiosis, Caenorhabditis elegans have evolved a third version of condensin called dosage compensation condensin (condensin DC). Condensin DC shares 4/5 subunits with condensin I, which is largely cytoplasmic and associates with chromosomes after nuclear envelope breakdown to accomplish condensation before segregation. Unlike condensin I, condensin DC remains bound on X chromosomes in hermaphrodite (XX) throughout interphase to repress the expression from each X chromosome by roughly half, thereby equalizing X expression between hermaphrodites (XX) and males (XO). We are interested in how these condensins have evolved to participate in these two distinct cellular processes. Structurally, condensin DC differs from condensin I by a single subunit DPY-27, a variant of SMC4. Despite minor differences, the main functional motifs of SMC proteins, including SMC-4 and DPY-27, are highly conserved throughout evolution. To study how condensins have evolved in C. elegans, we have generated lines with heat-shock inducible and GFP tagged SMC-4 and DPY-27 with mutations in highly conserved motifs and compared our microscopy data with those of other organisms with the same conserved mutations. We have preliminary data suggesting that, despite the strong structural conservation among SMC proteins in various organisms, the functional consequence of mutating the same conserved motif is different in C. elegans. We observed that the mutation in Walk-B motif of the ATPase domain, which slows down the rate of ATP hydrolysis, in SMC-4 and DPY-27, resulted in their complete loss of binding to DNA. In contrast, others have observed that the same mutations in SMC proteins in Xenopus and chicken cells did not disrupt their ability to bind DNA but resulted in their failure to condense DNA. We conclude that the need for a third version of condensin puts a unique evolutionary constraint on the SMC proteins of C. elegans.

Kim, J. et al. (2017) International Worm Meeting "Late endosomes and lysosomes are potential target sites for a diabetes drug metformin."

Lee, H-Y., You, Y-J., Kim, J., Hyun, M., Ahn, J., Lee, I., Min, K-J.

[International Worm Meeting, 2017]

Diabetes is one of the most impactful diseases worldwide. The most commonly prescribed anti-diabetic drug is metformin. Metformin lowers blood glucose level by decreasing hepatic glucose production in the liver and by increasing glucose uptake in the muscle. In the cell, it activates AMP dependent kinase (AMPK) and inhibits mTOR, creating a low-energy state. Recent studies show that metformin induces cell death in certain cancer cell lines by interfering with the metabolism of the cancer cells. Therefore, understanding of the action mechanisms for metformin will provide insights into the pathophysiology of diabetes and other metabolic disorders as well as better ways to treat them. Metformin has many known targets, yet none of them were shown to directly bind to metformin. Given metformin's broad effects on metabolism, it could act on multiple targets. In order to identify new targets, we performed an unbiased screen in C. elegans using a metformin-induced phenotype in L1 longevity. It has been shown that in Drosophila, metformin treatment (100 mM) reduces life span. We discovered that metformin (100 mM) also reduces a certain type of lifespan, L1 longevity. An unbiased screen for a mutant that survives better in the presence of metformin identified a mutation in an endosomal Na+/H+ exchanger (eNHE), NHX-5. The NHEs exchange one extracellular (or extra-organelle) Na+ for one intracellular (or intra-organelle) H+, making electroneutral exchange. The movement of H+ results in pH changes critical for controlling cell volume and intracellular organelle trafficking. Metformin inhibits oxygen consumption in C. elegans in NHX-5 dependent manner, suggesting a conserved action mechanism of metformin in mitochondria in C. elegans. The same NHE homolog exists in flies where it too mediates the effects of metformin. Interestingly, knockout mice that lack NHE6, the mammalian homolog of C. elegans NHX-5, show Angelman-like syndrome in which the lysosome becomes dysfunctional, mimicking lysosomal storage disorders. Consistent with the lysosomal defects shown in NHE6 knockout mice, metformin treatment misregulated autophagy during L1 starvation, suggesting that metformin could act on the endo-lysosomal compartment via endosomal NHEs and regulate the endocytic cycle and autophagy. Considering the newly discovered roles of endo-lysosomes in metabolism, such as in mTOR or AMPK activation, our study could suggest a novel link between metabolic disorders and the endocytic cycle.

Kim, J. et al. (2019) International Worm Meeting "Computational investigation of locomotion through integration of connectomics, neural dynamics and biomechanics in C. elegans."

Santos, J., Kim, J., Shlizerman, E.

[International Worm Meeting, 2019]

The ability to discern how the brain orchestrates behavior relies on the development of successful computational approaches to link and analyze outcomes of multi-pronged investigations of the nervous system. We developed the Neural Interactome framework to provide a computational platform to simulate the connectome and neural dynamics of C. elegans in real time. The framework combines graph visualization with the simulation of neural dynamics, or experimentally recorded multi neural time series, to allow application of stimuli to neurons and to examine network responses. In addition, Neural Interactome supports structural changes, such as disconnection of neurons from the network (ablation). To explore the role of connectomics and neural interactions in forming neural dynamics the Neural Interactome supports integration of additional connectome connections and various models of neural interaction. In addition, we have taken a further step and extended the platform to examine how neural dynamics shape behaviors. In particular, we include layers which translate neural activity to muscle forces and muscle impulses to body postures. The model also implements an inverse integration procedure which modulates neural dynamics according to external forces on the body. We are able to generate locomotion behaviors typical to the nematode (forward and backward) from neural stimuli or body forces. The characteristics of the movements are similar to experimentally identified ones. Furthermore, the neural dynamics associated with distinct movements can be mapped and classified using low dimensional embeddings. Our results show that our model, and thus the connectome along with neural dynamics, encompass rhythmic activity and locomotion behaviors. Utilizing the inverse integration approach we simulate the effect of environmental forces acting on the body through proprioceptive feedback and show that feedback can entrain and sustain movements initiated by neural or mechanical triggers. Taken together, our results show that the structure of the connectome sets specific movement patterns for the organism. These patterns are enabled by neural dynamics guided by the connectome.

J Kim et al. (2004) European Worm Meeting "FURTHER ANALYSIS OF UBIQUITIN DEPENDENT REGULATION OF THE MYOSIN ASSEMBLY CHAPERONE UNC-45"

C Kaehler, J Kim, T Hoppe

[European Worm Meeting, 2004]

Protein degradation by the ubiquitin/proteasome system plays an important role for many cellular processes such as cell cycle progression, induction of inflammatory response or general protein turnover and is severely dependent on stringent regulation for immaculate biological function. Selective ubiquitylation of target substrate proteins requires a cascade of enzymes consisting of an E1 ubiquitin-activating enzyme, an E2 ubiquitin-conjugating enzyme and finally an E3 enzyme which acts as a ubiquitin-protein ligase. In some instances a fourth enzyme, E4, is required for final multiubiquitylation of the target protein. The progression of this cascade often results in terminal protein degradation by the 26S proteasome. Recent studies in our lab have revealed that UFD-2 and CHN-1 form a novel complex with E3/E4 activity in C. elegans. This complex is necessary and sufficient to multiubiquitylate and subsequently downregulate the myosin chaperone UNC-45. It has been shown that CHIP, the human ortholog of CHN-1, and UNC-45 interact with the chaperones Hsp90 and Hsp70, respectively. Thus, it appears that co-chaperone activity of CHN-1 and UNC-45 in concomitance with E4 dependent degradation result in a tightly regulated mechanism for proper myosin assembly. We are looking for further interaction partners of CHN-1 and UNC-45 which will be cloned for interaction studies to investigate how potential chaperone activity may influence ubiquitylation in vitro. In addition, we will perform a genetic screen for suppression of two different unc-45(ts) alleles we have used for our previous studies as well as generate double mutants to explicate this regulation in detail. We are currently investigating if this mechanism of myosin assembly regulation is also conserved in mammals by studying the identified mouse and human UNC-45 homologs. In this regard, we are cloning the mammalian unc-45 isoforms into expression vectors to perform rescue experiments with the different unc-45(ts) mutants and subsequently with chn-1(by155); unc-45(ts) double mutants to elucidate if they also represent functional orthologs.

J Kim et al. (1999) International C. elegans Meeting "Genetic analysis of lj22, lj21 and lj1 genes involved in nicotine adaptation in C. elegans"

J Kim, W Schafer

[International C. elegans Meeting, 1999]

Long-term nicotine treatment causes adaptation, characterized by desensitization, attenuation, and functional inactivation of receptors while levels are increased. We wish to use genetic analysis in C. elegans to study the molecular mechanism behind nicotine adaptation. Our lab has identified two ways to characterize nicotine adaptation in C. elegans : by a muscle phenotype mediated by nAChRs in body muscle, and an egg-laying phenotype mediated by receptors in both the vulval muscles and the VC neurons (see L. Waggoner poster). Acute treatment of wild type worms with nicotine or levamisole causes complete paralysis and muscle contraction, while long-term treatment renders them tolerant, restoring their muscle function and normal body length. Acute treatment with agonist also stimulates egg-laying in wild type worms, while long-term treatment diminishes this response. The lev-1 mutant shows weak resistance to the effects of nicotine. By using the muscle phenotype, our lab has identified mutants that suppress the lev-1 resistance and restore nicotine sensitivity. lj21 , lj22 and lj10 are such lev-1 suppressor mutants. The lev-1 background has been recently crossed out of these mutants, and we see that lj21 and lj22 display hypersensitivity to nicotine and defects in ability to adapt to its muscle phenotypes. We plan to study next the egg-laying phenotype of all three mutants. Of particular interest is lj22 because it has been shown that this mutant displays defects in both the muscle and egg-laying behaviors. Additionally, lj22 has a very interesting movement phenotype: it generates motion, yet is unable to travel over a distance. This phenotype is mostly observed during larval and early-adult stages while eggs are being laid, and is no longer observed in older adults. lj22 is also characterized by a shorter length and dumpier body, and males show defects in mating. This gene has been shown to be linked to the X-chromosome, and further mapping studies will be performed with the intent to clone the gene. Hopefully, this will enable us to further our understanding of nicotine adaptation on the molecular level.

Kendall Wu et al. (2004) West Coast Worm Meeting "Identification of Genetic Biomarkers and Regulators of Aging in C. elegans"

James Lund, Min Jiang, John Wang, Yueyi Liu, Stuart Kim, Kendall Wu

[West Coast Worm Meeting, 2004]

Despite the wealth of knowledge about conditions and mutations that cause worms to live longer, we still have an incomplete understanding of what varies from young to old worms -- especially at the molecular level. A long-term goal of our laboratory is to determine the molecular changes that occur as an organism ages in order to better understand the aging process and its regulation. Using the nematode Caenorhabditis elegans ( C. elegans ) as a genetic model for aging, we have used DNA microarrays to identify a common set of genes regulated throughout several age-related conditions: genes that change in old age (Lund et al. , 2002), genes that change in the exit from the dauer state (Wang and Kim, 2003), and genes that change expression in long-lived mutants of the insulin-like pathway such as the age-1 and daf-16 mutants (unpublished data). Analysis of the upstream regions of these age-regulated genes showed that they are heavily enriched for a GATA DNA consensus sequence suggesting that a GATA transcription factor may be involved in regulating the expression of these genes. This GATA motif may represent a novel regulatory pathway of the aging process that might act either together or separately from the daf-2 /insulin-like pathway. Using GFP reporters of these genes, we have begun to determine how these genes are age-regulated and to identify the key tissues regulating lifespan. We have also used RNAi to abrogate the expression of these genes to determine their role in longevity. The results of these studies will define a set of molecular biomarkers that will allow a more accurate description of the aging process. In addition, these biomarkers will allow identification of new aging mutants perhaps leading to identification of mutants with accelerated aging. Lund, J., Tedesco, P., Duke, K., Wang, J., Kim, S. K., and Johnson, T. E. (2002). Transcriptional profile of aging in C. elegans. Curr Biol 12, 1566-1573. Wang, J., and Kim, S. K. (2003). Global analysis of dauer gene expression in Caenorhabditis elegans. Development 130, 1621-1634.

Philipp Christoph Janiesch et al. (2008) European Worm Meeting "The Ubiquitin-Selective Chaperone CDC-48 Links Myosin Assembly in C. elegans to Human Myopathy"

Hanns Lochmuller, Thorsten Hoppe,, Philipp Christoph Janiesch, Giuseppe Cassata, Roja Barikbin, Johnny Kim, Sabine Krause, Julien Mouysset

[European Worm Meeting, 2008]

Protein degradation in eukaryotes often requires the ubiquitin-selective. chaperone p97 for substrate recruitment and ubiquitin chain assembly.. However, the physiological relevance of p97 and its role in developmental. processes are still unclear. Here, we discovered an unanticipated function. of CDC-48/p97 in myosin assembly and myofibril organization both in C.. elegans and humans. The developmentally regulated assembly of a CDC-48/UFD-. 2/CHN-1 complex links turnover of the myosin-directed chaperone UNC-45 to. functional muscle formation. Our data suggest a similarly conserved pathway. regulating myosin assembly in man. Remarkably, mutations in human p97,. known to cause hereditary inclusion body myopathy, abrogate UNC-45. degradation and result in severely disorganized myofibrils, detrimental. towards sarcomeric function. These results identify a key role of CDC-. 48/p97 in the process of myofiber differentiation and maintenance, which is. abolished during pathological conditions leading to protein aggregation and. inclusion body formation in human skeletal muscle. To our knowledge this is. the first study implicating functional myosin assembly and maintenance in. the pathophysiology of human inclusion body myopathy associated with Paget. disease of bone and frontotemporal dementia (IBMPFD).. The ubiquitin-selective chaperone CDC-48/p97 links myosin assembly to human. myopathy. Janiesch P.C., Kim J., Mouysset J., Barikbin R., Lochmuller H.,. Cassata G., Krause S., and Hoppe T. (2007). Nat. Cell Biol. 9, 379-390.. Multiubiquitylation by E4 enzymes: one size doesn''t fit all.. Hoppe T. (2005). Trends Biochem. Sci. 30, 183-187.

Pedersen J S et al. (2010) Aging, Metabolism, Stress, Pathogenesis, and Small RNAs, Madison, WI "A study of ADan and ABri aggregation and toxicity in C. elegans models"

Morimoto R I, Wang N, Pedersen J S, Eichel K

[Aging, Metabolism, Stress, Pathogenesis, and Small RNAs, Madison, WI, 2010]

Familial Danish- and Familial British Dementias (FDD/FBD) are rare dominant neurodegenerative disorders mapped to mutations near the stop codon of the BRI2 gene that result in the synthesis of C-terminally elongated amyloidogenic peptides ADan and ABri, instead of the wild-type Bri23 peptide1. Neuropathological lesions in these disorders are similar to Alzheimer's disease (AD), including neurofibrillary tangles, parenchymal amyloid and pre-amyloid deposits. These similarities lead to the hypothesis that protein aggregation in general, irrespective of sequence can result in similar neuropathology. However, FDD and FBD are also manifested by early onset of cataract, deafness and/or ataxia not seen in AD. To further investigate the aggregation and toxicity of ADan and ABri, we have generated transgenic C. elegans animals expressing these peptides as YFP fusions. Animals expressing ADan and ABri exhibit massive, very early onset formation of brightly fluorescent YFP aggregates, while the Bri23 peptide fusions remain soluble. Despite this, behavioral studies indicate that the effects of ADan and ABri aggregation in neurons and muscle cells are significantly different from what is seen for Abeta1-42 and polyglutamine. Crosses between the different models reveal that Abeta1-42 forms heterogeneous co-aggregates with ADan, but not with Q40, bringing the attached CFP/YFP pair into FRET distances similar to that observed for homogeneous aggregates. Recent results indicate that wild-type BRI2-protein may inhibit production and aggregation of the Abeta peptide2, suggesting that loss of function as well as disruptive effects of ADan/ABri aggregation could directly or indirectly affect aggregation of Abeta, leading to the observed early onset of AD-like pathology. 1. A. Rostagno, Y. Tomidokoro, T. Lashley et al., Cell Mol Life Sci 62 (16), 1814 (2005); S. H. Kim, J. W. Creemers, S. Chu et al., The Journal of biological chemistry 277 (3), 1872 (2002). 2. S. Matsuda, L. Giliberto, Y. Matsuda et al., J Neurosci 28 (35), 8668 (2008); J. Kim, V. M. Miller, Y. Levites et al., J Neurosci 28 (23), 6030 (2008).