Li Z et al. (2015) Worm "How are necrotic cells recognized by their predators?"

Zhou Z, Li Z

[Worm, 2015]

Necrosis is a type of cell death often caused by cell injury and is linked to human diseases including neuron degeneration, stroke, and cancer. Cells undergoing necrosis are engulfed and degraded by engulfing cells, their predators. The mechanisms by which necrotic cells are recognized and removed remain elusive. Here we comment on our recent findings that reveal new molecular mechanisms of necrotic-cell recognition. Through studying the C. elegans touch neurons undergoing excitotoxic necrosis, we identified a receptor/ligand pair that enables engulfing cells to recognize necrotic neurons. The phagocytic receptor CED-1 is activated through interaction with its ligand phosphatidylserine (PS), exposed on the surface of necrotic cells. Furthermore, against the common belief that necrotic cells have ruptured plasma membrane, we found that necrotic C. elegans touch neurons actively present PS on their outer surfaces while maintaining plasma membrane integrity. We further identified 2 mechanisms governing the presentation of PS, one of which is shared with cells undergoing apoptosis, a "cell suicide" event, whereas the other is unique to necrotic neurons. The influx of Ca(2+), a key necrosis-triggering factor, is implicated in activating a neuronal PS-scramblase for PS exposure. We propose that the mechanisms controlling PS-exposure and necrotic-cell recognition by engulfing cells are likely conserved from worms to humans.

Li Z et al. (2016) Neuron "An Elegant Circuit for Balancing Risk and Reward."

Iliff AJ, Xu XZ, Li Z

[Neuron, 2016]

Animals constantly encounter conflicting cues in natural environments. To survive and thrive, they must make appropriate behavioral decisions. In this issue, Ghosh etal. (2016) identified a neural circuit underlying multisensory threat-reward decision making using an elegant C.elegans model.

Li, Z. et al. (2017) International Worm Meeting "Fine tuning of a motor behavior by a dual role interneuron."

Li, Z., Wani, K., Zhou, J., Yu, T., Liu, J., Piggott, BJ., Xu, S.

[International Worm Meeting, 2017]

Animals produce an incredible repertoire of motor actions. How genes and neural circuits regulate motor behaviors is poorly understood. Using C. elegans locomotion behavior as a model, by employing multidisciplinary approaches including optogenetics and in vivo calcium imaging, we found that a single rneuron RIM can both promote and suppress reversals during locomotion. Activation of RIM by optogenetics triggers reversals, and this promotion effect is mediated by the command interneurons AVA/AVE and gap junctions connecting AVA/AVE and RIM. By contrast, blocking chemical transmission in RIM increases reversal frequency, indicating that chemical transmitter release from RIM inhibits reversal initiation. Our findings suggest that motor behaviors are fine-tuned by motor circuits, and a single neuron in the circuitry may have complex roles in the regulation of motor output.

Li Z et al. (2013) Methods Mol Biol "Monitoring the clearance of apoptotic and necrotic cells in the nematode Caenorhabditis elegans."

Li Z, He X, Lu N, Zhou Z

[Methods Mol Biol, 2013]

The nematode Caenorhabditis elegans is an excellent model organism for studying the mechanisms -controlling cell death, including apoptosis, a cell suicide event, and necrosis, pathological cell deaths caused by environmental insults or genetic alterations. C. elegans has also been established as a model for understanding how dying cells are cleared from animal bodies. In particular, the transparent nature of worm bodies and eggshells make C. elegans particularly amenable for live-cell microscopy. Here we describe methods for identifying apoptotic and necrotic cells in living C. elegans embryos, larvae, and adults and for monitoring their clearance during development. We further discuss specific methods to distinguish engulfed from unengulfed apoptotic cells, and methods to monitor cellular and molecular events occurring during phagosome maturation. These methods are based on Differential Interference Contrast (DIC) microscopy or fluorescence microscopy using GFP-based reporters.

Li Z et al. (2022) Curr Biol "Chemosensation: Corollary discharge filters out self-generated chemical cues."

Li Z, Xu SXZ

[Curr Biol, 2022]

Corollary discharge allows organisms to discriminate external sensory inputs from self-generated cues. However, the underlying synaptic and molecular mechanisms are not well understood. A new study has identified a tyraminergic corollary discharge signal that extrasynaptically modulates chemosensory neurons in Caenorhabditis elegans.

Li Z et al. (2019) J Vis Exp "Using Caenorhabditis elegans for Studying Trans- and Multi-Generational Effects of Toxicants."

Ai F, Yu Z, Zhang J, Yin D, Li Z

[J Vis Exp, 2019]

Information about toxicities of chemicals are essential in their application and waste management. For chemicals at low concentrations, the long-term effects are very important in judging their consequences in the environment and on human health. In demonstrating long-term influences, effects of chemicals over generations in recent studies provide new insight. Here, we describe protocols for studying effects of chemicals over multiple generations using free-living nematode Caenorhabditis elegans. Two aspects are presented: (1) trans-generational (TG) and (2) multi-generational effect studies, the latter of which is separated to multi-generational exposure (MGE) and multi-generational residual (MGR) effect studies. The TG effect study is robust with a simple purpose to determine whether chemical exposure to parents can result in any residual consequences on offspring. After the effects are measured on parents, sodium hypochlorite solutions are used to kill the parents and keep the offspring so as to facilitate effect measurement on the offspring. The TG effect study is used to determine whether the offspring are affected when their parent is exposed to the pollutants. The MGE and MGR effect study is systematical used to determine whether continuous generational exposure can result in adaptive responses in offspring over generations. Careful pick-up and transfer are used to distinguish generations to facilitate effect measurement on each generation. We also combined protocols to measure locomotion behavior, reproduction, lifespan, biochemical and gene expression changes. Some example experiments are also presented to illustrate the trans- and multi-generational effect studies.

Li Z et al. (2014) J Biol Chem "Glutathione-dependent one-electron transfer reactions catalyzed by a B12 trafficking protein."

Gherasim C, Banerjee R, Lesniak NA, Li Z

[J Biol Chem, 2014]

CblC is involved in an early step in cytoplasmic cobalamin processing following entry of the cofactor into the cytoplasm. CblC converts the cobalamin cargo arriving from the lysosome to a common cob(II)alamin intermediate, which can be subsequently converted to the biologically active forms. Human CblC exhibits glutathione (GSH)-dependent alkyltransferase activity and flavin-dependent reductive decyanation activity with cyanocobalamin (CNCbl). In this study, we discovered two new GSH-dependent activities associated with the Caenorhabditis elegans CblC for generating cob(II)alamin: decyanation of CNCbl and reduction of aquocobalamin (OH2Cbl). We subsequently found that human CblC also catalyzes GSH-dependent decyanation of CNCbl and reduction of OH2Cbl, albeit efficiently only under anaerobic conditions. The air sensitivity of the human enzyme suggests interception by oxygen during the single-electron transfer step from GSH to CNCbl. These newly discovered GSH-dependent single-electron transfer reactions expand the repertoire of catalytic activities supported by CblC, a versatile B12-processing enzyme.

Li Z et al. (2014) Cell "Encoding of both analog- and digital-like behavioral outputs by one C. elegans interneuron."

Xu XZ, Li Z, Zheng M, Liu J

[Cell, 2014]

Model organisms usually possess a small nervous system but nevertheless execute a large array of complex behaviors, suggesting that some neurons are likely multifunctional and may encode multiple behavioral outputs. Here, we show that the C. elegans interneuron AIY regulates two distinct behavioral outputs: locomotion speed and direction-switch by recruiting two different circuits. The "speed" circuit is excitatory with a wide dynamic range, which is well suited to encode speed, an analog-like output. The "direction-switch" circuit is inhibitory with a narrow dynamic range, which is ideal for encoding direction-switch, a digital-like output. Both circuits employ the neurotransmitter ACh but utilize distinct postsynaptic ACh receptors, whose distinct biophysical properties contribute to the distinct dynamic ranges of the two circuits. This mechanism enables graded C. elegans synapses to encode both analog- and digital-like outputs. Our studies illustrate how an interneuron in a simple organism encodes multiple behavioral outputs at the circuit, synaptic, and molecular levels.

Li, Z. et al. (2019) International Worm Meeting "Visualizing C. elegans transcripts reveals distinct dynamics of membrane-associated genes."

Zhang, P., Zhang, H., Zhang, R., Li, Z.

[International Worm Meeting, 2019]

Localized mRNAs contribute to targeting proteins to discrete sites and are required for diverse cellular and developmental events, including translation, cell division, cell motility and cell-fate determination. The ability to visualize mRNA in real time at high resolution in live animals are critical for full understanding the spatiotemporal dynamics of gene regulation and function. Here we report a method to visualize mRNA using PP7-PCP system in C. elegans, in which mRNAs containing bacteriophage PP7 binding sites are labeled by the PP7 coat protein fused to fluorescent reporters. Using this approach, we construct a collection of tissue-specific, differentially expressed, membrane-associated mRNA toolkit strains and detect distinct mRNA dynamics and localizations through monitoring single RNP particles. We have applied this system to dissect the cis-regulatory element for mRNA localization and explore development- and tissue-dependent mRNA-protein localization correlations. Furthermore, we attempt to use the PP7-PCP RNA labeling system to uncover the possible link between mRNA localization and epithelial polarity in C. elegans. Together our results outline a powerful methodology to monitor mRNA through its life cycle. This approach offers important advantages compared with other gene expression techniques already available in this experimentally tractable model organism. This work is supported by Multi-Year Research Grant, University of Macau (MYRG) project 2017-00082-FHS.

Li Z et al. (2023) PNAS Nexus "A robotic system for automated genetic manipulation and analysis of Caenorhabditis elegans."

He S, Kassouni A, Bowlin PD, Fang-Yen C, Teng C, Raizen DM, Du A, Li Z, Ji H, Fan Y, Fouad AD, Chang MC

[PNAS Nexus, 2023]

The nematode <i>Caenorhabditis elegans</i> is one of the most widely studied organisms in biology due to its small size, rapid life cycle, and manipulable genetics. Research with <i>C. elegans</i> depends on labor-intensive and time-consuming manual procedures, imposing a major bottleneck for many studies, especially for those involving large numbers of animals. Here, we describe a general-purpose tool, WormPicker, a robotic system capable of performing complex genetic manipulations and other tasks by imaging, phenotyping, and transferring <i>C. elegans</i> on standard agar media. Our system uses a motorized stage to move an imaging system and a robotic arm over an array of agar plates. Machine vision tools identify animals and assay developmental stage, morphology, sex, expression of fluorescent reporters, and other phenotypes. Based on the results of these assays, the robotic arm selectively transfers individual animals using an electrically self-sterilized wire loop, with the aid of machine vision and electrical capacitance sensing. Automated <i>C. elegans</i> manipulation shows reliability and throughput comparable with standard manual methods. We developed software to enable the system to autonomously carry out complex protocols. To validate the effectiveness and versatility of our methods, we used the system to perform a collection of common <i>C. elegans</i> procedures, including genetic crossing, genetic mapping, and genomic integration of a transgene. Our robotic system will accelerate <i>C. elegans</i> research and open possibilities for performing genetic and pharmacological screens that would be impractical using manual methods.