Liu, Z. et al. (2015) International Worm Meeting "Existence of a nematode alarm pheromone."

Zhong, W., Nguyen, J., Aleman-Meza, B., Liu, Z., Jung, S.

[International Worm Meeting, 2015]

It is crucial for animal survival to detect dangers such as predators. A good indicator of dangers is injury of conspecifics. It was reported that in C. elegans when one worm exploded upon a glass needle, nearby worms would move away from the victim (Thomas and Horvitz, 1988). Using three newly-designed quantitative methods, we confirmed the existence of an alarm pheromone released by injured nematodes. C. elegans avoided the alarm pheromone in a dose-dependent pattern. We then explored the molecular properties of the alarm pheromone and found that it was not any of the known avoidance chemicals. The alarm pheromone appeared to be ascaroside-independent because ascaroside-synthesis mutants still generated the alarm pheromone and ascaroside-receptor mutants still detected the alarm pheromone. The alarm pheromone was an endogenous factor present in all developmental stages, including embryos, larvae, adults and dauers, and only released to the environment upon injury. The alarm pheromone appeared to be nematode specific, because C. elegans did not avoid injured fruit fly larvae, but avoided alarm pheromones from multiple nematode species, including the insect parasite Steinernema. Interestingly, Steinernema only avoided the alarm pheromone from Steinernema but not C. elegans. We further examined the effects of the alarm pheromone on C. elegans. Constant exposure to the alarm pheromone did not reduce the lifespan of C. elegans, suggesting that it imposed no physical damage to the worms. Avoidance of the alarm pheromone was not affected by gender, developmental stage or feeding status. Males, hermaphrodites, dauers, and starved worms showed equal avoidance of this alarm pheromone, suggesting that avoidance of the alarm pheromone was a basic response important for animal survival in the wild environment. Multiple neurotransmitters were required for the alarm response. Exposure to the anti-anxiety drug Fluoxetine (Prozac) abolished C. elegans avoidance to the alarm pheromone. Finally, we investigated the molecular and cellular pathways used by C. elegans to detect the alarm pheromone. Mutant analysis showed that detection of the alarm pheromone was modulated through the cGMP-gated ion channels TAX-2 and TAX-4, guanylyl cyclases DAF-11, ODR-1, GCY-5, GCY-9 and GCY-22, and G proteins GOA-1, GPA-3, GPA-7 and GPA-14. Genetic and laser ablation of neurons showed that the amphid neurons AWC, ASI, and ASK were required for the detection of the alarm pheromone. .

Liu Z et al. (2017) J Parasitol Res "Identification, Characterization, and Structure of Tm16 from Trichuris muris."

Liu Z, Hotez PJ, Asojo OA, Wei J, Bottazzi ME, Pollet J, Tabb S, Zhan B, Kelleher A

[J Parasitol Res, 2017]

Trichuriasis is a disease of poverty for which excretory and secretory (ES) products that induce the protective immunity are being investigated as candidate vaccines antigens. In this study, ES products of T. muris and immune sera were produced. The immune sera recognized more than 20 proteins on a 2D-gel of ES products of T. muris adult worms. Tm16 was one of the proteins identified by mass spectrometry. Tm16 shares 57% sequence identity with Ov16, an immunodominant diagnostic antigen from Onchocerca volvulus. Recombinant Tm16 with a carboxyl terminal hexahistidine was produced using Pichia pastoris. Polyclonal antibodies against rTm16 were generated by one-prime and two-boost immunization of three female Balb/c mice with 25g of recombinant Tm16 emulsified with ISA720 adjuvant. These polyclonal antibodies confirmed that Tm16 is localized to the ES products and the soluble fraction of the adult worm. Additionally, the high-resolution crystal structure of Tm16 was solved by molecular replacement. Tm16 belongs to the phosphatidylethanolamine-binding-like protein (PEBP1) family and this is the first structure of a PEBP1 from a parasite.

Liu Z et al. (2018) Nat Commun "Predator-secreted sulfolipids induce defensive responses in C. elegans."

Pribadi AK, Tong A, Bose N, Srinivasan J, Kariya MJ, Schroeder FC, Curran KP, Chalasani SH, Leinwand SG, Liu Z, Chute CD

[Nat Commun, 2018]

Animals respond to predators by altering their behavior and physiological states, but the underlying signaling mechanisms are poorly understood. Using the interactions between Caenorhabditis elegans and its predator, Pristionchus pacificus, we show that neuronal perception by C. elegans of a predator-specific molecular signature induces instantaneous escape behavior and a prolonged reduction in oviposition. Chemical analysis revealed this predator-specific signature to consist of a class of sulfolipids, produced by a biochemical pathway required for developing predacious behavior and specifically induced by starvation. These sulfolipids are detected by four pairs of C. elegans amphid sensory neurons that act redundantly and recruit cyclic nucleotide-gated (CNG) or transient receptor potential (TRP) channels to drive both escape and reduced oviposition. Functional homology of the delineated signaling pathways and abolishment of predator-evoked C. elegans responses by the anti-anxiety drug sertraline suggests a likely conserved or convergent strategy for managing predator threats.

Liu, Z. et al. (2017) International Worm Meeting "Sulfolipids drive C. elegans defensive responses to a predator."

Pribadi, A., Srinivasan, J., Liu, Z., Curran, K., Leinwand, S., Schroeder, F., Bose, N., Chalasani, S., Tong, A., Chute, C., Kariya, M.

[International Worm Meeting, 2017]

Animals respond to predator threats by implementing characteristic behavioural and physiological changes, but it is not known how the underlying neuronal ensembles process these threats. The simple, well-defined nervous system of the nematode, Caenorhabditis elegans facilitates uncovering conserved principles in neuronal processing. Here we show that C. elegans responds to excretions from a nematode predator, Pristionchus pacificus with instantaneous escape behaviour and a prolonged reduction of oviposition. Chemical analysis of the P. pacificus exo-metabolome revealed a series of novel sulfated lipids that instruct these C. elegans responses. These sulfolipids are specific to the predator and originate from a biosynthetic pathway that ties into endocrine signalling required for predacious behavior.

Liu Z et al. (2015) PLoS Genet "Promotion of bone morphogenetic protein signaling by tetraspanins and glycosphingolipids."

Sheth S, Amin NM, Tian C, He J, Kubba S, Aydin T, Liu Z, Zimmerman S, Constas K, Liu D, Ranjan S, Shwartz N, McMahon DE, Plavskin Y, Schwarz EM, Krause MW, Liu J, Schaeffer A, Alam E, Dad NA, Lindy A, Smith H, Shi H, Yun S, Szymczak LC

[PLoS Genet, 2015]

Bone morphogenetic proteins (BMPs) belong to the transforming growth factor (TGF) superfamily of secreted molecules. BMPs play essential roles in multiple developmental and homeostatic processes in metazoans. Malfunction of the BMP pathway can cause a variety of diseases in humans, including cancer, skeletal disorders and cardiovascular diseases. Identification of factors that ensure proper spatiotemporal control of BMP signaling is critical for understanding how this pathway is regulated. We have used a unique and sensitive genetic screen to identify the plasma membrane-localized tetraspanin TSP-21 as a key new factor in the C. elegans BMP-like "Sma/Mab" signaling pathway that controls body size and postembryonic M lineage development. We showed that TSP-21 acts in the signal-receiving cells and genetically functions at the ligand-receptor level. We further showed that TSP-21 can associate with itself and with two additional tetraspanins, TSP-12 and TSP-14, which also promote Sma/Mab signaling. TSP-12 and TSP-14 can also associate with SMA-6, the type I receptor of the Sma/Mab pathway. Finally, we found that glycosphingolipids, major components of the tetraspanin-enriched microdomains, are required for Sma/Mab signaling. Our findings suggest that the tetraspanin-enriched membrane microdomains are important for proper BMP signaling. As tetraspanins have emerged as diagnostic and prognostic markers for tumor progression, and TSP-21, TSP-12 and TSP-14 are all conserved in humans, we speculate that abnormal BMP signaling due to altered expression or function of certain tetraspanins may be a contributing factor to cancer development.

Liu Z et al. (2013) Development "The micronutrient element zinc modulates sperm activation through the SPE-8 pathway in Caenorhabditis elegans."

Miao L, Shang Y, Liu Z, Chen L, Huang P

[Development, 2013]

Immotile spermatids produced in the testis must undergo a series of poorly understood morphological, physiological and biochemical processes called sperm activation to become motile, fertilization-competent spermatozoa. In Caenorhabditis elegans, the spe-8 group contains sperm-specific genes active in both males and hermaphrodites, although their activity is required only for hermaphrodite self-sperm activation. The activating signal upstream of the SPE-8 signaling cascade remains unknown. Here, we show that the micronutrient zinc is sufficient to trigger sperm activation in vitro, and that extracellular zinc induces the intracellular redistribution of labile zinc. We demonstrate that other activating signals promote the similar redistribution of labile zinc, indicating that zinc might have first and/or second messenger roles during sperm activation. Moreover, zinc-induced sperm activation is SPE-8 pathway dependent. Labile zinc was enriched in the spermatheca, the normal site for self-sperm activation in hermaphrodites. High levels of zinc were also found in the secretory cells in the male gonad, suggesting that zinc might be secreted from these cells during copulation and become a component of seminal fluid, to modulate sperm activation post-copulation. These data indicate that zinc regulates sperm activation in both male and hermaphrodite C. elegans, a finding with important implications for understanding hermaphroditic evolution.

Liu Z et al. (2022) PLoS Genet "The C. elegans TspanC8 tetraspanin TSP-14 exhibits isoform-specific localization and function."

Shi H, Liu J, Liu Z

[PLoS Genet, 2022]

Tetraspanin proteins are a unique family of highly conserved four-pass transmembrane proteins in metazoans. While much is known about their biochemical properties, the in vivo functions and distribution patterns of different tetraspanin proteins are less understood. Previous studies have shown that two paralogous tetraspanins that belong to the TspanC8 subfamily, TSP-12 and TSP-14, function redundantly to promote both Notch signaling and bone morphogenetic protein (BMP) signaling in C. elegans. TSP-14 has two isoforms, TSP-14A and TSP-14B, where TSP-14B has an additional 24 amino acids at its N-terminus compared to TSP-14A. By generating isoform specific knock-ins and knock-outs using CRISPR, we found that TSP-14A and TSP-14B share distinct as well as overlapping expression patterns and functions. While TSP-14A functions redundantly with TSP-12 to regulate body size and embryonic and vulva development, TSP-14B primarily functions redundantly with TSP-12 to regulate postembryonic mesoderm development. Importantly, TSP-14A and TSP-14B exhibit distinct subcellular localization patterns. TSP-14A is localized apically and on early and late endosomes. TSP-14B is localized to the basolateral cell membrane. We further identified a di-leucine motif within the N-terminal 24 amino acids of TSP-14B that serves as a basolateral membrane targeting sequence, and showed that the basolateral membrane localization of TSP-14B is important for its function. Our work highlights the diverse and intricate functions of TspanC8 tetraspanins in C. elegans, and demonstrates the importance of dissecting the functions of these important proteins in an intact living organism.

Liu Z et al. (2014) Biochim Biophys Acta "Calcium signaling and the MAPK cascade are required for sperm activation in Caenorhabditis elegans."

Liu Z, Miao L, Wang B, Zhao Y, He R

[Biochim Biophys Acta, 2014]

In nematode, sperm activation (or spermiogenesis), a process in which the symmetric and non-motile spermatids transform into polarized and crawling spermatozoa, is critical for sperm cells to acquire fertilizing competence. SPE-8 dependent and SPE-8 independent pathways function redundantly during sperm activation in both males and hermaphrodites of Caenorhabditis elegans. However, the downstream signaling for both pathways remains unclear. Here we show that calcium signaling and the MAPK cascade are required for both SPE-8 dependent and SPE-8 independent sperm activation, implying that both pathways share common downstream signaling components during sperm activation. We demonstrate that activation of the MAPK cascade is sufficient to activate spermatids derived from either wild-type or spe-8 group mutant males and that activation of the MAPK cascade bypasses the requirement of calcium signal to induce sperm activation, indicating that the MAPK cascade functions downstream of or parallel with the calcium signaling during sperm activation. Interestingly, the persistent activation of MAPK in activated spermatozoa inhibits Major Sperm Protein (MSP)-based cytoskeleton dynamics. We demonstrate that MAPK plays dual roles in promoting pseudopod extension during sperm activation but also blocking the MSP-based, amoeboid motility of the spermatozoa. Thus, though nematode sperm are crawling cells, morphologically distinct from flagellated sperm, and the molecular machinery for motility of amoeboid and flagellated sperm is different, both types of sperm might utilize conserved signaling pathways to modulate sperm maturation.

Liu Z et al. (2008) BMC Genomics "Sequence space coverage, entropy of genomes and the potential to detect non-human DNA in human samples."

Liu Z, Maley CC, Venkatesh SS

[BMC Genomics, 2008]

ABSTRACT: BACKGROUND: Genomes store information for building and maintaining organisms. Complete sequencing of many genomes provides the opportunity to study and compare global information properties of those genomes. RESULTS: We have analyzed aspects of the information content of Homo sapiens, Mus musculus, Drosophila melanogaster, Caenorhabditis elegans, Arabidopsis thaliana, Saccharomyces cerevisiae, and Escherichia coli (K-12) genomes. Virtually all possible (>98%) 12bp oligomers appear in vertebrate genomes while <2% of 19bp oligomers are present. Other species showed different ranges of >98% to <2% of possible oligomers in D. melanogaster (12-17bp), C. elegans (11-17bp), A. thaliana (11-17bp), S. cerevisiae (10-16bp) and E. coli (9-15bp). Frequencies of unique oligomers in the genomes follow similar patterns. We identified a set of 2.6M 15-mers that are more than 1 nucleotide different from all 15-mers in the human genome and so could be used as probes to detect microbes in human samples. In a human sample, these probes would detect 100% of the 433 currently fully sequenced prokaryotes and 75% of the 3065 fully sequenced viruses. The human genome is significantly more compact in sequence space than a random genome. We identified the most frequent 5- to 20-mers in the human genome, which may prove useful as PCR primers. We also identified a bacterium, Anaeromyxobacter dehalogenans, which has an exceptionally low diversity of oligomers given the size of its genome and its GC content. The entropy of coding regions in the human genome is significantly higher than non-coding regions and chromosomes. However chromosomes 1, 2, 9, 12 and 14 have a relatively high proportion of coding DNA without high entropy, and chromosome 20 is the opposite with a low frequency of coding regions but relatively high entropy. CONCLUSIONS: Measures of the frequency of oligomers are useful for designing PCR assays and for identifying chromosomes and organisms with contain hidden structure that had not been previously recognized. This information may be used to detect novel microbes in human tissues.

Liu Z et al. (2014) J Biomed Opt "Real-time brightfield, darkfield, and phase contrast imaging in a light-emitting diode array microscope."

Liu Z, Waller L, Liu S, Tian L

[J Biomed Opt, 2014]

We demonstrate a single-camera imaging system that can simultaneously acquire brightfield, darkfield, and phase contrast images in real time. Our method uses computational illumination via a programmable light-emitting diode (LED) array at the source plane, providing flexible patterning of illumination angles. Brightfield, darkfield, and differential phase contrast images are obtained by changing the LED patterns, without any moving parts. Previous work with LED array illumination was only valid for static samples because the hardware speed was not fast enough to meet real-time acquisition and processing requirements. Here, we time multiplex patterns for each of the three contrast modes in order to image dynamic biological processes in all three contrast modes simultaneously. We demonstrate multicontrast operation at the maximum frame rate of our camera (50 Hz with 2160 x 2560 pixels).