Zhang C et al. (2012) RNA Biol "New insights into siRNA amplification and RNAi."

Zhang C, Ruvkun G

[RNA Biol, 2012]

In the nematode Caenorhabditis elegans (C. elegans), gene inactivation by RNA interference can achieve remarkable potency due to the amplification of initial silencing triggers by RNA-dependent RNA polymerases (RdRPs). RdRPs catalyze the biogenesis of an abundant species of secondary small interfering RNAs (siRNAs) using the target mRNA as template. The interaction between primary siRNAs derived from the exogenous double-stranded RNA (dsRNA) trigger and the target mRNA is required for the recruitment of RdRPs. Other genetic requirements for RdRP activities have not been characterized. Recent studies have identified the RDE-10/RDE-11 complex which interacts with the primary siRNA bound target mRNA and acts upstream of the RdRPs. rde-10 and rde-11 mutants show an RNAi defective phenotype because the biogenesis of secondary siRNAs is completely abolished. In addition, the RDE-10/RDE-11 complex plays a similar role in the endogenous RNAi pathway for the biogenesis of a subset of siRNAs targeting recently acquired, duplicated genes.

Zhang C et al. (2019) Cells "Membrane-Bound Meet Membraneless in Health and Disease."

Rabouille C, Zhang C

[Cells, 2019]

Membraneless organelles (MLOs) are defined as cellular structures that are not sealed by a lipidic membrane and are shown to form by phase separation. They exist in both the nucleus and the cytoplasm that is also heavily populated by numerous membrane-bound organelles. Even though the name membraneless suggests that MLOs are free of membrane, both membrane and factors regulating membrane trafficking steps are emerging as important components of MLO formation and function. As a result, we name them biocondensates. In this review, we examine the relationships between biocondensates and membrane. First, inhibition of membrane trafficking in the early secretory pathway leads to the formation of biocondensates (P-bodies and Sec bodies). In the same vein, stress granules have a complex relationship with the cyto-nuclear transport machinery. Second, membrane contributes to the regulated formation of phase separation in the cells and we will present examples including clustering at the plasma membrane and at the synapse. Finally, the whole cell appears to transit from an interphase phase-separated state to a mitotic diffuse state in a DYRK3 dependent manner. This firmly establishes a crosstalk between the two types of cell organization that will need to be further explored.

Zhang C et al. (2013) Biotechnol Adv "The olfactory signal transduction for attractive odorants in Caenorhabditis elegans."

Chen Y, Zhang C, Huang X, Zhang K, Yan J, Chen C

[Biotechnol Adv, 2013]

Olfaction in Caenorhabditis elegans is a versatile and sensitive strategy to seek food and avoid danger by sensing volatile chemicals emitted by the targets. The ability to sense attractive odor is mainly accomplished by the AWA and AWC neurons. Previous studies have shown the components of the olfaction signal pathway in these two amphid chemosensory neurons, but integration of the individual signaling components requires further elucidation. Here we review the progresses in our understanding of signal pathways for attractive olfaction involving AWA and AWC neurons, and discuss how the different signal molecules might employ the common molecular cascades to transduce the olfactory system and guide behavior in each neuron.

Zhang C et al. (2012) Curr Biol "The Caenorhabditis elegans RDE-10/RDE-11 complex regulates RNAi by promoting secondary siRNA amplification."

Fischer SE, Montgomery TA, Ruvkun G, Fahlgren N, Zhang C, Riedel CG, Sullivan CM, Garcia SM, Carrington JC

[Curr Biol, 2012]

BACKGROUND: In nematodes, plants, and fungi, RNAi is remarkably potent and persistent due to the amplification of initial silencing signals by RNA-dependent RNA polymerases (RdRPs). In Caenorhabditis elegans (C. elegans), the interaction between the RNA-induced silencing complex (RISC) loaded with primary small interfering RNAs (siRNAs) and the target messenger RNA (mRNA) leads to the recruitment of RdRPs and synthesis of secondary siRNAs using the target mRNA as the template. The mechanism and genetic requirements for secondary siRNA accumulation are not well understood. RESULTS: From a forward genetic screen for C. elegans genes required for RNAi, we identified rde-10, and through proteomic analysis of RDE-10-interacting proteins, we identified a protein complex containing the new RNAi factor RDE-11, the known RNAi factors RSD-2 and ERGO-1, and other candidate RNAi factors. The RNAi defective genes rde-10 and rde-11 encode a novel protein and a RING-type zinc finger domain protein, respectively. Mutations in rde-10 and rde-11 genes cause dosage-sensitive RNAi deficiencies: these mutants are resistant to low dosage but sensitive to high dosage of double-stranded RNAs. We assessed the roles of rde-10, rde-11, and other dosage-sensitive RNAi-defective genes rsd-2, rsd-6, and haf-6 in both exogenous and endogenous small RNA pathways using high-throughput sequencing and qRT-PCR. These genes are required for the accumulation of secondary siRNAs in both exogenous and endogenous RNAi pathways. CONCLUSIONS: The RDE-10/RDE-11 complex is essential for the amplification of RNAi in C. elegans by promoting secondary siRNA accumulation.

Zhang C et al. (2016) J Biol Chem "The signaling pathway of C. elegans mediates chemotaxis response to the attractant 2-heptanone in a 'Trojan Horse'- like pathogenesis."

Zhang D, Zhao N, Huang X, Yan J, Chen Y, Zou W, Zhang C, Zhang K

[J Biol Chem, 2016]

The nematode Caenorhabditis elegans exhibits behavioral responses to a wide range of odorants associated with food and pathogens. A previous study described a Trojan Horse like strategy of pathogenesis whereby the bacterium Bacillus subtilis emits the volatile organic compound 2-heptanone to trap C. elegans for successful infection. Here, we further explored the receptor for 2-heptanone as well as the pathway involved in signal transduction in C. elegans. Our experiments showed that 2-heptanone sensing depended on the function of AWC neurons and a GPCR encoded by str-2. Consistent with the above observation, the HEK293 cells expressing STR-2 on their surfaces showed a transient elevation in intracellular Ca2+ levels after 2-heptanone applications. After combining the assays of RNA interference and gene mutants, we also identified the G subunits and their downstream components in the olfactory signal cascade that are necessary for responding to 2-heptanone, including G subunits of egl-30 and gpa-3, phospholipase C of plc-1and egl-8, and the calcium channel of cmk-1 and cal-1. Our work demonstrates for the first time that an integrated signaling pathway for 2-heptanone response in C. elegans involves recognition by GPCR STR-2, activation by G subunits of egl-30/gpa-3 and transfer to the PLC pathway, indicating that a potentially novel olfactory pathway exists in AWC neurons. Meanwhile, since 2-heptanone, a metabolite from the pathogenic bacterium B. nematocida B16, can be sensed by C. elegans and thus strongly attract its host, our current work also suggested coevolution between the pathogenic microorganism and the chemosensory system in C. elegans.

Zhang C et al. (2020) Insect Sci "A new bacteria-free strategy induced by MaGal2 facilitates pinewood nematode escape immune response from its vector beetle."

Sun J, Zhang C, Zhao L, Wickham JD

[Insect Sci, 2020]

Symbiotic microbes play a crucial role in regulating parasite-host interactions; however, the role of bacterial associates in parasite-host interactions requires elucidation. In this study, we showed that, instead of introducing numerous symbiotic bacteria, dispersal of fourth-stage juvenile (J_IV ) pinewood nematodes (PWNs), Bursaphelenchus xylophilus, only introduced few bacteria to its vector beetle, Monochamus alternatus (Ma). J_IV showed weak binding ability to five dominant bacteria species isolated from the beetles' pupal chamber. This was especially the case for binding to the opportunistic pathogenic species Serratia marcescens; the nematodes' bacteria binding ability at this critical stage when it infiltrates Ma for dispersal was much weaker compared with Caenorhabditis elegans, Diplogasteroides asiaticus, and propagative-stage PWN. The associated bacterium S. marcescens, which was isolated from the beetles' pupal chambers, was unfavorable to Ma, because it caused a higher mortality rate upon injection into tracheae. In addition, S. marcescens in the tracheae caused more immune effector disorders compared with PWN alone. Ma_Galectin2 (MaGal2), a pattern-recognition receptor, was up-regulated following PWN loading. Recombinant MaGal2 protein formed aggregates with five dominant associated bacteria in vitro. Moreover, MaGal2 knockdown beetles had up-regulated prophenoloxidase gene expression, increased phenoloxidase activity, and decreased PWN loading. Our study revealed a previously unknown strategy for immune evasion of this plant pathogen inside its vector, and provides novel insights into the role of bacteria in parasite-host interactions. This article is protected by copyright. All rights reserved.

Zhang C et al. (2011) Proc Natl Acad Sci U S A "mut-16 and other mutator class genes modulate 22G and 26G siRNA pathways in Caenorhabditis elegans."

Fahlgren N, Zhang C, Ruvkun G, Phillips CM, Carrington JC, Montgomery TA, Gabel HW, Sullivan CM, Fischer SE

[Proc Natl Acad Sci U S A, 2011]

Argonaute-associated siRNAs and Piwi-associated piRNAs have overlapping roles in silencing mobile genetic elements in animals. In Caenorhabditis elegans, mutator (mut) class genes mediate siRNA-guided repression of transposons as well as exogenous RNAi, but their roles in endogenous RNA silencing pathways are not well-understood. To characterize the endogenous small RNAs dependent on mut class genes, small RNA populations from a null allele of mut-16 as well as a regulatory mut-16(mg461) allele that disables only somatic RNAi were subjected to deep sequencing. Additionally, each of the mut class genes was tested for a requirement in 26G siRNA pathways. The results indicate that mut-16 is an essential factor in multiple endogenous germline and somatic siRNA pathways involving several distinct Argonautes and RNA-dependent RNA polymerases. The results also reveal essential roles for mut-2 and mut-7 in the ERGO-1 class 26G siRNA pathway and less critical roles for mut-8, mut-14, and mut-15. We show that transposons are hypersusceptible to mut-16-dependent silencing and identify a requirement for the siRNA machinery in piRNA biogenesis from Tc1 transposons. We also show that the soma-specific mut-16(mg461) mutant allele is present in multiple C. elegans laboratory strains.

Zhang C et al. (2008) J Theor Biol "Microsatellites that violate Chargaff's second parity rule have base order-dependent asymmetries in the folding energies of complementary DNA strands and may not drive speciation."

Zhang C, Wei JF, Xu S, Forsdyke DR

[J Theor Biol, 2008]

Models for meiotic recombination based on Crick''s "unpairing postulate" require symmetrical extrusion of stem-loop structures from homologous DNA duplexes. The potential for such extrusion is abundant in many species and, for a given single-strand segment, can be quantitated as the "folding of natural sequence" (FONS) energy value. This, in turn, can be decomposed into base order-dependent and base composition-dependent components. The FONS values of top and bottom strands in most Caenorhabditis elegans segments are close, as are the corresponding base order-dependent and base composition-dependent components; any discrepancies are in the base composition-dependent component. This suggests that the strands would extrude with similar kinetics. However, interspersed among these segments and at the ends of chromosomes (telomeres) are segments containing short tandem repeats (microsatellites) which, by virtue of their high variability, have been postulated to inhibit the pairing of homologous chromosomes and hence drive speciation. In these segments, there are usually wide discrepancies between the FONS values of top and bottom strands, mainly attributable to differences in base order-dependent components. Analyses of artificial microsatellites of different unit sizes and base compositions show that this asymmetrical distribution of folding potential is greatest for microsatellites when the units are short and violate Chargaff''s second parity rule. It is proposed that when there is folding asymmetry, recombination proceeds by special, strand-biased, somatic mechanisms analogous to those operating with Chi sequences in Escherichia coli. If meiotic recombination in the germ-line requires extrusion symmetry, then a general inhibitory influence of microsatellite-containing segments could mask the antirecombinational influence of their variability. Thus, microsatellites may not have driven speciation.

W Zhang et al. (1999) International C. elegans Meeting "Analysis of several mutations that disrupt vulval development"

M Han, W Zhang, A Mapes

[International C. elegans Meeting, 1999]

We have conducted ts and sterile mutant screens for mutations that disrupt various steps of vulval development. Analyses of several mutations will be reported. ku260 and ku272 . Mutations have similar vulval phenotype likely due to defects in VPC generation. Mutants display either an everted vulva (Evl or Pvl) or fail entirely to differentiate vulval tissue. Nomarski analysis indicates that such mutants are often missing more than one Pn.p cell, resulting in either abnormal or no vulval invaginations. Using an unc-47::GFP as a marker, we observed that many GABAergic neurons in the ventral cord are absent (approximately 50% missing). ku260 homozygotes are Unc, likely due to the missing VD neurons. Both ku260 and ku272 mutants have an abnormal somatic gonad and contain no oocytes, leading to a sterile phenotype. ku260 was mapped to LGIV between unc-24 and unc-129 and is uncovered by eDf19 . Pools of cosmids are being injected in order to rescue the mutant phenotype. ku260 fails to complement evl-24(ar124) which was previously isolated by Seydoux and Greenwald as a class III Evl mutation(1). ku272 has been mapped to right arm of LGI. ku259, ku255 and ku270 . These mutations appear to cause defects in both vulval lineage and morphogenesis and display an Evl phenotype (subtle for ku270 ). We also detect a low to moderate percentage of double invaginations and protrusions in ku259 and ku255 mutant worms. Nomarski analysis indicates that the abnormal or double invaginations observed are at least partly due to lineage defects. All three mutations also cause a sterile phenotype (incomplete for ku270 ). ku259 was mapped to LGIII between dpy-1 and daf-2 . DNA-mediated transformation is currently being performed to identify the affected gene. ku255 was mapped between unc-13 and nob-3 on LGI and failed to complement evl-17(ar94) (1). ku270 has been mapped to the right of lin-25 on LGV. The cloning of both genes is in progress. 20-30% of ku273 homozygous mutants are Egl and some contain two or three vulval protrusions. Nomarski analysis reveals that the Egl and Muv phenotype are caused by defects in vulval lineage and morphogenesis. We are currently mapping this gene to a chromosomal location. (1) Seydoux, G., C. Savage, and Greenwald. I. (1993). Dev. Biol. 157,423-436.

Shenghyuan Zhang et al. (2001) International C. elegans Meeting "Suppressors that recover daf-7::gfp expression in tph-1(mg280)"

Shenghyuan Zhang, Ji Ying Sze

[International C. elegans Meeting, 2001]

tph-1 encodes a tryptophan hydroxylase, the key enzyme for serotonin biosynthesis. Our previous studies show that daf-7::gfp expression is reduced 6-fold in the deletion mutant tph-1(mg280). This suggests that serotonin may normally up-regulate daf-7 expression. To isolate genes that upregulate daf-7 expression in response to serotonin signaling, we have been screening mutagenized tph-1(mg280) worms for suppressors that recover daf-7::gfp expression. We have clonally screened 3,870 haploid genomes and isolated 41 suppressors, 15 of them are being back-crossed. DiI staining of back-crossed suppressors showed a normal dye filling pattern, suggesting that the mutations do not grossly compromise the integrity of the chemosensory neurons. Further examination of the back-crossed suppressors has led us to classify the suppressors into two classes. The first class of the suppressors shows a strong daf-7::gfp expression only in L2 and L3 tph-1(mg280). The second class of the suppressors shows a strong daf-7::gfp expression even in the adults. The results of genetic and molecular characterization of the suppressors will be discussed.