Zhang, Chi et al. (2009) International Worm Meeting "rde-10 is required for efficient RNAi."

Zhang, Chi, Mak, Ho Yi, Yang, Huan, Ruvkun, Gary

[International Worm Meeting, 2009]

To identify new genes required for RNAi as well as for the trafficking of the silencing signals, we established a reporter strain that allows easy visualization of an efficient RNAi response. We generated constructs that express gfp hairpin dsRNAs under the control of neuron-specific promoters (e.g., snb-1 or ric-19). We then introduced these constructs into sur-5::gfp transgenic worms expressing GFP in most somatic cells. Although the neuronally expressed dsRNAs do not affect GFP fusion gene expression cell-autonomously, they potently silence GFP expression in intestine, muscle and hypodermal cells in wild type animals. We carried out a large-scale ethyl methanesulfonate (EMS) mutagenesis screen for mutations that reanimate GFP expression in the somatic tissues. From this screen we identified rde-10, a gene that is conserved in the Caenorhabditis genus but not in other organisms. The rde-10 mutant is resistant to feeding dsRNAs targeting many genes expressed in either germline or somatic tissues (e.g. pos-1, elt-2, lin-29 and nhr-23). However, this mutant is not completely deficient in RNAi; it is still sensitive to feeding unc-22 dsRNA, as well as injected pos-1 dsRNA. Similar to rde-1 and rde-4, and in contrast to mutators, rde-10 mutations do not cause other phenotypic abnormalities. The mutant has normal brood size at various growth temperatures, normal germline cytology, and does not have high incidence of chromosome loss during meiosis. In addition to its role in the exogenous RNAi pathway, rde-10 is also required for the biogenesis and/or stability of certain classes of endogenous small RNAs. For example, the level of X-cluster tiny non-coding RNAs derived from a unique region on the X chromosome, is significantly reduced in the rde-10 mutant. rde-10 function is being further characterized using a combination of genetic, molecular, cellular and biochemical approaches. Studies are underway to determine whether rde-10 is required for the biogenesis or activity of small interference RNAs (siRNAs) derived from the exogenous long dsRNA triggers. We are conducting experiments to identify its interacting protein factors and associated small RNAs. We will also explore how rde-10 mutations affect endogenous small RNA levels by Solexa deep sequencing technology.

Wu Y et al. (2003) Filaria J "Human immune responses to infective stage larval-specific chitinase of filarial parasite, Onchocerca volvulus, Ov-CHI-1."

Nutman TB, Bianco AE, McCarthy JS, Wu Y, Egerton G

[Filaria J, 2003]

BACKGROUND: Ov-CHI-1 is a chitinase specifically expressed in the infective stage larvae of the human filarial parasite Onchocerca volvulus. Evidence has show that it could be a vaccine candidate, however, there is no data available regarding the immunological status of people naturally exposed to infective stage larvae and thus provoked by this antigen. METHOD: We analysed the Ov-CHI-1-specific immune response present in four endemic foci of human onchocerciasis (Ecuador, Nigeria, Togo and Cameroon) by enzyme-linked immunosorbent assays and T-cell proliferation assays. RESULTS: In these foci of infection, antibodies to Ov-CHI-1 were found to be present in only 22% of individuals from Ecuador, but were detected in 42-62% of infected individuals in the three foci from West Africa (Nigeria, Togo and Cameroon). There was found to be no relationship between antibody level and age, gender, or infection intensity as indicated by microfilarial density and numbers of skin nodules. The isotype response to Ov-CHI-1 was dominated by the presence of IgG3, IgG1 was present to a lesser extent. Our results show a positive correlation between N- and C-termini of Ov-CHI-1 in their ability to provoke humoral and cellular immune responses in the human. Peripheral blood mononuclear cell (PBMC) proliferative responses to Ov-CHI-1 when assayed, were found to be significantly higher in the individuals from endemic areas and there was a statistically elevated response to Ov-CHI-1 in the infected individuals when compared to putative immune individuals. CONCLUSION: Ov-CHI-1 is an antigen that we have found strongly induces both humoral and cellular immune responses in humans.

Wu Y et al. (2001) J Biol Chem "Expression and secretion of a larval-specific chitinase (family 18 glycosyl hydrolase) by the infective stages of the parasitic nematode, Onchocerca volvulus."

Sakuda S, Egerton G, Bianco AE, Underwood AP, Wu Y

[J Biol Chem, 2001]

A recently reported chitinase gene, expressed in the infective, third-stage (L3) larvae of the human filarial parasite Onchocerca volvulus, belongs to the family 18 glycosyl hydrolases and has been designated Ov-chi-1. The gene product of Ov-chi-1 is chitinolytic. Allosamidin ablates activity of the native enzyme in a dose-dependent manner but did not significantly inhibit the moulting of L3 larvae. Mono-specific antibodies were used to characterize Ov-CHI-1 as a 60-kDa protein expressed almost exclusively in L3 stages. Immunoelectron microscopy showed that Ov-CHI-1 expression is initiated in late L2 larvae and increases markedly in infective, L3 larvae. It is synthesized exclusively in the glandular esophagus and stored within discrete secretory granules. Secretion occurs through de-granulation during post-infective development, and the primary route of transport appears to be via the pseudo-coelom. An orthologue of Ov-chi-1 was detected in Caenorhabditis elegans by BLAST analysis. It is constitutively expressed at a low level and is overexpressed in dauer larvae and embryonated eggs. It is chitinolytic. We conclude that Ov-CHI-1 is a highly stage-specific enzyme that may have a role in infectivity of the parasite, aiding escape from the vector or participating in early post-infective migration and/or development. The identification of an orthologue in C. elegans opens the way for further studies into the biological function(s) of this intriguing parasite product.

Shi C et al. (2016) Genes Dev "Feeding the germline."

Murphy CT, Shi C

[Genes Dev, 2016]

Nucleotides are required in order to replicate DNA in the developing germline. Here, Chi and colleagues (pp. 307-320) have used Caernohabditis elegans to identify a GLP-1-dependent checkpoint that senses food (bacterially)-supplied nucleotide levels, arresting reproductive development in the absence of sufficient nucleotide supplies.

Irazoqui JE (2015) Immunity "Why worms watch their hemidesmosomes and why you should, too."

Irazoqui JE

[Immunity, 2015]

Hemidesmosomes are cellular attachment structures of great importance to the epidermis. In this issue of Immunity, Zhang et al. (2015) have discovered that in addition to having structural functions, invertebrate and human hemidesmosomes are actively monitored by the cell as a novel mechanism for detecting pathogenic infection.

Kumsta C et al. (2019) J Cell Biol "Getting under the skin: Cuticle damage elicits systemic autophagy response in C. elegans."

Hansen M, Kumsta C

[J Cell Biol, 2019]

In this issue, Zhang et al. (2019. <i>J. Cell. Biol.</i> https://doi.org/10.1083/jcb.201907196) describe a molecular mechanism by which cuticular damage in the nematode <i>C. elegans</i> leads to systemic induction of autophagy by signals propagated from sensory neurons via the TGF- signaling pathway.

Svendsen JM et al. (2018) Dev Cell "piRNA Rules of Engagement."

Svendsen JM, Montgomery TA

[Dev Cell, 2018]

piRNAs are known to silence transposable elements, but not all piRNAs match transposon sequences. Recent studies from Shen etal. (2018) and Zhang etal. (2018) identify rules for piRNA target recognition in Caenorhabditis elegans. Permissive pairing rules allow targeting of essentially all germline mRNAs, while protective mechanisms prevent silencing self-genes.

McDiarmid, Troy A et al. (2020) MicroPubl Biol

Kepler, Lexis D, McDiarmid, Troy A, Rankin, Catharine H

[MicroPubl Biol, 2020]

The Auxin-Inducible Degradation (AID) system is a powerful technique in the C. elegans toolkit that enables conditional and reversible protein depletion with high temporal and spatial specificity (Zhang et al. 2015; Martinez et al. 2020; Ashley et al. 2020; Martinez and Matus 2020). This system relies on tagging a gene of interest with a short AID degron sequence and transgenic expression of TIR1, an inducible E3 ubiquitin ligase normally found only in plants (Nishimura et al. 2009; Zhang et al. 2015). Upon exposure to the plant-derived hormone Auxin, TIR1 is activated and targets AID-tagged proteins for proteasomal degradation (Nishimura et al. 2009; Zhang et al. 2015) (Figure 1A). While there are qualitative reports that Auxin does not overtly affect the morphology or behavior of wild-type C. elegans (Zhang et al. 2015), this has not been quantitatively assessed. Determining whether Auxin significantly affects C. elegans morphology and behavior, even in subtle ways, is important given the C. elegans communitys rapid uptake of the AID system (Kasimatis et al. 2018; Nance and Frkjr-Jensen 2019; Ashley et al. 2020; McDiarmid et al. 2020). Here, we use our high-throughput machine vision tracking system, the Multi-Worm Tracker (MWT) (Swierczek et al. 2011), to investigate whether exposure to Auxin affects a suite of morphological, locomotor, mechanosensory, and short-term habituation learning phenotypes in our labs derivate of Bristol N2 wild-type worms and the CGC wild-type reference strain, PD1074 (Yoshimura et al. 2019) (Figure 1B).

Robertson FM (2010) IDrugs "High-Content Analysis--CHI's Seventh Annual Conference--Developments in HCA and Pathway Analysis."

Robertson FM

[IDrugs, 2010]

CHI''s Seventh Annual Conference on High-Content Analysis (HCA), held in San Francisco, incorporated topics covering new developments in the field of HCA, including hardware and software updates, new biological models for HCA and pathway analysis. This conference report highlights selected presentations on the use of HCA for the characterization of stem cells, cell-colony analysis, the validation of disease models and the identification of antiparasitic compounds.

Fischer, Sylvia E. J. et al. Worm Breeder's Gazette "Prevalent, spontaneously occurring mutations at the eri-6/7 locus result in an Eri phenotype"

Fischer, Sylvia E. J., Ruvkun, Gary, Zhang, Chi

[Worm Breeder's Gazette]