Feng XP et al. (2002) J Neurochem "Study of the nematode putative GABA type-A receptor subunits: evidence for modulation by ivermectin."

Prichard RK, Hayashi J, Feng XP, Beech RN

[J Neurochem, 2002]

Two alleles of the HG1 gene, which encodes a putative GABA receptor alpha/gamma subunit, were isolated from Haemonchus contortus. These two alleles were shown previously to be associated with ivermectin susceptibility (HG1A) and resistance (HG1E), respectively. Sequence analysis indicates that they differ in four amino acids. To explore the functional properties of the two alleles, a full-length cDNA encoding the beta subunit, a key functional component of the GABA receptor, was isolated from Caenorhabditis elegans (gab-1, corresponding to the GenBank locus ZC482.1) and coexpressed in Xenopus oocytes with the HG1 alleles. When gab-1 was coexpressed with either the HG1A allele or the HG1E allele in Xenopus oocytes, gamma-aminobutyric acid (GABA)-responsive channels with different sensitivity to the agonist were formed. The effects of ivermectin on the hetero-oligomeric receptors were determined. Application of ivermectin alone had no effect on the receptors. However, when coapplied with 10 micro m GABA, ivermectin potentiated the GABA-evoked current of the GAB-1/HG1A receptor, but attenuated the GABA response of the GAB-1/HG1E receptor. We demonstrated that the coexpressed HG1 and GAB-1 receptors are GABA-responsive, and provide evidence for the possible involvement of GABA receptors in the mechanism of ivermectin resistance.

Li T et al. (2022) STAR Protoc "Live imaging of postembryonic developmental processes in C.elegans."

Wang X, Zou Y, Li T, Feng Z

[STAR Protoc, 2022]

Live imaging is an important tool to track dynamic processes such as neuronal patterning events. Here, we describe a protocol for time-lapse microscopy analysis using neuronal migration and dendritic growth as examples. This protocol can provide detailed information for understanding cellular dynamics during postembryonic development in Caenorhabditis elegans (C. elegans). For complete details on the use and execution of this protocol, please refer to Feng etal. (2020), Li etal. (2021), and Wang etal. (2021).

J Astin et al. (2004) European Worm Meeting "CHARACTERISATION OF AN XPA-1 MUTANT IN C. ELEGANS."

J Astin, P Kuwabara, N O'Neil

[European Worm Meeting, 2004]

Many pathways have been identified in eukaryotes that are responsible for the repair of damaged DNA. In humans, repair of UV-induced photolesions is dependent on the multi step nucleotide excision repair (NER) system. An absence of NER activity in humans results in the disorder xeroderma pigmentosum (XP). XP patients suffer from extreme photosensitivity and a high incidence of skin cancers. XPA is a DNA-binding protein that is essential in the early steps of both global and transcription-coupled NER. Loss of XPA results in the most severe form of XP. Most studies involving NER have employed yeast or somatic cell lines; hence, we are interested in understanding how loss of XPA affects the development and survival of a multicellular organism such as C. elegans. A search of the C. elegans genome has led to the identification of a predicted gene, xpa-1, that encodes a protein with 42% overall amino acid identity to human XPA. We have obtained an xpa-1 mutant from the KO consortium that truncates the protein from a predicted 241 to 49 amino acids. The truncated protein lacks the conserved XPA domains, so the mutation is likely to be null. We compared the UV sensitivity of xpa-1 to N2 after exposure to different fluences of UV irradiation and found that the xpa-1 mutant is hyper-sensitive to UV at all stages of development. In addition to a decrease in survival, UV irradiation of xpa-1 mutants leads to a growth arrest. Irradiated xpa-1 embryos, larvae and dauers fail to reach adulthood. We believe the growth arrest and death of xpa-1 following UV is due to a general loss of transcription due to stalling of RNA polymerases on unrepaired UV photolesions.

Davo Martinez, Carlota et al. (2021) International Worm Meeting "Persistent DNA repair complex binding in the absence of DNA damage excision impairs neuron functionality"

Ribeiro-Silva, Cristina, Muniesa Vargas, Alba, Theil, Arjan, Davo Martinez, Carlota, Vermeulen, Wim, Lans, Hannes

[International Worm Meeting, 2021]

Nucleotide excision repair (NER) is a major DNA repair pathway that removes a large variety of helix-distorting DNA damage. Hereditary mutations in NER genes that encode proteins that make up the core NER machinery, such in endonucleases XPF and XPG, can give rise to the cancer prone disorder xeroderma pigmentosum (XP) or to more severe disorders in which XP is combined with progressive neurodegeneration or progeroid Cockayne syndrome (CS) features, called XP-CS. Intriguingly, mutations in other factors of the core NER complex give rise to a milder phenotype. It is not properly understood why mutations in genes that act in the same DNA repair pathway cause different types of disease. Here, we use C. elegans as a model to study the phenotypic impact of different mutations in the evolutionary conserved NER pathway. We found that XPF-1 or XPG-1 deletion mutants show severe developmental arrest and neuronal dysfunction upon induction of UV damage. In the absence of XPF-1 or XPG-1, core NER-intermediates accumulate at DNA damage and stay stably bound to DNA. We hypothesized that this could shield the lesion from other DNA repair pathways and block transcription, which could cause this severe phenotype. Intriguingly, we found that inhibiting the binding of the core NER-intermediates to DNA in XPF-1 or XPG-1 worms alleviated the severe phenotype, confirming that the accumulation of repair intermediates can indeed be more toxic for cells than the damage itself. Together, these results strongly suggest that the persistence of NER intermediates adversely affects cell functionality, which may be a plausible explanation for why mutations in XPF or XPG can lead to more severe disease features than mutations in other core subunits.

Oh, Jun Young et al. (2019) MicroPubl Biol

Wang, Han, Sternberg, Paul W, Gharib, Shahla, Liu, Jonathan, Oh, Jun Young

[MicroPubl Biol, 2019]

cGAL, a recently developed temperature-robust bipartite GAL4-UAS system in C. elegans, consists of two components: a cGAL driver that expresses the cGAL protein in specific cells using a promoter (i.e. neuron-specific or tissue-specific), and an effector that carries a gene of interest downstream of UAS (Wang et al., 2017). Crossing or combining a driver with an effector leads to the expression of the gene of interest in a cell-specific or tissue-specific manner.Here we report a new cGAL driver for the DVC interneuron. The ceh-63 promoter was chosen due to its restricted expression in the DVC neuron (Feng et al. 2012). The DVC interneuron driver construct containing the ceh-63 promoter (646 bp upstream of ATG translation start site) was injected into N2 and an integrated DVC driver line was generated. When crossed with the UAS-GFP effector strain (PS6843), the ceh-63 cGAL driver dictated GFP expression in the single DVC neuron (Figure 1), in addition to GFP in the coelomocyte from Punc-122::gfp co-injection marker. We did not observe GFP expression in uterus as reported by Feng et al., 2012.

Rauthan, M. et al. (2017) International Worm Meeting "MicroRNA regulation of nicotine-dependent behavior in C. elegans."

Rauthan, M., Ronan, E.A., Gong, J., Xu, X.Z.S., Liu, J., Wescott, S.

[International Worm Meeting, 2017]

Tobacco smoking is the leading cause of preventable death in developed nations. Nicotine is the principle addictive substance in cigarettes. Chronic exposure to nicotine up-regulates nAChRs and is thought to play a critical role in the primary steps of nicotine dependence, but the underlying mechanisms are not well understood. We have previously developed a C. elegans model of nicotine dependent behavior, and shown that the nAChR gene acr-15 is required for acute response to nicotine (Feng et al., 2006). Here we identify a key role for microRNA in regulating nicotine-dependent behavior by modulating nAChR expression in C. elegans. Specifically, we show that chronic nicotine treatment down-regulates microRNA machinery, leading to up-regulation of another nAChR gene that is specifically required for nicotine withdrawal behavior. This effect is mediated by a microRNA that recognizes the 3'UTR of nAChR transcripts. These observations uncover an interesting phenomenon that different nAChRs mediate distinct aspects of nicotine dependence in C. elegans. Our results reveal a functional link between nicotine, microRNA, nAChRs, and nicotine-dependent behavior. Reference(s): 1. Feng, Z., Li, W., Ward, A., Piggott, B.J., Larkspur, E., Sternberg, P.W., and Xu, X.Z.S. (2006). A C. elegans model of nicotine-dependent behavior: regulation by TRP family channels. Cell 127, 621-633.

Wang X et al. (2021) STAR Protoc "Invivo imaging of a PVD neuron in Caenorhabditis elegans."

Zhong R, Feng Z, Li T, Wang X, Yang X, Hu J, Zou Y, Nie W

[STAR Protoc, 2021]

The nematode <i>Caenorhabditis elegans</i> nociceptive PVD neurons have highly ordered dendritic branches, making this an ideal model to study the development and organization of dendrites. A <i>ser-2</i>-promoter-driven GFP reporter line <i>wyIs592</i>[<i>ser-2prom-3p::myr-</i>GFP] provides a comprehensive visualization of PVD anatomy. Here, we describe the detailed procedures for imaging a PVD neuron using <i>wyIs592</i> at late L4 larval stage <i>in vivo</i> by confocal microscopy. This protocol can also be applied to imaging other cells in <i>C.elegans</i>. For complete details on the use and execution of this protocol, please refer to Feng etal. (2020).

Liu J et al. (2001) Cell "Defective interplay of activators and repressors with TFIH in xeroderma pigmentosum."

Wang XW, Chuikov S, Reinberg D, Liu J, Ge H, Conaway JW, Weber A, Libutti D, Akoulitchev S, Levens D, Harris CC, Conaway RC

[Cell, 2001]

Inherited mutations of the TFIIH helicase subunits xeroderma pigmentosum (XP) B or XPD yield overlapping DNA repair and transcription syndromes. The high risk of cancer in these patients is not fully explained by the repair defect. The transcription defect is subtle and has proven more difficult to evaluate. Here, XPB and XPD mutations are shown to block transcription activation by the FUSE Binding Protein (FBP), a regulator of c-myc expression, and repression by the FBP Interacting Repressor (FIR). Through TFIIH, FBP facilitates transcription until promoter escape, whereas after initiation, FIR uses TFIIH to delay promoter escape. Mutations in TFIIH that impair regulation by FBP and FIR affect proper regulation of c-myc expression and have implications in the development of malignancy.

Arico JK et al. (2011) PLoS Genet "Epigenetic patterns maintained in early Caenorhabditis elegans embryos can be established by gene activity in the parental germ cells."

van der Vlag J, Arico JK, Kelly WG, Katz DJ

[PLoS Genet, 2011]

Epigenetic information, such as parental imprints, can be transmitted with genetic information from parent to offspring through the germ line. Recent reports show that histone modifications can be transmitted through sperm as a component of this information transfer. How the information that is transferred is established in the parent and maintained in the offspring is poorly understood. We previously described a form of imprinted X inactivation in Caenorhabditis elegans where dimethylation on histone 3 at lysine 4 (H3K4me2), a mark of active chromatin, is excluded from the paternal X chromosome (Xp) during spermatogenesis and persists through early cell divisions in the embryo. Based on the observation that the Xp (unlike the maternal X or any autosome) is largely transcriptionally inactive in the paternal germ line, we hypothesized that transcriptional activity in the parent germ line may influence epigenetic information inherited by and maintained in the embryo. We report that chromatin modifications and histone variant patterns assembled in the germ line can be retained in mature gametes. Furthermore, despite extensive chromatin remodeling events at fertilization, the modification patterns arriving with the gametes are largely retained in the early embryo. Using transgenes, we observe that expression in the parental germline correlates with differential chromatin assembly that is replicated and maintained in the early embryo. Expression in the adult germ cells also correlates with more robust expression in the somatic lineages of the offspring. These results suggest that differential expression in the parental germ lines may provide a potential mechanism for the establishment of parent-of-origin epigenomic content. This content can be maintained and may heritably affect gene expression in the offspring.

L. Stergiou et al. (2006) European Worm Meeting "Gene Requirements for UV-Induced Apoptosis"

M.O. Hengartner, K. Doukoumetzidis, L. Stergiou, A. Sendoel

[European Worm Meeting, 2006]

L. Stergiou1, K. Doukoumetzidis1,2, A. Sendoel1 and M.O. Hengartner1 Activation of checkpoints following DNA damage is important to ensure efficient repair processes. The physiological importance of these response pathways is illustrated by a number of genetic disorders, such as ataxia telangiectasia (A-T) or xeroderma pigmentosum (XP), which result in multiple pathological features including predisposition to cancer. We studied the effects on the C. elegans germ line of ultraviolet light, which causes a pattern of DNA damage distinct from the one observed following ionizing radiation. We identified a signaling pathway that senses UV-induced damage, and conveys this information to the apoptotic machinery. Recognition of UV lesions by the nucleotide excision repair (NER) machinery is required not only for DNA repair, but also to activate downstream signaling pathways that lead to apoptosis. We will also present our results about the interplay between the NER pathway and other DNA damage response pathways that participate in the UV response in C. elegans.