Wallace, Sean W et al. (2011) International Worm Meeting "Glial regulation of chemosensation."

Shaham, Shai, Wallace, Sean W

[International Worm Meeting, 2011]

Glia play active roles in regulating nervous system development and function; however, little is known about the regulation of neuronal sensory transduction by glia at sensory organs. Ablation of glia in the C. elegans amphid results in functional defects in the associated sensory neurons, suggesting glia-derived factors regulate sensory neuron function (Bacaj et al., 2008). We aim to identify and characterize the functions of such factors. Previous work from our lab showed that the amphid sheath glial gene fig-1 is required for amphid sensory neurons to take up lipophilic but not water-soluble dyes from the extracellular media, raising the possibility that fig-1 may alter neuronal membrane properties. Furthermore, fig-1 mutants display defects in 1-Octanol avoidance behavior. fig-1 is predicted to encode a secreted protein containing two thrombospondin domains. Glial proteins with similar domains play roles in synapse establishment in vertebrates. To understand how FIG-1 protein functions, we plan to use genetic screens to determine how FIG-1 secretion from glia is regulated, and to identify neuronal proteins that may be targets of its activity. To identify additional glial regulators of sensory neuron function, 298 amphid sheath glial genes previously identified in our lab will be screened by RNA interference (RNAi) for defects in chemotaxis and avoidance behaviors. Sensory function is critical for animal survival, and loss of sensory function results in decreased quality of life in humans. The presence of glia at sensory structures is a conserved feature of animal evolution, yet their contribution to sensory function is poorly studied. Furthermore, the receptive endings of chemosensory neurons resemble synapses in that they contain dendrites that respond to similar chemicals and that are intimately associated with glia. Thus, our studies are likely to inform us about the roles glia play at sensory structures, and have the potential to provide general insights into the synaptic functions of glia.

Wallace, Sean et al. (2021) International Worm Meeting "Nuclear hormone receptors mediate adaptive responses to the mold Penicillium brevicompactum"

Shaham, Shai, Wallace, Sean

[International Worm Meeting, 2021]

Xenobiotic metabolizing enzymes (XMEs) play a major role in detoxification. Previous studies of XMEs in C. elegans have primarily revealed a role in the gut. By carrying out transcriptome analysis of amphid sheath (AMsh) glial cells, we have found many detoxifying enzymes to be expressed in glia, including cytochrome P450 oxygenases (CYPs), UDP-glucoronosyltransferases (UGTs) and glutathione-S-transferases (GSTs). Intriguingly, high expression levels of XMEs are also observed in the sustentacular glial cells of the mammalian olfactory epithelium, the analogous cell-type to AMsh glia, but the functions of these enzymes in olfactory tissues are poorly characterized. We have found that expression of the cytochrome P450 gene cyp-33C2, in both glia and gut, is induced by exposure to Penicillium brevicompactum, a toxin-producing mold. Penicillium species are common laboratory contaminants, and they are also found naturally in environments that wild isolates of C. elegans inhabit, yet there have been surprisingly few studies exploring the interactions between C. elegans and Penicillium species. We have found that the nuclear hormone receptors nhr-45 and nhr-156 are required cell-autonomously for mold-dependent induction of cyp-33C2. nhr-45 has been previously implicated in mitochondrial stress induced by the pathogen Pseudomonas aeruginosa. We have found that the mold P. brevicompactum causes mitochondrial stress in the gut and that this is greatly enhanced in nhr-45 mutant animals that are unable to induce expression of detoxifying enzymes. This elevated mitochondrial stress is accompanied by severe developmental defects, which can be rescued by restoring nhr-45 expression in the gut. These findings suggest that nuclear hormone receptor-dependent expression of detoxifying enzymes in the gut is required to clear mold toxins. We are currently exploring similar mechanisms in AMsh glia. The nuclear hormone receptor family has undergone massive expansion and diversification in the C. elegans genome. It has been proposed that this expansion allows NHRs to act as receptors for varying environmental cues. Intriguingly, naturally occurring polymorphisms in the nhr-156 gene in wild isolates of C. elegans, including the Hawaiian isolate CB4856, result in a reduced transcriptional response in AMsh glia upon mold exposure. Remarkably, using CRISPR to substitute the nhr-156 locus from CB4856 in to the N2 background is sufficient to generate the observed phenotypic diversity. While the implications of these findings are still under investigation, they support the idea that diversification of nuclear hormone receptor sequences in C. elegans allows for adaptations to environmental conditions.

Wallace, Sean et al. (2015) International Worm Meeting "The transcription factor pros-1 is expressed in glia and regulates the morphology and function of sensory neurons."

Lu, Yun, Shaham, Shai, Wallace, Sean

[International Worm Meeting, 2015]

Glia are found associated with neurons throughout animal nervous systems, and interactions between glia and neurons are critical for proper neuronal function. We are using the amphid sense organ as a model to study how glia regulate neuronal morphology and function. We previously showed that ablation of amphid sheath (AMsh) glia results in structural defects in sensory neuron receptive endings and corresponding behavioral defects in chemosensation. To investigate the nature of the glial-neuronal interactions underlying these defects, we carried out a post-embryonic RNAi screen of candidate glial genes, and found that downregulation of the prospero-related transcription factor pros-1 results in defects similar to those observed with AMsh glia ablation, including loss of the glial-dependent amphid channel, through which ciliated sensory neurons are exposed to the outside environment, and defective AWC wing-like cilia morphology. The Nemo-like kinase LIT-1, which normally promotes amphid channel expansion and is localized to the channel surface is mislocalized in pros-1 mutants, explaining, in part, the channel defects, and supporting a role for pros-1 in controlling expression of proteins that promote LIT-1 channel localization. pros-1 is expressed in AMsh glia, but not in the associated sensory neurons. We have used cell-specific RNAi and mosaic rescue experiments to show that pros-1 acts in AMsh glia to control these aspects of neuronal function. Prospero is a conserved homeodomain transcription factor with well characterized roles in cell fate determination during Drosophila nervous system development. Importantly, the defects we observe after pros-1 post-embryonic RNAi are not a consequence of changes in glial cell fate or morphology, allowing us to uncover novel functions for this transcription factor in differentiated glia. To identify transcriptional targets of pros-1, we isolated AMsh glia from larvae by cell dissociation and fluorescence associated cell sorting (FACS) and carried out RNAseq analysis of gene expression. Using this approach we identified many genes expressed in AMsh glia, a subset of which depend on pros-1 for their expression. We are currently testing the role of these candidate pros-1 targets with secondary screens. Homologs of pros-1 are expressed in glia in a number of animals including mammals. The transcriptional network we have uncovered downstream of pros-1 in C. elegans has allowed us to further characterize the molecular nature of the conserved interactions between glia and neurons that are essential for nervous system function.

Prasad BC et al. (1997) International C. elegans Meeting "THE C. ELEGANS unc-3 GENE ENCODES A HOMOLOGUE OF THE O/E FAMILY OF MAMMALIAN TRANSCRIPTION FACTORS"

Prasad BC, Reed RR, Seydoux GC

[International C. elegans Meeting, 1997]

The expression of specialized signal transduction components in mammalian olfactory neurons is thought to be regulated by the O/E (Olf-1/EBF) family of transcription factors. The O/E proteins are expressed in cells of the olfactory neuronal lineage throughout development, and are also expressed transiently in neurons in the developing nervous system during embryogenesis. We have identified a C. elegans homologue of the mammalian O/E proteins, which displays greater than 80% similarity over 350 amino acids. The CeO/E cDNA maps to cosmid F42D1, previously shown to encode unc-3(1). We demonstrate that CeO/E is the product of the unc-3 gene, mutations in which cause defects in the axonal outgrowth of motor neurons as well as defects in dauer formation, a process requiring chemosensory input. Like its mammalian homologues, CeO/E is expressed in certain chemosensory neurons (ASI amphid neurons) throughout development, and is also expressed transiently in developing motor neurons when these cells undergo axonal outgrowth. Currently, we are defining the DNA sequence binding specificity of the CeO/E protein. These observations suggest that the O/E family of transcription factors play a central and evolutionarily conserved role in the expression of proteins essential for axonal pathfinding and neuronal differentiation. (1) Sean Eddy, WBG 12(5):86 (February 1, 1993)

Jon Pierce-Shimomura et al. (2004) West Coast Worm Meeting "Swimming as a distinct form of locomotion in C. elegans: Analysis of the switch between crawling and swimming"

Steve McIntire, Jon Pierce-Shimomura

[West Coast Worm Meeting, 2004]

C. elegans crawls on a flat surface and swims in liquid. Both forms of locomotion are initiated as a series of alternating dorsal-ventral head bends. The DV bends propagate backward along the body to generate forward thrust. The two behaviors are distinct in that swimming bends are deeper and more rapid than crawling bends (Crawl ~0.5 Hz, Swim ~2.0 Hz). These differences underlie the characteristic body waveforms for each behavior. When the head is bent maximally to the ventral or dorsal side, the body forms a 'C' shape when swimming, and an 'S' shape when crawling. The motion of C. elegans in liquid has previously been called 'thrashing,' implying that the worm flails its body in an uncontrolled fashion. We find, however, that C. elegans makes efficient progress when swimming over a surface to orient up an attractive salt gradient. Therefore, for C. elegans , as for other nematode species (Wallace, 1958 Ann Applied Bio ), movement through liquid can be a goal-oriented form of locomotion that may be better described as 'swimming'. Swimming may have important adaptive value for C. elegans in its natural environment of damp soil where it must traverse through pockets of water. C. elegans rapidly switches between crawl and swim kinematics when it enters and exits a puddle. This switch is reminiscent of how other animals switch between distinct forms of locomotion called gaits. In some animals, gaits are known to be controlled by neural networks that are distinct in either the sense that separate networks control each gait, or a single locomotion network operates in different 'gait' states. We wondered whether distinct neural networks may also control crawling and swimming in C. elegans . To better understand swimming behavior and the switch from crawling to swimming, we screened for mutants that are normal for crawling, but abnormal for the initiation or maintenance of swimming, what we term 'Swa' ( Sw imming a bnormal). We isolated over 40 swa mutants and divided them into different classes. Interestingly, all of these mutants exhibit normal crawling. Four Swa classes related to the maintenance of swimming included those with smaller amplitude of head bends, irregular swim pattern, extra left-right bends, and frequent stopping. Two Swa classes related to the initiation and maintenance of swimming included (1) those that in liquid exhibit persistent crawling, and (2) those that exhibit delayed initiation together with frequent stopping. This second class included several mutants with defective touch responses suggesting importance of mechanosensory input in swimming. Analysis of mechanosensory mutants revealed that specific loss of function of the six main mechanosensory neurons (e.g. mec-3 and mec-4 ) results in only a mild irregular swim pattern. Mechanosensory mutants with defective mechanosensory and chemosensory function ( mec-1 , mec-2 and mec-8 ), however, exhibit delayed swim initiation, severely irregular swim pattern, and frequent stopping. Analysis of chemosensory mutants revealed that loss of function in taste (e.g. che-1 ), but not olfaction (e.g. odr-1 odr-7 ), produced a similar severe swim phenotype as observed in mec-2 mutants. Chemosensory input may be particularly important for the initiation and maintenance of swimming. Our findings demonstrate that crawling and swimming are distinct forms of locomotion for C. elegans . Moreover, both mechanosensory and chemosensory pathways appear to control the initiation and maintenance of swimming. Analysis of the switch between crawling and swimming in C. elegans may help to uncover the general neurogenetic mechanisms that underlie the control of patterned neural output.