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Neurons and epithelia are viewed as fundamentally different cell types, yet some sensory neurons exhibit hallmarks of epithelial cells. For example, they use tight junctions to form a diffusion barrier continuous with the skin or other epithelia and they exhibit bona fide apical-basal polarity, with an outward-facing apical surface that is biochemically and functionally distinct from their inward-facing basolateral surface. Yet they are unmistakeably neurons with axon-dendrite polarity. Examples include olfactory receptor neurons and photoreceptors. In this review, I highlight how viewing these neurons as specialized epithelial cells informs our understanding of their development and raises intriguing questions about the establishment of apical-basal and axon-dendrite polarity.

Glia shape the development and function of the C. elegans nervous system, especially its sense organs and central neuropil (nerve ring). Cell-type-specific promoters allow investigators to label or manipulate individual glial cell types, and therefore provide a key tool for deciphering glial function. In this technical resource, we compare the specificity, brightness, and consistency of cell-type-specific promoters for C. elegans glia. We identify a set of promoters for the study of seven glial cell types (F16F9.3, amphid and phasmid sheath glia; F11C7.2, amphid sheath glia only; grl-2, amphid and phasmid socket glia; hlh-17, cephalic (CEP) sheath glia; and grl-18, inner labial (IL) socket glia) as well as a pan-glial promoter (mir-228). We compare these promoters to promoters that are expressed more variably in combinations of glial cell types (delm-1 and itx-1). We note that the expression of some promoters depends on external conditions or the internal state of the organism, such as developmental stage, suggesting glial plasticity. Finally, we demonstrate an approach for prospectively identifying cell-type-specific glial promoters using existing single-cell sequencing data, and we use this approach to identify two novel promoters specific to IL socket glia (col-53 and col-177).

The extracellular matrix (ECM) guides and constrains the shape of the nervous system. InC. elegans, DIG-1 is a giant ECM component that is required for fasciculation of sensory dendrites during development and for maintenance of axon positions throughout life. We identified four novel alleles ofdig-1in three independent screens for mutants affecting disparate aspects of neuronal and glial morphogenesis. First, we find that disruption of DIG-1 causes fragmentation of the amphid sheath glial cell in larvae and young adults. Second, it causes severing of the BAG sensory dendrite from its terminus at the nose tip, apparently due to breakage of the dendrite as animals reach adulthood. Third, it causes embryonic defects in dendrite fasciculation in inner labial (IL2) sensory neurons, as previously reported, as well as rare defects in IL2 dendrite extension that are enhanced by loss of the apical ECM component DYF-7, suggesting that apical and basolateral ECM contribute separately to dendrite extension. Our results highlight novel roles for DIG-1 in maintaining the cellular integrity of neurons and glia, possibly by creating a barrier between structures in the nervous system.

During convergent differentiation, multiple developmental lineages produce a highly similar or identical cell type. However, few molecular players that drive convergent differentiation are known. Here, we show that the C. elegans Forkhead transcription factor UNC-130 is required in only one of three convergent lineages that produce the same glial cell type. UNC-130 acts transiently as a repressor in progenitors and newly-born terminal cells to allow the proper specification of cells related by lineage rather than by cell type or function. Specification defects correlate with UNC-130:DNA binding, and UNC-130 can be functionally replaced by its human homolog, the neural crest lineage determinant FoxD3. We propose that, in contrast to terminal selectors that activate cell-type specific transcriptional programs in terminally differentiating cells, UNC-130 acts early and specifically in one convergent lineage to produce a cell type that also arises from molecularly distinct progenitors in other lineages.

Neuronal activity often leads to alterations in gene expression and cellular architecture. The nematode Caenorhabditis elegans, owing to its compact translucent nervous system, is a powerful system in which to study conserved aspects of the development and plasticity of neuronal morphology. Here we focus on one pair of sensory neurons, termed URX, which the worm uses to sense and avoid high levels of environmental oxygen. Previous studies have reported that the URX neuron pair has variable branched endings at its dendritic sensory tip. By controlling oxygen levels and analyzing mutants, we found that these microtubule-rich branched endings grow over time as a consequence of neuronal activity in adulthood. We also find that the growth of these branches correlates with an increase in cellular sensitivity to particular ranges of oxygen that is observable in the behavior of older worms. Given the strengths of C. elegans as a model organism, URX may serve as a potent system for uncovering genes and mechanisms involved in activity-dependent morphological changes in neurons and possible adaptive changes in the aging nervous system.

To sense the outside world, some neurons protrude across epithelia, the cellular barriers that line every surface of our bodies. To study the morphogenesis of such neurons, we examined theC. elegansamphid, in which dendrites protrude through a glial channel at the nose. During development, amphid dendrites extend by attaching to the nose via DYF-7, a type of protein typically found in epithelial apical ECM. Here, we show that amphid neurons and glia exhibit epithelial properties, including tight junctions and apical-basal polarity, and develop in a manner resembling other epithelia. We find that DYF-7 is a fibril-forming apical ECM component that prevents rupture of the tube-shaped glial channel, reminiscent of roles for apical ECM in other narrow epithelial tubes. We also identify a role for FRM-2, a homolog of EPBL15/moe/Yurt which promote epithelial integrity in other systems. Finally, we show that other environmentally-exposed neurons share a requirement for DYF-7. Together, our results suggest that these neurons and glia can be viewed as part of an epithelium continuous with the skin, and are shaped by mechanisms shared with other epithelia.

Neurons develop elaborate morphologies that provide a model for understanding cellular architecture. By studyingC. eleganssensory dendrites, we previously identified genes that act to promote the extension of ciliated sensory dendrites during embryogenesis. Interestingly, the nonciliated dendrite of the oxygen-sensing neuron URX is not affected by these genes, suggesting it develops through a distinct mechanism. Here, we use a visual forward genetic screen to identify mutants that affect URX dendrite morphogenesis. We find that disruption of the MAP kinase MAPK-15 or the βH-spectrin SMA-1 causes a phenotype opposite to what we had seen before: dendrites extend normally during embryogenesis but begin to overgrow as the animals reach adulthood, ultimately extending up to 150% of their normal length. SMA-1 is broadly expressed and acts non-cell-autonomously, while MAPK-15 is expressed in many sensory neurons including URX and acts cell-autonomously. MAPK-15 acts at the time of overgrowth, localizes at the dendrite ending, and requires its kinase activity, suggesting it acts locally in time and space to constrain dendrite growth. Finally, we find that the oxygen-sensing guanylate cyclase GCY-35, which normally localizes at the dendrite ending, is localized throughout the overgrown region, and that overgrowth can be suppressed by overexpressing GCY-35 or by genetically mimicking elevated cGMP signaling. These results suggest that overgrowth may correspond to expansion of a sensory compartment at the dendrite ending, reminiscent of the remodeling of sensory cilia or dendritic spines. Thus, in contrast to established pathways that promote dendrite growth during early development, our results reveal a distinct mechanism that constrains dendrite growth throughout the life of the animal, possibly by controlling the size of a sensory compartment at the dendrite ending.AUTHOR SUMMARYLewis Carroll’s Alice told the Caterpillar, “Being so many different sizes in a day is very confusing.” Like Alice, the cells of our bodies face a problem in size control – they must become the right size and remain that way throughout the life of the organism. This problem is especially relevant for nerve cells (neurons), as the lengths of their elaborate dendrites determine the connections they can make. To learn how neurons control their size, we turned not to a Caterpillar but to a worm: the microscopic nematodeC. elegans, in which single neurons can be easily visualized and the length of each dendrite is highly predictable across individuals. We focused on a single dendrite, that of the oxygen-sensing neuron URX, and we identified two genes that control its length. When these genes are disrupted, the dendrite develops correctly in embryos but then, like Alice, grows too much, eventually extending up to 1.5x its normal length. Thus, in contrast to known pathways that promote the initial growth of a dendrite early in development, our results help to explain how a neuron maintains its dendrite at a consistent length throughout the animal’s life.

Coordination of neurite morphogenesis with surrounding tissues is crucial to the establishment of neural circuits, but the underlying cellular and molecular mechanisms remain poorly understood. We show that neurons in a C. elegans sensory organ, called the amphid, undergo a collective dendrite extension to initiate formation of the sensory nerve. The amphid neurons first assemble into a multicellular rosette. The vertex of the rosette, which becomes the dendrite tips, is attached to the anteriorly migrating epidermis and carried to the sensory depression, extruding the dendrites away from the neuronal cell bodies. Multiple adhesion molecules including DYF-7, SAX-7, HMR-1 and DLG-1 function redundantly in rosette-to-epidermis attachment. PAR-6 is localized to the rosette vertex and dendrite tips, and promotes DYF-7 localization and dendrite extension. Our results suggest a collective mechanism of neurite extension that is distinct from the classical pioneer-follower model and highlight the role of mechanical cues from surrounding tissues in shaping neurites.

Glia constitute a major, understudied population of cells in the nervous system. Currently, it is appreciated that these cells exhibit vast morphological, functional, and molecular diversity, but our understanding of glial biology is limited. Some key unanswered questions include how glial diversity is generated during development and what functions distinct glial subtypes serve in the mature nervous system. The nematode Caenorhabditis elegans contains a defined set of glia, which have clear morphological and molecular differences, and thus provides a simplified model for understanding glial diversity. In addition, recent experiments suggest that the molecular mechanisms underlying the generation of glial diversity in C. elegans are conserved with those in mammals. In this review, we summarize the surprising diversity of glial subtypes present in this simple organism, and highlight current thinking about what roles they perform in the nervous system. We emphasize how genetic approaches may be used to identify the mechanistic origins of glial diversity, which is key to understanding how glia function in health and disease. WIREs Dev Biol 2015, 4:151–160. doi: 10.1002/wdev.171 For further resources related to this article, please visit the WIREs website. Conflict of interest: The authors have declared no conflicts of interest for this article.

Primary cilia are ubiquitous sensory organelles that mediate diverse signaling pathways. Cilia position on the cell surface is determined by the location of the basal body (BB) that templates the cilium. The mechanisms that regulate BB positioning in the context of ciliogenesis are largely unknown. Here we show that the conserved signaling and scaffolding protein Girdin localizes to the proximal regions of centrioles and regulates BB positioning and ciliogenesis in Caenorhabditis elegans sensory neurons and human RPE-1 cells. Girdin depletion alters localization of the intercentriolar linker and ciliary rootlet component rootletin, and rootletin knockdown in RPE-1 cells mimics Girdin-dependent phenotypes. C. elegans Girdin also regulates localization of the apical junction component AJM-1, suggesting that in nematodes Girdin may position BBs via rootletin- and AJM-1-dependent anchoring to the cytoskeleton and plasma membrane, respectively. Together, our results describe a conserved role for Girdin in BB positioning and ciliogenesis.

A dendrite grows by sprouting filopodia, some of which mature into stable dendrite branches that bear synapses and sprout filopodia of their own. Recent work has shown that a filopodium begins deciding to become a stable branch within one minute of contacting a presynaptic partner, but what triggers this decision remains unknown. We consider the evidence for three possible triggers: activity of neurotransmitter receptors, signaling through adhesion proteins, and heightened membrane tension as the filopodium attempts to retract but is held in place by adhesive contacts with the target. Of these, membrane tension-induced signaling is especially appealing, as it would serve as a general reporter of attachment, independent of which specific adhesion molecules are used.

During development, biomechanical forces contour the body and provide shape to internal organs. Using genetic and molecular approaches in combination with a FRET-based tension sensor, we characterized a pulling force exerted by the elongating pharynx (foregut) on the anterior epidermis during C. elegans embryogenesis. Resistance of the epidermis to this force and to actomyosin-based circumferential constricting forces is mediated by FBN-1, a ZP domain protein related to vertebrate fibrillins. fbn-1 was required specifically within the epidermis and FBN-1 was expressed in epidermal cells and secreted to the apical surface as a putative component of the embryonic sheath. Tiling array studies indicated that fbn-1 mRNA processing requires the conserved alternative splicing factor MEC-8/RBPMS. The conserved SYM-3/FAM102A and SYM-4/WDR44 proteins, which are linked to protein trafficking, function as additional components of this network. Our studies demonstrate the importance of the apical extracellular matrix in preventing mechanical deformation of the epidermis during development. DOI: http://dx.doi.org/10.7554/eLife.06565.001, eLife digest For an animal embryo to develop, its cells must organize themselves into tissues and organs. For example, skin and the lining of internal organs—such as the lungs and gut—are made from cells called epithelial cells, which are tightly linked to form flat sheets. In a microscopic worm called Caenorhabditis elegans, the outermost layer of epithelial cells (called the epidermis) forms over the surface of the embryo early on in embryonic development. Shortly afterwards, the embryonic epidermis experiences powerful contractions along the surface of the embryo. The force generated by these contractions converts the embryo from an oval shape to a roughly cylindrical form. These contractions also squeeze the internal tissues and organs, which correspondingly elongate along with the epidermis. It has been known for decades that such ‘mechanical’ forces are important for the normal development of embryos. However, it remains poorly understood how these forces generate tissues and organs of the proper shape—partly because it is difficult to measure forces in living embryos. It is also not clear how the mechanical properties of specific tissues are controlled. Now, Kelley, Yochem, Krieg et al. have analyzed the development of C. elegans' embryos and discovered a novel mechanical interplay between the feeding organ (called the pharynx) and the worm's epidermis. The experiments involved studying several mutant worms that perturb epidermal contractions and disrupt the attachment of the pharynx to the epidermis. These studies suggested that the pharynx exerts a strong inward pulling force on the epidermis during development. Using recently developed methods, Kelley, Yochem, Krieg et al. then measured mechanical forces within intact worm embryos and demonstrated that greater forces were experienced in cells that were being pulled by the pharynx. Kelley, Yochem, Krieg et al. further analyzed how the epidermis normally resists this pulling force from the pharynx and implicated a protein called FBN-1. This worm protein is structurally related to a human protein that is affected in people with a disorder called Marfan Syndrome. Worm embryos without the FBN-1 protein become severely deformed because they are unable to withstand mechanical forces at the epidermis. FBN-1 is normally synthesized and then transported to the outside of the worm embryo by epidermal cells, where it is thought to assemble into a meshwork of long fibers. This provides a strong scaffold that attaches to the epidermis to prevent the epidermis from undergoing excessive deformation while it experiences mechanical forces. The work of Kelley, Yochem, Krieg et al. provides an opportunity to understand how FBN-1 and other fiber-forming proteins are produced and transported to the cell surface. Moreover, these findings may have implications for human diseases and birth defects that result from an inability of tissues to respond appropriately to mechanical forces. DOI: http://dx.doi.org/10.7554/eLife.06565.002

The ciliated receptive endings of sensory cells and the dendrites of other neurons are shaped by adhesive interactions, many of which depend on machinery also present in epithelia. Sensory cells are shaped by interactions with support cells through adhesion junctions via the Crumbs complex, tight junction components such as claudins, as well as interactions with apical extracellular matrix composed of zona pellucida domain proteins. Neuronal dendrites are shaped by adhesion machinery that includes cadherins, catenins, afadin, L1CAM, CHL1, Sidekicks, Contactin and Caspr, many of which are shared with epithelia. This review highlights this shared machinery, and suggests that mechanisms of epithelial morphogenesis may thus provide a guide to understanding dendrite morphogenesis.

Under mating conditions, yeast cells adopt a characteristic pear-shaped morphology, called a “shmoo,” as they project a cell extension toward their mating partners. Mating partners make contact at their shmoo tips, dissolve the intervening cell wall, and fuse their plasma membranes. We identified mutations in ERG4 , encoding the enzyme that catalyzes the last step of ergosterol biosynthesis, that impair both shmoo formation and cell fusion. Upon pheromone treatment, erg4 Δ mutants polarized growth, lipids, and proteins involved in mating but did not form properly shaped shmoos and fused with low efficiency. Supplementation with ergosterol partially suppressed the shmooing defect but not the cell fusion defect. By contrast, removal of the Erg4 substrate ergosta-5,7,22,24(28)-tetraenol, which accumulates in erg4 Δ mutant cells and contains an extra double bond in the aliphatic chain of the sterol, restored both shmooing and cell fusion to wild-type levels. Thus, a two-atom change in the aliphatic moiety of ergosterol is sufficient to obstruct cell shape remodeling and cell fusion.

The nematode Caenorhabditis elegans has a simple nervous system with glia restricted primarily to sensory organs. Some of the activities that would be provided by glia in the mammalian nervous system are either absent or provided by non-glial cell types in C. elegans, with only a select set of mammalian glial activities being similarly provided by specialized glial cells in this animal. These observations suggest that ancestral roles of glia may be to modulate neuronal morphology and neuronal sensitivity in sensory organs.

The molecular machines that mediate cell fusion are unknown. Previously, we identified a multispanning transmembrane protein, Prm1 (pheromone-regulated membrane protein 1), that acts during yeast mating (Heiman, M.G., and P. Walter. 2000. J. Cell Biol. 151:719–730). Without Prm1, a substantial fraction of mating pairs arrest with their plasma membranes tightly apposed yet unfused. In this study, we show that lack of the Golgi-resident protease Kex2 strongly enhances the cell fusion defect of Prm1-deficient mating pairs and causes a mild fusion defect in otherwise wild-type mating pairs. Lack of the Kex1 protease but not the Ste13 protease results in similar defects. Δkex2 and Δkex1 fusion defects were suppressed by osmotic support, a trait shared with mutants defective in cell wall remodeling. In contrast, other cell wall mutants do not enhance the Δprm1 fusion defect. Electron microscopy of Δkex2-derived mating pairs revealed novel extracellular blebs at presumptive sites of fusion. Kex2 and Kex1 may promote cell fusion by proteolytically processing substrates that act in parallel to Prm1 as an alternative fusion machine, as cell wall components, or both.

Cell fusion occurs throughout development, from fertilization to organogenesis. The molecular mechanisms driving plasma membrane fusion in these processes remain unknown. While yeast mating offers an excellent model system in which to study cell fusion, all genes previously shown to regulate the process act at or before cell wall breakdown; i.e., well before the two plasma membranes have come in contact. Using a new strategy in which genomic data is used to predict which genes may possess a given function, we identified PRM1, a gene that is selectively expressed during mating and that encodes a multispanning transmembrane protein. Prm1p localizes to sites of cell-cell contact where fusion occurs. In matings between Deltaprm1 mutants, a large fraction of cells initiate zygote formation and degrade the cell wall separating mating partners but then fail to fuse. Electron microscopic analysis reveals that the two plasma membranes in these mating pairs are tightly apposed, remaining separated only by a uniform gap of approximately 8 nm. Thus, the phenotype of Deltaprm1 mutants defines a new step in the mating reaction in which membranes are juxtaposed, possibly through a defined adherence junction, yet remain unfused. This phenotype suggests a role for Prm1p in plasma membrane fusion.

Neurons are the pedestals of human consciousness, yet from a cell biological view they are little more than specialized epithelia. Indeed, as highlighted in the “Neuronal Development and Degeneration” Minisymposium, neurons inhabit conventional cell biology at an extreme: in their diversity of cell types, the specificity of their cell–cell junctions and cell–cell signaling, their degree of asymmetric polarization, their rate and volume of membrane traffic, the size and stability of their cytoskeletal assemblies, and their metabolic activity and ion flux. This extreme lifestyle requires extreme upkeep and, although neurons can be extreme in their longevity, if any of the above processes are compromised then neurodegenerative diseases can ensue. This session highlighted what can be learned by studying cell biology at the extremes, beginning with neuronal development and ending in studies of neurodegenerative disease.

SummaryCells are devices whose structures delimit function. For example, in the nervous system, neuronal and glial shapes dictate paths of information flow. To understand how cells acquire their shapes, we examined the formation of a sense organ in C. elegans. Using time-lapse imaging, we found that sensory dendrites form by stationary anchoring of dendritic tips during cell-body migration. A genetic screen identified DEX-1 and DYF-7, extracellular proteins required for dendritic tip anchoring, which act cooperatively at the time and place of anchoring. DEX-1 and DYF-7 contain, respectively, zonadhesin and zona pellucida domains, and DYF-7 self-associates into multimers important for anchoring. Thus, unlike other dendrites, amphid dendritic tips are positioned by DEX-1 and DYF-7 without the need for long-range guidance cues. In sequence and function, DEX-1 and DYF-7 resemble tectorins, which anchor stereocilia in the inner ear, suggesting that a sensory dendrite anchor may have evolved into part of a mechanosensor.

Neurons develop elaborate morphologies that provide a model for understanding cellular architecture. By studying C. elegans sensory dendrites, we previously identified genes that act to promote the extension of ciliated sensory dendrites during embryogenesis. Interestingly, the nonciliated dendrite of the oxygen-sensing neuron URX is not affected by these genes, suggesting it develops through a distinct mechanism. Here, we use a visual forward genetic screen to identify mutants that affect URX dendrite morphogenesis. We find that disruption of the MAP kinase MAPK-15 or the βH-spectrin SMA-1 causes a phenotype opposite to what we had seen before: dendrites extend normally during embryogenesis but begin to overgrow as the animals reach adulthood, ultimately extending up to 150% of their normal length. SMA-1 is broadly expressed and acts non-cell-autonomously, while MAPK-15 is expressed in many sensory neurons including URX and acts cell-autonomously. MAPK-15 acts at the time of overgrowth, localizes at the dendrite ending, and requires its kinase activity, suggesting it acts locally in time and space to constrain dendrite growth. Finally, we find that the oxygen-sensing guanylate cyclase GCY-35, which normally localizes at the dendrite ending, is localized throughout the overgrown region, and that overgrowth can be suppressed by overexpressing GCY-35 or by genetically mimicking elevated cGMP signaling. These results suggest that overgrowth may correspond to expansion of a sensory compartment at the dendrite ending, reminiscent of the remodeling of sensory cilia or dendritic spines. Thus, in contrast to established pathways that promote dendrite growth during early development, our results reveal a distinct mechanism that constrains dendrite growth throughout the life of the animal, possibly by controlling the size of a sensory compartment at the dendrite ending. ispartof: Plos Genetics vol:14 issue:6 ispartof: location:United States status: published