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WormBook,
2013]
Cell isolation and culture are essential tools for the study of cell function. Isolated cells grown under controlled conditions can be manipulated and imaged at a level of resolution that is not possible in whole animals or even tissue explants. Recent advances have allowed for large-scale isolation and culture of primary C. elegans cells from both embryos and all four larval stages. Isolated cells can be used for single-cell profiling, electrophysiology, and high-resolution microscopy to assay cell autonomous development and behavior. This chapter describes protocols for the isolation and culture of C. elegans embryonic and larval stage cells. Our protocols describe isolation of embryonic and L1 stage cells from nematodes grown on high-density NA22 bacterial plates and isolation of L2 through L4 stage cells from nematodes grown in axenic liquid culture. Both embryonic and larval cells can be isolated from nematode populations within 3 hours and can be cultured for several days. A primer on sterile cell culture techniques is given in the appendices.
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FEBS J,
2021]
Glia make up roughly half of all cells in the mammalian nervous system and play a major part in nervous system development, function and disease. Although research in the past few decades have shed light on their morphological and functional diversity, there is still much to be known about key aspects of their development such as the generation of glial diversity and the factors governing proper morphogenesis. Glia of the nematode C. elegans possess many developmental and morphological similarities with their vertebrate counterparts and can potentially be used as a model to understand certain aspects of glial biology owing to advantages such as its genetic tractability and fully mapped cell lineage. In this review we summarize recent progress in our understanding of genetic pathways that regulate glial development in C. elegans and discuss how some of these findings may be conserved.
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Legouis R, Chang JT, O'Rourke EJ, Sato K, Lapierre LR, Kumsta C, Guo B, Zhang H, Wu F, Lin L, Melendez A, Kovacs AL, Hansen M, Lu Q, Jia K, Wang X, Sato M
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Autophagy,
2015]
The cellular recycling process of autophagy has been extensively characterized with standard assays in yeast and mammalian cell lines. In multicellular organisms, numerous external and internal factors differentially affect autophagy activity in specific cell types throughout the stages of organismal ontogeny, adding complexity to the analysis of autophagy in these metazoans. Here we summarize currently available assays for monitoring the autophagic process in the nematode C. elegans. A combination of measuring levels of the lipidated Atg8 ortholog LGG-1, degradation of well-characterized autophagic substrates such as germline P granule components and the SQSTM1/p62 ortholog SQST-1, expression of autophagic genes and electron microscopy analysis of autophagic structures are presently the most informative, yet steady-state, approaches available to assess autophagy levels in C. elegans. We also review how altered autophagy activity affects a variety of biological processes in C. elegans such as L1 survival under starvation conditions, dauer formation, aging, and cell death, as well as neuronal cell specification. Taken together, C. elegans is emerging as a powerful model organism to monitor autophagy while evaluating important physiological roles for autophagy in key developmental events as well as during adulthood.
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Dev Dyn,
2010]
Hemidesmosomes are evolutionarily conserved attachment complexes linked to intermediate filaments that connect epithelial cells to the extracellular matrix. They provide tissue integrity and resistance to mechanical forces. Alterations in hemidesmosome structures are responsible for skin blistering, carcinoma invasion, and wound-healing defects. Valuable information about hemidesmosome assembly and disassembly has been obtained from in vitro cell culture studies. However, how these processes take place in vivo still remains elusive. Here, we discuss recent data about the formation and reorganization of hemidesmosomes in several in vivo model systems, particularly zebrafish and Caenorhabditis elegans, focusing on various factors affecting their dynamics. Mechanisms found in different organisms reveal that hemidesmosome formation and maintenance in vivo are carefully controlled by ECM protein folding, ECM-receptor expression and trafficking, and by post-translational modification of hemidesmosome components. These findings validate and extend the in vitro studies, and shed light on our understanding about hemidesmosomes across species.
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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.
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Ageing Res Rev,
2003]
Caspases are a group of cysteine dependent aspartate-specific proteases. Originally found as the homolog of Ced-3 in C. elegans, 14 caspases have now been identified in mammals to date. Caspases play important roles in both the intrinsic and extrinsic apoptotic pathways and interact with the non-caspase apoptotic pathways. A number of recent published observations have suggested a strong association between apoptosis, age-related diseases and aging. Findings from our group and others reveal a strong correlation between alterations in caspase activity and aging. In this view point, we summarize current knowledge of the connection between caspases and aging observed in a variety of model systems from cultured cells in vitro to the in vivo models of rodents and humans.
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Mech Ageing Dev,
2002]
Ageing is accompanied by a general decline of physiological function, especially at later stages, and significant increases in the incidence of cancer and other degenerative diseases. It has recently been hypothesized that alterations in apoptosis may contribute to these age-associated changes. However, whether there is a role for apoptosis in the ageing process and how ageing may modify the regulatory machinery of apoptosis remains obscure. Although the literature addressing these issues is scarce, research in this area is gaining momentum. Molecules involved in apoptosis signaling in mammals have been found to regulate ageing in organisms such as Caenorhabditis elegans and Drosophila melanogaster. Caloric restriction studies in a wide variety of organisms, ranging from yeast to mammals, suggest the conserved nature of the ageing regulatory systems. It seems very likely that signals that regulate ageing will impact apoptosis and the extent of apoptosis may then impact ageing. However, to date, there has been no direct evidence supporting the existence of such cross-communication between ageing and apoptosis in mammalian system. Here we review progress in the field.
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Curr Opin Microbiol,
2008]
Individuals interact with environment through different neuronal functions, such as olfaction and mechanosensation; experience shapes these physiological functions. It is not well understood how an individual senses and processes multiple cues of natural stimuli in the environment and how experience modulates these physiological mechanisms. Recent molecular genetics and behavioral studies on the interactions of the genetic model organism Caenorhabditis elegans with pathogenic bacteria have provided insights on the molecular and cellular mechanisms underlying these regulatory processes.
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Cell Host Microbe,
2009]
Communication between the nervous and immune systems is fundamental to animal physiology. However, the complicated anatomy and signaling pathways of these systems in mammals challenge the understanding of the neural-immune interaction at molecular, cellular, and organismic levels. Caenorhabditis elegans has been valuable in this regard because of its simple, well-defined nervous system and accessibility to genetic, molecular, and behavioral analyses. Recent studies in C. elegans have identified neuronal pathways that regulate signaling cascades in innate immune responses, including a neuroendocrine network, a TGF-beta pathway and dopaminergic neurotransmission, illuminating how specific neuronal signaling molecules and circuits control integrative immune responses.
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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.