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Programmed cell death is a common cell fate in most if not all multicellular organisms. Apoptosis, which will be used as a synonym for programmed cell death throughout this chapter, occurs extensively during development as well as during later life. The development of the nematode worm Caenorhabditis elegans provides a good example of the extensive use of programmed cell death.
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[
WormBook,
2007]
The intestine is one of the major organs in C. elegans and is largely responsible for food digestion and assimilation as well as the synthesis and storage of macromolecules. In addition, the intestine is emerging as a powerful experimental system in which to study such universal biological phenomena as vesicular trafficking, biochemical clocks, stress responses and aging. The present chapter describes some of these many and varied properties of the C. elegans intestine: the embryonic cell lineage, intestine morphogenesis, structure and physiology of the intestinal cell and, finally, the transcription factor network controlling intestine development and function.
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[
WormBook,
2005]
Nematodes are the most abundant type of animal on earth, and live in hot springs, polar ice, soil, fresh and salt water, and as parasites of plants, vertebrates, insects, and other nematodes. This extraordinary ability to adapt, which hints at an underlying genetic plasticity, has long fascinated biologists. The fully sequenced genomes of Caenorhabditis elegans and Caenorhabditis briggsae, and ongoing sequencing projects for eight other nematodes, provide an exciting opportunity to investigate the genomic changes that have enabled nematodes to invade many different habitats. Analyses of the C. elegans and C. briggsae genomes suggest that these include major changes in gene content; as well as in chromosome number, structure and size. Here I discuss how the data set of ten genomes will be ideal for tackling questions about nematode evolution, as well as questions relevant to all eukaryotes.
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[
1990]
The free-living nematode Caenorhabditis elegans is a small and unpretentious organism. It may thrive unnoticed in the cabbage patch in your backyard or the flower pot on your balcony. In their natural habitat soil nematodes live in a thin film of water. In the laboratory C. elegans dwells on Petri dishes in the liquid film on the top of an agar layer, but can also be grown in liquid culture. As in other nematodes the liquid-filled body cavity (pseudocoelom) functions as a hydroskeleton. When the worm dries out, the hydroskeleton collapses and the animal inevitably dies. In a loose sense C. elegans may therefore be considered as a kind of aquatic animal. Because of this and because C. elegans is particularly well suited to the study of certain aspects of development, the following chapter is included in this book on Experimental Embryology of Aquatic Organisms. The intention of this contribution is to serve as an introduction and as a reference source rather than as a complete summary of present knowledge in the field. As indicated by the title, the review will focus on embryonic cell lineages, pattern formation in the embryo and the analysis of mutants affecting early
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[
Methods Cell Biol,
1995]
Genetic balancers are genetic constructs or chromosomal rearrangements that allow lethal or sterile mutations to be stably maintained in heterozygotes. In this chapter we use the term balancer primarily to refer to chromosomal duplications or rearrangements that suppress crossing over. In addition, we define lethal as any mutation that blocks survival or reproduction. Phenotypes associated with lethal mutations in Caenorhabditis elegans range from egg or larval lethality to adult sterility and maternal effect lethality, and can include conditional effects such as temperature sensitivity. The number of essential genes in C. elegans (those identified by lethal mutations) may range as high as 7000 according to genetic estimates. Thus, lethal mutations constitute a rich source of information about basic biological processes in this nematode.
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[
Methods Mol Biol,
2012]
Cellular effects of primary mitochondrial dysfunction, as well as potential mitochondrial disease therapies, can be modeled in living animals such as the microscopic nematode, Caenorhabditis elegans. In particular, molecular analyses can provide substantial insight into the mechanism by which genetic and/or pharmacologic manipulations alter mitochondrial function. The relative expression of individual genes across both nuclear and mitochondrial genomes, as well as relative quantitation of mitochondrial DNA content, can be readily performed by quantitative real-time PCR (qRT-PCR) analysis of C. elegans. Additionally, microarray expression profiling offers a powerful tool by which to survey the global genetic consequences of various causes of primary mitochondrial dysfunction and potential therapeutic interventions at both the single gene and integrated pathway level. Here, we describe detailed protocols for RNA and DNA isolation from whole animal populations in C. elegans, qRT-PCR analysis of both nuclear and mitochondrial genes, and global nuclear genome expression profiling using the Affymetrix GeneChip C. elegans Genome Array.
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[
WormBook,
2005]
The regulation of transcription in C. elegans shares many similarities to transcription in other organisms. The details of how specific transcription factors bind to target promoters and act as either activators or repressors are still being examined in many cases, but an increasing number of factors and their binding sites are being characterized. This chapter reviews the general concepts that have emerged with regards to promoter function in C. elegans. Included are the methods that have been successfully employed as well as limitations encountered to date. Specific cis-acting promoter elements from
myo-2 ,
hlh-1 and
lin-26 are discussed as examples of complex promoters regulated by multiple sequence elements. In addition, examples of organ-, tissue-, and cell type-specific mechanisms for generating spatial specificity in gene expression are discussed.
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[
Methods Cell Biol,
2012]
In Caenorhabdatis elegans as in other animals, fat regulation reflects the outcome of behavioral, physiological, and metabolic processes. The amenability of C. elegans to experimentation has led to utilization of this organism for elucidating the complex homeostatic mechanisms that underlie energy balance in intact organisms. The optical advantages of C. elegans further offer the possibility of studying cell biological mechanisms of fat uptake, transport, storage, and utilization, perhaps in real time. Here, we discuss the rationale as well as advantages and potential pitfalls of methods used thus far to study metabolism and fat regulation, specifically triglyceride metabolism, in C. elegans. We provide detailed methods for visualization of fat depots in fixed animals using histochemical stains and in live animals by vital dyes. Protocols are provided and discussed for chloroform-based extraction of total lipids from C. elegans homogenates used to assess total triglyceride or phospholipid content by methods such as thin-layer chromatography or used to obtain fatty acid profiles by methods such as gas chromatography/mass spectrometry. Additionally, protocols are provided for the determination of rates of intestinal fatty acid uptake and fatty acid breakdown by -oxidation. Finally, we discuss methods for determining rates of de novo fat synthesis and Raman scattering approaches that have recently been employed to investigate C. elegans lipids without reliance on invasive techniques. As the C. elegans fat field is relatively new, we anticipate that the indicated methods will likely be improved upon and expanded as additional researchers enter this field.
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[
1990]
Nematodes have been used as biological models of aging for some twenty years, and a large number of reviews have appeared both as a chapter in the previous edition of this handbook and in other sources. Major advantages and disadvantages in the use of nematodes as model organisms have been well reviewed. It is clear that for some purposes, such as the identification of genetic variants in length of life, which will be reviewed here, nematodes are an invaluable model. Genetic variants of Caenorhabditis elegans have recently been isolated that have life span extensions of more than 70%; these strains offer an exceptional new avenue for the dissection of aging processes. With the exception of dietary restriction and selectively bred long-lived strains of Drosophila melanogaster, there are no other techniques for lengthening life, thereby allowing the study of associated changes in other physiological systems. This chapter will concentrate on C. elegans and will review the genetic techniques used to study againg as well as methodological advances in other areas of C. elegans genetics. The possibilities for the study of physiological alterations associated with aging through the use of such genetic variants are not yet being widely exploited, leaving open a wide variety of potential research areas.
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[
Methods Mol Biol,
2016]
The development of genomics and next generation sequencing platforms has dramatically improved our insight into chromatin structure and organization and its fine interplay with gene expression. The nuclear envelope has emerged as a key component in nuclear organization via extensive contacts between the genome and numerous proteins at the nuclear periphery. These contacts may have profound effects on gene expression as well as cell proliferation and differentiation. Indeed, their perturbations are associated with several human pathologies known as laminopathies or nuclear envelopathies. However, due to their dynamic behavior the contacts between nuclear envelope proteins and chromatin are challenging to identify, in particular in intact tissues. Here, we propose the DamID technique as an attractive method to globally characterize chromatin organization in the popular model organism Caenorhabditis elegans. DamID is based on the in vivo expression of a chromatin-associated protein of interest fused to the Escherichia coli DNA adenine methyltransferase, which produces unique identification tags at binding site in the genome. This marking is simple, highly specific and can be mapped by sensitive enzymatic and next generation sequencing approaches.