Cell Death in C. elegans Development

Jennifer Zuckerman Malin; Shai Shaham1    Laboratory of Developmental Genetics, The Rockefeller University, New York, USA
1 Corresponding author: email address: shaham@rockefeller.edu

1 Introduction

Cell death is a widespread process that is essential for life. Tissue sculpting, organ morphogenesis, and organ size control are but a few of the developmental events that integrally utilize programmed cell death to generate a functioning adult animal. It is therefore not surprising that many things go wrong when cell death goes awry (Fuchs & Steller, 2011). Indeed, neurodegeneration and tumorigenesis, disease states against which armies of researchers have been amassed, result from too much or too little cell culling, respectively (Hanahan & Weinberg, 2011; Youle & van der Bliek, 2012). While the hypothesis that cell death is a regulated phenomenon in animal development was first experimentally addressed in vertebrates (Hamburger & Levi-Montalcini, 1949) and insects (Lockshin & Williams, 1965), the first systematic studies aimed at deciphering the molecular program promoting cell demise employed the free-living soil nematode Caenorhabditis elegans (Horvitz, 2003). Early observations of the cellular complement of adult C. elegans revealed little variation in the number and position of cells between individuals of similar ages, leading to the proposal that cell lineage in this animal may be invariant. This prediction was largely borne out by taking advantage of the transparent cuticle of the animal to observe cell divisions in vivo (Kimble & Hirsh, 1979; Sulston, Albertson, & Thomson, 1980; Sulston & Horvitz, 1977; Sulston, Schierenberg, White, & Thomson, 1983). This heroic effort culminated in a complete cell lineage tree documenting a generally predictable pattern of divisions that generate adult somatic tissue from the zygote. These studies demonstrated that precisely 1090 and 1178 somatic cells must be generated to produce a C. elegans hermaphrodite and male, respectively.

Among the generated cells, a small but substantial set (~ 12%) are eliminated. These cells become refractile under differential interference contrast (DIC) optics (Fig. 1), acquire a rounded morphology, and eventually disappear. Ultrastructural studies reveal that these dying cells are engulfed by neighboring cells (Sulston et al., 1983), and possess characteristics of apoptotic cell death, such as condensed nuclear chromatin, and reduced cytoplasmic volume (Shaham & Horvitz, 1996b; Sulston et al., 1983) (Fig. 1). Like the lineage itself, these cell death events are essentially invariant between individuals and target the same cells at the same time in development. In the hermaphrodite and male, 131 and 147 somatic cells are eliminated, respectively. Subsequent studies demonstrated that cell death is highly prevalent during germline development and maintenance, with roughly 50% of female meiosis products succumbing to apoptosis (Gumienny, Lambie, Hartwieg, Horvitz, & Hengartner, 1999). Developmental death of germ cells in C. elegans differs from somatic cell death in that the identities of dying cells are not ascribed to their lineage (Gumienny et al., 1999; Sulston et al., 1983). C. elegans therefore offers two arenas for understanding cell death control: one in which cell death and lineage are tightly coupled, and one in which stochastic processes apparently determine life and death. Studies of the former revealed a core pathway controlling apoptotic cell death from C. elegans to mammals.

Here, we describe these key components and their interactions and explore current understanding of the lineage-dependent mechanisms that trigger the activation of these killer genes and proteins. We also discuss a group of genes important for the clearance of dying cells and their relation to cell death execution and delve into a number of mysteries that remain unanswered and which have the potential to expand and modify our understanding of why and how cells die. We end by describing a novel nonapoptotic C. elegans cell death program that promotes dismantling of the male-specific linker cell.

2 Core Apoptosis Regulators in C. elegans

Most cell death in C. elegans is controlled by the proteins CED-3, CED-4, CED-9, and EGL-1, whose functions and interactions have been worked out in some detail (Fig. 2). All four components of this canonical cell death pathway are conserved across disparate animal species, but are apparently absent from bacteria, fungi, and plants. Thus, it is likely that this pathway arose early on in the animal lineage.

2.1 CED-3/Caspase

The most downstream core component of the apoptotic cell death pathway is the protein CED-3, encoded by the ced-3 gene. The role of ced-3 in cell death was initially revealed from genetic studies. Animals mutant for the gene ced-1 (see below) accumulate unengulfed cell corpses during development that are easily detectable using DIC microscopy. A suppressor screen for animals lacking these corpses identified the recessive mutant ced-3(n717), in which most cells fated to die fail to do so. Characterization of the mutant revealed widespread inhibition of cell death, resulting in animals with extra cells (Ellis & Horvitz, 1986). While lineage studies suggest that few, if any, of these extra cells divide (Hoeppner, Hengartner, & Schnabel, 2001), some are able to differentiate (Shaham & Bargmann, 2002), incorporate into neural circuits (White, Southgate, & Thomson, 1991), and even substitute for their surviving sister cells, if those are experimentally ablated (Avery & Horvitz, 1987). ced-3 mutant animals are alive, suggesting that at least under laboratory conditions cell death is not essential (Ellis & Horvitz, 1986). However, some ced-3 animals exhibit defects in chemotaxis to attractive odors, and some exhibit pronounced developmental delay (Ellis, Yuan, & Horvitz, 1991), suggesting that in the wild, cell death likely confers a survival advantage.

The CED-3 protein is a founding member, together with mammalian caspase-1, of the caspase family of proteases (Yuan, Shaham, Ledoux, Ellis, & Horvitz, 1993). Active CED-3 is derived from a precursor that is cleaved to generate three fragments. The N-terminal fragment, which has sequence homology to caspase recruitment domains (CARD), is not required for protease...