Einhard Schierenberg

The embryonic cell lineage of the nematode Caenorhabditis elegans.

The embryonic cell lineage of Caenorhabditis elegans has been traced from zygote to newly hatched larva, with the result that the entire cell lineage of this organism is now known. During embryogenesis 671 cells are generated; in the hermaphrodite 113 of these (in the male 111) undergo programmed death and the remainder either differentiate terminally or become postembryonic blast cells. The embryonic lineage is highly invariant, as are the fates of the cells to which it gives rise. In spite of the fixed relationship between cell ancestry and cell fate, the correlation between them lacks much obvious pattern. Thus, although most neurons arise from the embryonic ectoderm, some are produced by the mesoderm and a few are sisters to muscles; again, lineal boundaries do not necessarily coincide with functional boundaries. Nevertheless, cell ablation experiments (as well as previous cell isolation experiments) demonstrate substantial cell autonomy in at least some sections of embryogenesis. We conclude that the cell lineage itself, complex as it is, plays an important role in determining cell fate. We discuss the origin of the repeat units (partial segments) in the body wall, the generation of the various orders of symmetry, the analysis of the lineage in terms of sublineages, and evolutionary implications.

Cell lineages and developmental defects of temperature-sensitive embryonic arrest mutants in Caenorhabditis elegans.

The cellular phenotypes of 11 temperature-sensitive mutants in nine genes (emb-1-emb-9) arresting embryogenesis in Caenorhabditis elegans are described, including early embryonic cell lineages and developmental defects and the terminal phenotypes at the stage of arrest at the nonpermissive temperature (25°C), as well as residual phenotypes at the permissive temperature (16°C). By Nomarski microscopy of living embryos, the behavior of individual cells in mutant embryos is compared to that of the same cells in wild-type embryos. All mutants but one (emb-9) have visible defects before the 50-cell stage, even if they arrest much later. Cell proliferation continues in the absence of normal morphogenesis. The terminal phenotype of the late-arresting mutants looks grossly abnormal. The overall rate of cell division is faster than in the wild-type in two mutants, emb-3 and emb-5(hc67), and slower in five mutants, emb-2, emb-4, emb-5(hc61), emb-6, and emb-7. In five of these mutants, emb-3, emb-4, emb-5 (both alleles), and emb-7, the rate and the sequence of divisions of specific cell lines is altered. For all mutants with timing defects, maternal gene expression is at least sufficient. Gastrulation is abnormal in the two emb-5 mutants. A premature division of the intestine precursor cells occurs in close temporal coincidence with the defective execution stage (determined by temperature-shift experiments). The order of migration and division of the intestine precursor cells is reversed in gastrulation, and the cell pattern of the intestine primordium is abnormal. It seems possible that the timing error is responsible for the pattern defect. The cleavage behavior of emb-3 eggs indicates that germ line determinants are prelocalized in the egg of C. elegans.

Genetics and mode of expression of temperature-sensitive mutations arresting embryonic development in Caenorhabditis elegans

Eleven temperature-sensitive mutations causing arrest of embryogenesis in Caenorhabditis elegans have been mapped. The mutations define nine genes (emb-1 to emb-9) on four chromosomes. The functions of six genes seem to be required exclusively for embryogenesis. Mutants in these genes have no other detectable phenotype at the permissive (16°C) or nonpermissive (25°C) temperature. The function of the other three genes is also required for postembryonic development. As shown by progeny tests for parental effects, for seven genes, maternal gene expression is necessary and sufficient for normal embryogenesis; for one gene, emb-2, either maternal or zygotic expression is sufficient; for one gene, emb-9, zygotic expression is necessary and sufficient. The high proportion of emb genes with maternal expression is consistent with the model of intracellular preprogramming of the egg of C. elegans (U. Deppe, E. Schierenberg, T. Cole, C. Krieg, D. Schmitt, B. Yoder, and G. von Ehrenstein, 1978; Proc. Nat. Acad. Sci. USA 75, 376–380). Two developmental stages have been defined by temperature-shift experiments: (1) the normal execution stage indicating the time of execution of the normal event at the permissive temperature; (2) the defective execution stage indicating the time of the execution of an irreversible defect at the nonpermissive temperature. The classes of mutants defined by the progeny tests have corresponding execution stages, but the maternal necessary and sufficient class is subdivided into mutants executing during oogenesis or embryogenesis.

The cellular anatomy of embryos of the nematode Caenorhabditis elegans: Analysis and reconstruction of serial section electron micrographs

Abstract As described from light microscopy, embryogenesis of the free-living soil nematode Caenorhabditis elegans follows a strictly determinate cleavage pattern, producing a newly hatched juvenile with about 550 cells arranged quite predictably. In this communication we present results on the electron microscopy of C. elegans embryos and introduce methods for fixing, embedding, and serially sectioning embryos encased in the egg shell. Fixation at elevated temperature either with osmium tetroxide alone or with glutaraldehyde followed by osmium tetroxide gives reproducible results with embryos in all developmental stages so far tested, from the fertilized egg to hatching. Eighteen wild-type eggs at various stages have been sectioned to date. We have achieved using newly developed procedures for analyzing electron micrographs of serial sections detailed reconstructions of the cellular anatomy of complete embryos of a metazoan organism. Three-dimensional serial section reconstructions were made with a computer system. We characterize and map the 24 cells of an early-stage embryo in this report. Additionally, we can specify the lineage history of all cells of this embyro by matching the reconstructed three-dimensional arrangement of this series to a living embryo at this stage, where cell lineage has been observed with Nomarski optics and analyzed using videotape ( U. Deppe, E. Schierenberg, T. Cole, C. Krieg, D. Schmitt, B. Yoder, and G. von Ehrenstein, 1978 , Proc. Nat. Acad. Sci. USA 75, 376–380). In addition, cytoplasmic and nuclear morphological features such as incomplete membranes between sister cells, rounding-off of the cytoplasm, and chromatin condensation patterns have been correlated with cell division. Mapping of such structures presents a new method by which supplementary lineage information can be obtained directly from an electron micrographic series.

Altered cell-division rates after laser-induced cell fusion in nematode embryos.

In embryos of Caenorhabditis elegans adjacent cells belonging to different cell lines have been fused by disrupting the cell membrane between them with a single laser pulse. This leads to cytoplasmic mixing. Synchronous mitoses of the two nuclei result in the direct formation of four cells. Subsequent alteration of the cell cycle rhythm in some specific--but not all--descendants of the fused cells is observed, leading to abnormal cell patterns and often embryonic arrest.

Cellular development of a nematode: 3-D computer reconstruction of living embryos

SummaryEmbryos of the free-living soil nematodeCaenorhadditis elegans are capable of developing normally outside the mother; we have monitored this process in isolated embryos by light microscopy and recorded it on video tape. The size and position of each nucleus were entered into a computer at short time intervals from the 2- to 102-cell stages. Models were reconstructed in which nuclei are represented by spheres and assigned different colors and patterns according to lineage membership. Three-dimensional reconstructions aid visualization of the spatial arrangement of nuclei and demonstrate the small degree of positional variance among individuals. The dynamic processes of nuclear growth during the cell cycle, division, migration, and patern formation can be quantitatively analyzed. Our knowledge of the complete embryonic lineage allows the correlation of nuclear behavior with eventual cellular fate.

Laser-Induced Cell Fusion

In recent years a number of different techniques have been developed to fuse cells of various origin in order to obtain hybrids with qualities different from each of the original cells. In many cases, one wants to fuse large numbers of cells (e.g., to generate hybridomas), and fusion occurs more or less randomly (e.g., Kohler and Milstein, 1975). With other methods, individual cells have been fused under controlled conditions (e.g., McGrath and Solter, 1984). So far a prerequisite for fusing two preselected cells has been that they must be isolated away from other cells.

Computer-aided three-dimensional reconstruction of nematode embryos from EM serial sections.

Embryos of the nematode Caenorhabditis elegans were serially sectioned and photographed in the electron microscope (EM). The micrographs were used to produce three-dimensional (3D) reconstructions. Size and position of each nucleus were entered into a computer, displayed as spheres, and were color-coded to indicate lineage membership. Location in space and position in the cell cycle are generally adequate criteria to identify cells. The reconstructions allow visualization of lineage-related topographic patterns and ultrastructural analysis of differently determined cells.

Embryonic Cell Lineages and Segregation of Developmental Potential in Caenorhabditis elegans

Embryogenesis of the nematode Caenorhabditis elegans is determinate and virtually invariant from individual to individual. The fertilized egg develops into an anatomically relatively simple juvenile animal having a constant number of only 550 cells (or nuclei) at hatching. The complete embryonic cell lineage up to the 220-cell stage has been described previously, and some lineages have been followed considerably further (Deppe et al. 1978; Krieg et al. 1978; Schierenberg 1978; von Ehrenstein and Schierenberg 1980). During postembryonic development, the cell number increases to about 950 in mature hermaphrodites (including the 143 of the somatic gonad structures) and to about 1025 in males. These cells arise from about 50 blast cells which resume division after hatching. The lineages of all of these cells have been described (Sulston 1976; Sulston and Horvitz 1977; Kimble and Hirsh 1979; Sulston et al. 1980).