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First published online February 9, 2006
doi: 10.1242/10.1242/dev.02276

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Degrade to create: developmental requirements for ubiquitin-mediated proteolysis during early C. elegans embryogenesis

¹ Institute of Molecular Biology, University of Oregon, Eugene, OR 97403, USA.
² Institute of Biochemistry, ETH Zürich, Hönggerberg, 8093 Zürich, Switzerland.

View larger version (34K): [in a new window]   Fig. 1. Schematic representations of different cullin RING ubiquitin ligases and the APC/C. All cullin complexes are Nedd8 modified (see Box 2) and recruit E2-bound ubiquitin through their association with the RING-finger protein Rbx1. (A) The SCF complex is the best-characterized cullin ligase. Cullin 1 proteins bind at their N terminus to the BTB-fold protein Skp1. Skp1, in return, interacts with different F-box proteins, which specifically recognize substrates (blue). (B) Cullin 2-based ECS complexes recruit their substrate through their interaction with elongin B and elongin C and a variable SOCS-box protein, such as ZIF-1 (hence the name ECS complex). (C) Cullin 3-based complexes are unique in using only one protein to bind the cullin and the substrate. (D) The APC/C has many more subunits than the cullin ligases, many of which have not been assigned a function and are shown as a single yellow mass. Although not Nedd8 modified, APC2 has sequence similarity to cullin proteins and interacts with the APC11 RING-finger protein. Different WD40 repeat proteins, e.g. Cdc20 or Cdh1, mediate substrate recognition by the APC/C. APC/C, anaphase promoting complex/cyclosome; BTB, Bric-a-Brack, Tramtrack and Brahma; Cdc20, cell division cycle 20; Cdh1, Cdc20 homolog 1; ECS, elongin B/C, cullin 2, SOCS box; MEI-1, meiosis defective 1; MEL-26, maternal effect lethal; Rbx1, ring box protein 1; SCF, Skp1, cullin 1 and F-box subunits; Skp1, suppressor of kinetochore protein 1; ZIF-1, zinc-finger interacting factor 1.

View larger version (24K): [in a new window]   Fig. 2. Oogenesis and early embryogenesis in C. elegans. (A) A schematic of an adult C. elegans hermaphrodite to show the bilaterally symmetrical U-shaped gonad arms (grey). Anterior is towards the left, dorsal is upwards. The major anatomical features are indicated. (B) One arm of the U-shaped C. elegans gonad. After dividing mitotically in the distal arm of the gonad, germ cell nuclei (in blue), partially enclosed by membrane cubicles, migrate proximally, enter meiosis and cellularize into oocytes. Mature oocytes enter the spermatheca, are fertilized and exit as zygotes into the uterus. The fertilized zygotes form an eggshell, finish meiosis and undergo the first mitotic divisions in the uterus. (C) (a) Meiotic divisions I and II, and (b) the first two mitotic divisions of a C. elegans embryo. DNA, blue; meiotic and mitotic spindles, green. (a) After fertilization, the C. elegans embryo executes the first and second meiotic division in the anterior of the zygote, resulting in the extrusion of two polar bodies and the formation of a haploid female pronucleus (top). The female meiotic spindles are small and lack centrosomes. After meiosis, the male and female pronuclei fuse, and the zygote enters mitosis. (b) The mitotic spindle is organized by two centrosomes (green circles) at the spindle poles. The first division is asymmetric, producing larger anterior and smaller posterior daughter cells that differ in cell cycle timing and mitotic spindle orientation during the next round of mitosis.

View larger version (60K): [in a new window]   Fig. 3. Meiosis and mitosis in early C. elegans embryos. (A-C) C. elegans embryos stained for tubulin (blue) and DNA (yellow). (A) Shortly after fertilization, a small meiotic spindle assembles at the anterior cortex. At this stage, most tubulin is not recruited to the spindle, but is scattered throughout the cytoplasm. (B) During the first mitosis, a large and asymmetric spindle forms. The first mitosis produces daughter cells of unequal size. (C) The anterior somatic precursor AB is shown undergoing anaphase of mitosis, while the posterior germline precursor P1 is still in metaphase.

View larger version (57K): [in a new window]   Fig. 4. A schematic of C. elegans embryos undergoing meiosis, axis specification and the first mitotic division in wild-type and different E3 ubiquitin ligase mutants. (A) During meiosis in wild-type embryos, cytoplasmic cell fate determinants are uniformly distributed in the cytoplasm and the PAR-3 protein (blue) occupies the entire cortex (a). After meiosis, the centrosomes that accompany the male pronucleus trigger the establishment of anteroposterior (AP) polarity: starting at the site of sperm aster-cortex contact, the posterior pole is specified and PAR-2 (red) spreads, replacing PAR-3 at the posterior cortex. (b,c) During this process, cytoplasmic cell fate determinants are also actively partitioned to the anterior (yellow; e.g. MEX-5 and MEX-6) and posterior (green; e.g. MEX-1, PIE-1, POS-1 and germline P granules) cytoplasm. (d) The first mitotic spindle aligns along the AP axis and cytokinesis cleaves the cell asymmetrically. (e) The respective cell fate determinants are confined to the anterior or posterior cell, and residual `mis-localized' determinants are degraded by the ubiquitin proteasome system late at the two-cell stage. (B) Partial loss-of-function APC/C mutants do not arrest in meiosis, but go on to divide mitotically. (a,b) In these embryos, the meiotic spindle often persists longer than in wild type, the sperm pronucleus does not properly associate with the cortex and an aberrant cortical PAR-2 crescent often forms. (c-e) Subsequently, polarity is lost and the embryo divides symmetrically. (C) (a-c) Loss of function of CUL-2 results in a delayed exit from meiosis II and aberrant establishment of the cortical PAR-2 domain. (c,d) In some embryos, this polarity reversal is corrected and the first division is executed properly. (d,e) Although cortical polarity is normal, the cytoplasm is not polarized properly, owing to impaired degradation of cell fate determinants. (D) No polarity defects are observed in embryos in which CUL-3 has been inactivated. (a,b) Rather, extensive cortical contractions are apparent during pronuclear migration, owing to ectopic activation of the acto-myosin cytoskeleton by MEL-26. (c,d) The failure to degrade MEI-1/katanin results in the severing of mitotic microtubules and spindle orientation defects. (e) During and after cytokinesis, MEL-26-dependent ectopic furrows appear again.