Worm Breeder's Gazette 14(1): 68 (October 1, 1995)
Dept. of Genetics, Washington University School of Medicine, St. Louis, MO 63110, firstname.lastname@example.org
We are interested in the regulation of the meiotic cell cycle in oocytes. Specifically, how does each C. elegans oocyte maintain its cell cycle in diakinesis of prophase I, and how is maturation (cell cycle progression) triggered? To address these questions, we have followed oocyte development, maturation, and ovulation by time-lapse Nomarksi microscopy of anesthetized animals 1. Some of the landmark events, shown in Figure 1, have been briefly described previously 2,3. Timing and standard deviations were compiled from observations of 45 N2 oocytes. Time zero is defined as the completion of ovulation into the spermatheca, which coincides closely with fertilization. Oocyte Development Oocytes develop while in diakinesis of meiotic prophase I. As they move proximally, cell and nuclear volumes increase, and chromosomes become more condensed. In hermaphrodites, the nucleolus disappears about 70 minutes before ovulation. Subsequently, the nucleus migrates to the distal surface of the cell. The distal surface of the oocyte can also bend to meet the nucleus, suggesting a physical connection under tension. Both the time when nuclear migration occurs and its rate are variable.4 Meiotic Maturation Progression from prophase to metaphase of meiosis I is termed meiotic maturation.5 The first indication of maturation is nuclear envelope breakdown (NEBD) which begins at -5.7 minutes. At -3.0 minutes, the oocyte begins to change shape from a cube to a sphere, a process we have termed oocyte cortical rearrangement. While vigorous sheath contractions occur at the same time, analyses of sheath ablated animals and mutants defective in sheath activity appear to indicate that oocyte cortical rearrangement does not depend on somatic activity. The events of maturation are highly reproducible with low standard deviations. Ovulation Ovulation, the exit of the oocyte from the gonad arm into the spermatheca, requires contraction of the myoepithelial sheath and dilation of the distal spermatheca. The rate of sheath contractions increases preceding NEBD and peaks at ovulation. The sheath appears to tonically contract as it pulls the dilating distal spermatheca over the most proximal oocyte. As the distal spermatheca closes, cytoplasmic streaming is visible in the oocyte, possibly indicating fertilization. Entry into the uterus occurs 4.2 minutes after ovulation. The meiosis I and II divisions occur in the uterus. Meiotic Prophase Arrest In unmated females, oocytes arrest in diakinesis, failing to undergo meiotic maturation and ovulation for hours or days. Numerous oocytes accumulate, each with an enlarged, distally positioned nucleus lacking a nucleolus. This phenotype defines the arrest point, and separates the events belonging to oocyte development from those of meiotic maturation. Additionally, sheath contractile activity in females is lower than the background level observed between ovulations in hermaphrodites. Oocytes in females do stochastically exit arrest and ovulate at a very low rate (approximately 1/15 the rate for hermaphrodites), perhaps reflecting an inability of C. elegans oocytes to maintain arrest indefinitely. In hermaphrodites, oocyte development, maturation, and ovulation occur in an assembly-line fashion with an ovulation rate of about once per 45 minutes while sperm are present. The time an oocyte spends in diakinesis may reflect a developmental requirement for the execution of certain events and thus might not be a true 'arrest' of the cell cycle. Prospects While the events of oocyte development, maturation, and ovulation occur in a reproducible order, the dependency relationships for these events are not yet defined. Mutant analysis may aid in inferring causality. For instance, we are currently investigating mutants which appear to interfere with the somatic events of ovulation even though meiotic maturation occurs normally within the oocyte. 1 Kirby, C., M. Kusch, K. Kemphues (1990). Dev. Bio., 142:203-215. 2 Ward, S. and J.S. Carrel (1979). Dev. Bio. 73:304-321. 3 Doniach, T (1988). WBG 10(2):64-65. 4 Figure 1 indicates the completion of slow migration (39% of oocytes) or fast migration (36% of oocytes). In 25% of oocytes, the nucleus did not migrate, and in 18%, the nucleus drifted from distal after migrating. Nuclear position is most variable in the first oocyte produced by the arm. 5 Meiotic Maturation is the term used in other systems for oocyte cell cycle progression (i.e. MPF = maturation promoting factor), Masui, Y. and H.J. Clark (1979). Int. Rev. Cytol. 57:185.