Behavior of centrosomes during fertilization and cell division in mouse oocytes and in sea urchin eggs.

Abstract

The forms and locations of centrosomes in mouse oocytes and in sea urchin eggs were followed through the whole course of fertilization and first cleavage by immunofluorescence microscopy. Centrosomes were identified with an autoimmune antiserum to centrosomal material. Staining of the same preparations with tubulin antibody and with the DNA dye Hoechst 33258 allowed the correlation of the forms of the centrosomes with the microtubule structures that they generate and with the stages of meiosis, syngamy, and mitosis. The results with sea urchin eggs conform to Boveri's view on the paternal origin of the functional centrosomes. Centrosomes are seen in spermatozoa and enter the egg at fertilization. Initially, the centrosomes are compact, but as the eggs enter the mitotic cycle the forms of the centrosomes go through a cycle in which they spread during interphase, apparently divide, and condense into two compact poles by metaphase. In anaphase, they spread to form flat poles. In telophase and during reconstitution of the daughter nuclei, the centrosomal material is dposed as hemispherical caps around the poleward surfaces of the nuclei. Mouse sperm lack centrosomal antigen. In the unfertilized mouse oocyte, the meiotic spindle poles are displayed as broad-beaded centrosomes. In addition, centrosomal material is detected in the cytoplasm as particles, about 16 in number, which are foci of small aster-like arrays of microtubules. The length and number of astral microtubules correlate with the size of the centrosomal foci. After sperm incorporation, as the pronuclei develop and more cytoplasmic microtubules assemble, a few of the foci associate with the peripheries of the nuclei. The number of foci multiplies during the first cell cycle. At the end of interphase, all of the centrosomal foci have concentrated on the nuclear peripheries and the cytoplasmic microtubules have disappeared. At prophase, the centrosomes are seen as two irregular clusters, marking the poles which, at metaphase and anaphase, appear as rough bands with foci, and the spindle is typically barrel-shaped. At telophase, the centrosomes are seen as arcs that lie on the nuclear peripheries after cleavage. The ordering of microtubules in all the stages reflects the shapes of the centrosomes. The findings on the sea urchin confirm the classical theory of the paternal origin of centrosomes and contrast with observations tracing the mitotic poles of the mouse egg to maternal centrosomal material. This evidence strengthens the conclusion that mouse centrosomes derive from the oocyte. Mouse and sea urchin fertilization was as described (6). Sea urchin eggs were extracted in a microtubule-stabilization buffer (7), and mouse egg cytoskeletons were stabilized with a similar mixture (4). The cells were affixed to polylysinecoated coverslips (8). Sea urchin eggs were fixed in methanol at -10TC and mouse eggs were fixed in 10 mM ethylene glycol bis(succinimidyl)succinate (9). Autoimmune centrosomal antiserum 5051 was derived from a patient with scleroder...

Bipolar, anastral spindle development in artificially activated sea urchin eggs

“…The general structure of the minispindles present in ammonia-activated sea urchin eggs is similar to the anastral meiotic spindles present in the female oocytes of a wide range of organisms, including Drosophila (Theurkauf and Hawley, 1992), Xenopus (Gard, 1992), and mice (Schatten et al, 1986). In terms of echinoderms, the mini-spindles are similar in shape to the single published image (to our knowledge) of a tubulin-stained sea urchin meitoic spindle present in the literature (Voronina et al, 2003), as well as to certain stages of the meiotic spindles in starfish (Shirai et al, 1990) and sea cucumber (Miyazaki et al, 2005) oocytes.…”

“…Earlier studies have shown that maternal centrosomal material is associated with monasters induced by ammonia activation (Schatten et al, 1992) and with the zygote centrosomes organized by the paternal centriole during normal fertilization Schatten, 1991a, 1997). Furthermore, maternal centrosomal material is associated with acentriolar spindles in other organisms such as the mouse oocyte (Schatten et al, 1986). Perhaps this maternal centrosomal material generates MTs in the mini-spindles in addition to the role of the female chromosomes.…”

“…The idea of a dominant paternal centrosome (or "division center") in sea urchin fertilization is a classic 19th century theory of Theodor Boveri that was promoted by E. B. Wilson (1925) and confirmed by numerous reports (including Sluder et al, 1985;Schatten et al, 1986). Studies have shown that the maternal cytoplasm does contribute components to the zygote centrosome Schatten, 1991a, 1997), however, the sperm centrosome is clearly dominant and provides the centrioles (Paweletz et al, 1984;Sluder et al, 1985).…”

Male Infertility and DNA Damage in Doppel Knockout and Prion Protein/Doppel Double-Knockout Mice

“…In the mouse the centrosome is provided by the oocyte, not the sperm. 42 Thus, the contribution that a sperm makes to embryonic development is primarily restricted to providing the paternal genome. If this DNA is damaged then early embryonic failure might be anticipated soon after the activation of embryonic gene transcription, because many forms of damage will prevent transcription.…”

Contribution of spermatozoal centrosomes to the microtubule-organizing centre in Antarctic minke whale (Balaenoptera bonaerensis)

“…During fertilization in mammals, the centrosome introduced into the oocyte by the spermatozoon plays a critical role in assembly of the microtubule network that brings both male and female pronuclei to the center of the newly formed zygote, as reported in human (Simerly et al, 1995), rhesus monkey (Hewitson et al, 1996), rabbit (Pinto-Correia et al, 1994) and cattle (Navara et al, 1994). On the other hand, the paternal inheritance of MTOC does not occur in mouse (Schatten et al, 1985(Schatten et al, , 1986) and rat (Woolley & Fawcett, 1973) and the microtubule network developed from multiple cytoplasmic asters, instead of a single sperm aster, is involved in the migration of pronuclei. Rodent spermatozoa are lacking centrioles and the majority of pericentriolar proteins after spermiogenesis (Woolley & Fawcett, 1973;Manandhar et al, 1998Manandhar et al, , 1999.…”