Giant sperm cells with accessory macrotubules in a neuropteran insect

The flagellar axoneme of the atypical spermatozoa (paraspermatozoa) of Mantispa perla (Neuroptera, Planipennia) contains accessory microtubules or rather macrotubules that are 55 nm in diameter and that has a wall consisting of about 40 protofilaments. The sperm tail further contains two giant mitochondrial derivatives, which during spermiogenesis store an electron dense material. The mature spermatozoon has a flattened acrosome and a elliptical nucleus. These giant spermatozoa may furnish nutrients to the functional spermatozoa (euspermatozoa) when they reach the female genital tracts or/and they function in sperm competition filling the spermatheca.     

Distinct mechanisms eliminate mother and daughter centrioles in meiosis of starfish oocytes

Centriole elimination is an essential process that occurs in female meiosis of metazoa to reset centriole number in the zygote at fertilization. How centrioles are eliminated remains poorly understood. Here we visualize the entire elimination process live in starfish oocytes. Using specific fluorescent markers, we demonstrate that the two older, mother centrioles are selectively removed from the oocyte by extrusion into polar bodies. We show that this requires specific positioning of the second meiotic spindle, achieved by dynein-driven transport, and anchorage of the mother centriole to the plasma membrane via mother-specific appendages. In contrast, the single daughter centriole remaining in the egg is eliminated before the first embryonic cleavage. We demonstrate that these distinct elimination mechanisms are necessary because if mother centrioles are artificially retained, they cannot be inactivated, resulting in multipolar zygotic spindles. Thus, our findings reveal a dual mechanism to eliminate centrioles: mothers are physically removed, whereas daughters are eliminated in the cytoplasm, preparing the egg for fertilization.     

Behavior of centrioles during meiosis in the male silkworm, Bombyx mori (Lepidoptera)

The behavior of centrioles during eupyrene and apyrene meiosis was examined in the silkworm, Bombyx mori , by transmission electron microscopy and indirect immunofluorescence for tubulin. In eupyrene spermatocytes the centrioles, accompanied by axonemes, attached temporarily to the nucleus at diplotene, then detached from the nucleus in diakinesis. After the separation, a beret-shaped structure consisting of a double membrane covered the proximal region of the pair of centrioles. The structure disappeared after breakdown of the nuclear membrane. The centriole, with the axoneme, reattached to the nucleus at telophase I. The process was repeated during meiosis II until the centrioles maintained their nuclear attachment in newly developed spermatids. In stark contrast to their eupyrene counterparts, apyrene spermatocytes were conspicuously devoid of any attachment of the centrioles to the nucleus. These eupyrene-specific and apyrene-specific relationships were consistently and repeatedly found between the nuclear membrane and centrioles, giving rise to suspicion that the behavioral phenomena may be related to differentiation of the dimorphic sperm types.     

Freewheeling centrioles

The century-old controversy over the reproduction and function of the centriole is examined to elucidate the conceptual and methodological issues that have made it so resistant to closure. The study of centrioles is situated in two distinct eras punctuated by the deployment of the electron microscope. From the late 19th century to the mid-20th century, centrioles were defined largely in functional terms- as self-reproducing 'central bodies' playing a directive role in mitosis. During this period, when their structure remained unknown, their universal presence in all cells was ambiguous, and their reality was seriously debated. A conceptual switch occurred after the mid-1950s. When the centriole was made visible under the electron microscope, it was defined in terms of its characteristic cart-wheel structure, but its function and manner of reproduction have remained enigmatic. The controversy over the nature of centrioles illustrates the dynamic interplay of techniques, theories, and background assumptions in the production of scientific knowledge. It also highlights the difficulties biologists face in coming to grips with problems of cell structure and intracellular morphogenesis.     

Flies without centrioles

Centrioles and centrosomes have an important role in animal cell organization, but it is uncertain to what extent they are essential for animal development. The Drosophila protein DSas-4 is related to the human microcephaly protein CenpJ and the C. elegans centriolar protein Sas-4. We show that DSas-4 is essential for centriole replication in flies. DSas-4 mutants start to lose centrioles during embryonic development, and, by third-instar larval stages, no centrioles or centrosomes are detectable. Mitotic spindle assembly is slow in mutant cells, and approximately 30% of the asymmetric divisions of larval neuroblasts are abnormal. Nevertheless, mutant flies develop with near normal timing into morphologically normal adults. These flies, however, have no cilia or flagella and die shortly after birth because their sensory neurons lack cilia. Thus, centrioles are essential for the formation of centrosomes, cilia, and flagella, but, remarkably, they are not essential for most aspects of Drosophila development.     

F-actin is a component of the karyosome in neuropteran oocyte nuclei

The principles underlying the assembly of intranuclear compartments are only beginning to be understood. The karyosome is an organelle typical of oocyte nuclei. It represents the tightly packed oocyte chromosomes, arrested at the diplotene of meiotic prophase. It has been known from several insect orders that a prominent capsule of unknown materials is built around the karyosome in the course of previtellogenesis and vitellogenesis. Here we show that F-actin, detected by dye-coupled phalloidin, is a major molecular component of the karyosome capsule in Neuroptera. We investigated capsule formation in six species belonging to the family Chrysopidae. Though F-actin was present in the capsules of all six species there were striking interspecific differences in the morphological array of actin filaments and the developmental dynamics of actin deposition in the capsule. The potential biological significance of the karyosome capsule is discussed with respect to the presence of extrachromosomal rDNA in neuropteran oocytes and the molecular functions known from F-actin. Our results corroborate the still controversial hypothesis of a role for actin as a nuclear protein.     

Male gametogenesis without centrioles

The orientation of the mitotic spindle plays a central role in specifying stem cell-renewal by enabling interaction of the daughter cells with external cues: the daughter cell closest to the hub region is instructed to self-renew, whereas the distal one starts to differentiate. Here, we have analyzed male gametogenesis in DSas-4 Drosophila mutants and we have reported that spindle alignment and asymmetric divisions are properly executed in male germline stem cells that lack centrioles. Spermatogonial divisions also correctly proceed in the absence of centrioles, giving rise to cysts of 16 primary spermatocytes. By contrast, abnormal meiotic spindles assemble in primary spermatocytes. These results point to different requirements for centrioles during male gametogenesis of Drosophila. Spindle formation during germ cell mitosis may be successfully supported by an acentrosomal pathway that is inadequate to warrant the proper execution of meiosis.     

Do centrioles contain ribosomes?

Centrioles induced via a number of parthenogenetic agents regularly reveal the presence of one or more granules within their central cores. Though not a new discovery, these centriolar granules have carefully been re-evaluated here. Considering various criteria, it is proposed that these granules are intra-centriolar ribosomes. More specifically, they are more comparable to 705 ribosomes than to 805 cytoribosomes. The data suggest that within the lumen of centrioles, certain centriolar proteins are synthesized on ribosomes that may be uniquely centriolar.     

Mitosis in a cell with multiple centrioles

N115 mouse neuroblastoma cells possess a large number of microtubule organizing centers (MTOCs) which can be identified ultrastructurally as single centrioles. The distribution and activity of these organizing centers can be followed through all stages of the cell cycle by labeling microtubules with anti-tubulin and chromatin with the Hoechst dye, Bisbenzimid. We have found that multiple MTOCs persist and continue to organize microtubules during mitosis. They exhibit a well- defined sequence of movements, starting from a loose cluster during interphase, proceeding to a widely and evenly dispersed arrangement in prophase, gathering into small clusters and chains during prometaphase, and residing in two ring-shaped groups at the mitotic poles during metaphase and anaphase. Despite their large number of centrioles, virtually all N115 cells show a normal bipolar mitosis, but often with unequal numbers of centrioles at the two poles. Such observations bring into question the importance of the centriole in establishing bipolar division in this cell type.     

TSKS concentrates in spermatid centrioles during flagellogenesis

Centrosomal coiled-coil proteins paired with kinases play critical roles in centrosomal functions within somatic cells, however knowledge regarding gamete centriolar proteins is limited. In this study, the substrate of TSSK1 and 2, TSKS, was localized during spermiogenesis to the centrioles of post-meiotic spermatids, where it reached its greatest concentration during the period of flagellogenesis. This centriolar localization persisted in ejaculated human spermatozoa, while centriolar TSKS diminished in mouse sperm, where centrioles are known to undergo complete degeneration. In addition to the centriolar localization during flagellogenesis, mouse TSKS and the TSSK2 kinase localized in the tail and acrosomal regions of mouse epididymal sperm, while TSSK2 was found in the equatorial segment, neck and the midpiece of human spermatozoa. TSSK2/TSKS is the first kinase/substrate pair localized to the centrioles of spermatids and spermatozoa. Coupled with the infertility due to haploinsufficiency noted in chimeric mice with deletion of Tssk1 and 2 (companion paper) this centriolar kinase/substrate pair is predicted to play an indispensable role during spermiogenesis.     

Life without centrioles: cilia in the spotlight

Centrioles are critical cellular components that form the architectural core of both centrosomes and basal bodies, the nucleating structures of cilia. New work, including a study in this issue (), highlights the unexpected finding that lack of centrioles does not impede development in the fruit fly. Rather, flies reach maturity but then die because their sensory neurons lack cilia.     

Nonequivalence of maternal centrosomes/centrioles in starfish oocytes: selective casting-off of reproductive centrioles into polar bodies

It is believed that in most animals only the paternal centrosome provides the division poles for mitosis in zygotes. This paternal inheritance of the centrosomes depends on the selective loss of the maternal centrosome. In order to understand the mechanism of centrosome inheritance, the behavior of all maternal centrosomes/centrioles was investigated throughout the meiotic and mitotic cycles by using starfish eggs that had polar body (PB) formation suppressed. In starfish oocytes, the centrioles do not duplicate during meiosis II. Hence, each centrosome of the meiosis II spindle has only one centriole, whereas in meiosis I, each has a pair of centrioles. When two pairs of meiosis I centrioles were retained in the cytoplasm of oocytes by complete suppression of PB extrusion, they separated into four single centrioles in meiosis II. However, after completion of the meiotic process, only two of the four single centrioles were found in addition to the pronucleus. When the two single centrioles of a meiosis II spindle were retained in the oocyte cytoplasm by suppressing the extrusion of the second PB, only one centriole was found with the pronucleus after the completion of the meiotic process. When these PB-suppressed eggs were artificially activated to drive the mitotic cycles, all the surviving single centrioles duplicated repeatedly to form pairs of centrioles, which could organize mitotic spindles. These results indicate that the maternal centrioles are not equivalent in their intrinsic stability and reproductive capacity. The centrosomes with the reproductive centrioles are selectively cast off into the PBs, resulting in the mature egg inheriting a nonreproductive centriole, which would degrade shortly after the completion of meiosis.     

Possible role of centrioles in stabilizing cytoplasmic microtubules

Small fragments of the peripheral cytoplasm were obtained from cytochalasine B-treated mouse embryo fibroblasts and studied for distribution of microtubules by indirect immunofluorescence. Microtubules were demonstrated to progressively depolymerize in these fragments which did not contain any tubules after 6 hours of incubation in the growth medium. This effect was specific for microtubules, since the distribution of intermediate filaments remained unchanged during incubation. The fragments remained viable during incubation, inasmuch no changes were detectable in the membrane potential of the mitochondria stained with rhodamine 123. Progressive destruction of microtubules in the tiny cell fragments is likely to be related to the lack of centrioles in such fragments.