Differential Mitosis Research Articles

GTPase Ran strongly accumulates at the kinetochores of somatic chromosomes in the spermatogonial mitoses of Acricotopus lucidus (Diptera, Chironomidae)

Unequal chromosome segregation and spindle formation occurs in the last gonial mitosis in the germ line of the chironomid Acricotopus lucidus. During this differential mitosis, all germ line-limited chromosomes (=Ks) migrate undivided to only one pole of the cell, while the somatic chromosomes (=Ss) first remain in the metaphase plane, and with the arrival of the Ks at the pole, they then separate equally. The evolutionarily conserved GTPase Ran plays a crucial role in many cellular processes. This includes the regulation of microtubule nucleation and stabilisation at kinetochores and of spindle assembly during mitosis, which is promoted by a RanGTP concentration gradient that forms around the mitotic chromosomes (Kalab et al. in Science 295:2452-2456, 2002, Nature 440:697-701, 2006). In the present study, a strong accumulation of Ran was detected by immunofluorescence at the kinetochores of the Ss in normal gonial and differential gonial mitoses of males of A. lucidus. In contrast, no Ran accumulation was observed at the kinetochores of the Ss in the metaphases of brain ganglia mitoses or of aberrant spermatocytes or in metaphases I and II of spermatocyte meiotic divisions. Likewise, there was no accumulation at the kinetochores of Drosophila melanogaster mitotic chromosomes from larval brains. The specific accumulation of Ran at the kinetochores of the Ss in differential gonial mitoses of A. lucidus strongly suggests that Ran is involved in a mechanism acting in this exceptional mitosis, which retains the Ss at the metaphase plane and prevents a premature separation and unequal segregation of the Ss during monopolar migration of the Ks.

CYTOCHEMICAL AND ULTRASTRUCTURAL STUDIES OF OOGENESIS IN THE DYTISCIDAE

SUMMARY Oogenesis of Dytiscidae is characterized by a remarkable phenomenon occurring in the germarium: each oogonium, after a series of normal mitoses, is transformed into a preoocyte characterized by a mass of extra- chromosomal chromatin called the Giardina mass or body or ring (GB), from the name of its Discoverer. The preoocyte undergoes four differential mitoses and the GB stays in only one cell, the oocyte, the other fifteen becoming nurse cells. The behaviour of this extrachromosomal formation is interesting but its significance is not completely clear, partly because it disintegrates and disappears during growth of the oocyte. Cytochemical study has shown that at its appearance the GB consists predominantly of DNA and at the time of its destruction there is an increasing synthesis of RNA. The results were obtained with a variety of cytochemical and autoradiographic techniques and have been confirmed by various Authors. The nurse cells grow too and their nuclei undergo numerous endomitoses. DNA sy...

Centrosome hyperamplification with the formation of multiple asters and programmed chromosome inactivation in aberrant spermatocytes during male meiosis in Acricotopus

In the germ line of the midge Acricotopus lucidus, an unequal chromosome segregation occurs in the last gonial mitosis prior to meiosis. This results in one daughter cell receiving only somatic chromosomes (Ss), whereas the other cell is given all the so-called germ line limited chromosomes (Ks) in addition to the Ss. The cytokinesis following this differential mitosis is incomplete and the daughter cells remain connected by a permanent cytoplasmic bridge. The cell with the Ss and Ks develops into a primary oocyte or spermatocyte, whereas the cell containing only Ss differentiates as a nurse cell in the female or as an aberrant spermatocyte in the male. When the primary spermatocyte enters meiosis, the Ss in the connected aberrant spermatocyte undergo chromosome condensation but the aberrant spermatocyte remains undivided, with the condensed metaphase status and inactivation of the Ss persisting during both meiotic divisions. These events indicate a programmed inactivation of all chromosomes in the aberrant spermatocyte at the beginning of meiosis. The alterations in the microtubule arrangements and of the distribution of mitochondria in the spermatocytes during meiosis have been followed via live-cell fluorescence labelling with the TubulinTracker and MitoTracker reagents and by transmission electron microscopy. The observations reveal a hyperamplification of the centrosomes and the formation of tetrapolar asters in the non-dividing aberrant spermatocytes containing the condensed Ss. The programmed inactivation of the Ss in the aberrant spermatocyte is suggested to have developed during evolution to inhibit the entry of the aberrant spermatocytes into meiosis, thereby preventing the formation of sperms containing only Ss but no Ks.

Asymmetric distribution of mitochondria and of spindle microtubules in opposite directions in differential mitosis of germ line cells in Acricotopus

Additional chromosomes present only in the germ line are a specific feature of the Orthocladiinae, a subfamily of the Chironomidae. During the complex chromosome cycle in the orthocladiid Acricotopus lucidus, about half of the germ-line-limited chromosomes (Ks) are eliminated in the first division of the primary germ cells. Following normal gonial mitoses, the reduction in the number of Ks is compensated for, in the last mitosis prior to meiosis, by a monopolar movement of the unseparated Ks, while the somatic chromosomes (Ss) segregate equally. This differential mitosis produces daughter cells with different chromosome constitutions and diverse developmental fates. A preferential segregation of mitochondria occurs to one pole associated with an asymmetric formation of the mitotic spindle. This has been detected in living gonial cells in both sexes by using MitoTracker probes and fluorochrome-labelled paclitaxel (taxol). In males, the resulting unequal partitioning of mitochondria to the daughter cells is equalised by the transport of mitochondria through a permanent cytoplasmic bridge from the aberrant spermatocyte to the primary spermatocyte. This asymmetry in the distribution and in the segregation of cytoplasmic components in differential gonial mitosis in Acricotopus may be involved in the process of cell-fate determination.

Immunofluorescence study of spindle microtubule arrangements during differential gonial mitosis of Acricotopus lucidus (Diptera, Chironomidae).

The changes in microtubule arrangements correlated with the behavior of the germ line limited and soma chromosomes were studied during the last unequal gonial mitosis, the so-called differential mitosis, of the chironomid Acricotopus lucidus by indirect immunofluorescence using a monoclonal anti-beta-tubulin antibody, and by simultaneous staining with the DNA-specific fluorescence dye DAPI. An impressive difference in microtubule density between both half spindles was determined in metaphase and during the monopolar anaphasic migration of the germ line limited chromosomes. In the following normal separation of the soma chromosomes, a similar microtubule density in both half spindles occurred. In each of the half spindles, chromosome movement and spindle elongation occurred independently, and in one half spindle two anaphasic chromosome movements ran off one after another, the second without a simultaneous spindle elongation.

Development of the male scent organ ofCreatonotos transiens (Lepidoptera, Arctiidae) during metamorphosis

(1) The male abdominal scent organ (corema) of the arctiid mothCreatonotos transiens consists of a basal bladder and four tubes. It can be everted from the sternal intersegmental membrane 7/8. Its scent hairs (scales) produce and release the pheromone hydroxydanaidal, which attracts both sexes. Pyrrolizidine alkaloids (PA) ingested by the larva with its food are not only precursors of the pheromone but also a morphogen, which quantitatively controls the growth of the pupal corema and, thus, its final size and number of hairs. (2) The coremata arise from epidermalanlagen at the anterior border of the 8th abdominal sternite. If male larvae are fed 1 mg PA these organs begin to develop from small vesicles, and four tubes then arise during the first 3 pupal days. The corresponding mitoses reach their peak at 36 h. During the next 2 days the tubes shorten, while the walls become thin and doubly folded. The total surface of the corema increases about 20 times because of the shape transformation of the epidermal cells from prismatic to very flat. (3) The scent hairs originate from trichogen cells, which arise together with their associated tormogen cells during the 1st pupal day by way of differential mitoses. As the trichogen cells grow, their nuclei enlarge by way of endomitoses, elongate distally, and thus produce the hairs that extend into the lumen of the corema. Tormogen cells degenerate by the 8th day at latest. The hairs in each tube form a thick, caudally oriented bundle. The hair cells are finally bottle-shaped and at day 6 they extend freely into the hemolymph space. They are probably also the pheromoneproducing cells in later pupal and early imaginal life. Mitoses that produce trichogen cells stop after the 1st day, those producing epithelial cells 2 days later. This delay shifts the ratio of the two cell types from about 1∶11 (18 h) to 1∶40. (4) The processes hitherto described refer to “normogenesis” with ample PA supply. “Control” coremata in PA-free or PA-deficient specimens develop in principle in the same way, but at a slower rate, with minimal hair cell numbers barely 1/10th of normal, or at any rates between, depending upon the earlier PA supply. The size of control coremata varies from very small to small; even the hair cells and the hairs are smaller. (5) PA regulates corema development quantitatively through the number of mitoses of its cells and of endomitotic steps of the hair cells. In PA-treated specimens the coremaanlage is already advanced prior to pupation, at about the time when its sensitivity to PA influence terminates, in the early prepupa. Since PA only affects the anlagen of the corema and not that of any other body part (not even the basal coremal bladder), we postulate a selective interaction of PA with the presumptive corema cells. We found earlier that ecdysone is also involved, since the respective cell numbers can only be realized if this hormone is present.

Sensitive periods for abnormal patterning on a leg segment inDrosophila melanogaster.

The development of a leg segment of the fruitflyDrosophila melanogaster was analyzed in order to determine whether the orderliness of the segment's bristle pattern originates via waves of cellular interactions, such as those that organize the retina. Fly development was perturbed at specific times by either teratogenic agents (gamma rays, heat shock, or the drug mitomycin C) or temperature-sensitive mutations (l(1)63, l(1) Notchts1, orl(1) shibire ts1 ), and the resulting abnormalities (e.g., missing or extra structures) were mapped within the pattern area. If bristles develop in a linear sequence across the pattern, then they should show sensitivity to perturbations in the same order, and wavefronts of cuticular defects should result. Contrary to this prediction, the maps reveal no evidence for any directional waves of sensitivity. Nevertheless, other clues were uncovered as to the nature and timing of patterning events. Chemosensory bristles show earlier sensitivities than mechanosensory bristles, and longer bristles precede shorter ones. The types and sequence of cuticular abnormalities imply the following stages of bristle pattern development: (1) scattered inception of bristle mother cells, each surrounded by an inhibitory field, (2) alignment of the mother cells into rows, (3) differential mitoses, (4) assignment of cuticular fates to the mitotic progeny, (5) polytenization of the bristle cells, (6) fine-tuning adjustments in bristle spacing, and (7) signalling from bristle cells to adjacent epidermal cells, inducing them to form "bracts".

Morphogenesis of the antenna of the male silkmoth, Antheraea polyphemus. II. Differential mitoses of ‘dark’ precursor cells create the Anlagen of sensilla

The antenna of the male silkmoth Antheraea polyphemus develops from a one-layered, flattened epidermal sac during the pupal phase. Within the first day post-apolysis (developmental stages 1 and 2), this epithelium differentiates into ‘sensillogenic’ and ‘nonsensillogenic’ regions, while numerous slender ‘dark cells’ interpreted as the precursor cells of sensilla arise in the former. Approximately between the first and second day post-apolysis (developmental stage 3), the dark cells retract to the apical pole of the epidermis, assume a round shape, and undergo a series of differential mitoses with spindles usually oriented parallel to the epidermal surface. These mitoses finally yield the Anlagen of the olfactory sensilla trichodea, each consisting of mostly 6–7 dark cells arranged side by side. In most of the Anlagen, 3–4 of these cells are situated more basally, each giving off a slender apical process which together are arranged in a fascicle. These are the prospective 2–3 sensory neurons plus the thecogen cell, which most probably is a sister cell of the former. Three additional cells are arranged more apically and partly enclose the fascicle of presumed sensory and thecogen cell processes. These are interpreted as the trichogen plus 2 tormogen cells, one of the latter degenerating later during development. In the basal region of the sensillogenic epidermis, massive signs of cell degeneration have been found. At stage 3, the basal epidermal feet in the non-sensillogenic regions have assumed a more uniform orientation as compared with the preceding stages.

Unusual germ line limited chromosomes in Acricotopus lucidus (Diptera, Chironomidae)

A small supernumerary polytene chromosome was found during the last 8 years in some rare cases in larval salivary gland cells of Acricotopus lucidus (Diptera, Chironomidae). The chromosome may be derived from the germ line restricted parts of the genome. It consists of a short heterochromatic segment and of euchromatic sections with banding patterns homologous to sections of the short arm of soma chromosome I. When examining male meiosis, an exceptional small germ line limited chromosome was found. It is believed that this chromosome was not always recognized during soma elimination as a germ line limited chromosome, probably because of its partial homology to one of the soma chromosomes, and was then polytenized in salivary gland cells. Another germ line limited chromosome with a characteristic morphology and with a special behavior in differential gonial mitosis was found to have existed for more than 12 years in a laboratory stock. In differential gonial mitosis this special germ line limited chromosome partly pairs with the long arm of soma chromosome I. The present results strongly support the idea that the germ line limited chromosomes of A. lucidus are derived from the soma chromosomes, and show that chromosomes of the germ line restricted part of the genome can persist for many generations in a laboratory stock in spite of complex chromosome elimination mechanisms in the primary germ cells. Key words: germ line limited chromosomes, supernumerary polytene chromosome, salivary gland, Acricotopus lucidus.

The spermatogenesis of Ephydatia fluviatilis (Porifera)

Spermatogensis of the freshwater sponge Ephydatia fluviatilis begins with differential mitosis of choanocytes in the flagellated chambers. The spermatogonia thus produced aggregate in the mesenchyme by amoeboid movement. They are surrounded by dendritic cells which eventually form the walls of spermatocysts, within which spermatogenesis takes place. Notable features are the early formation of flagella in first order spermatocytes and the appearance of two flagella in the metaphase of the second maturation division.

X-ray induced rearrangements between germ-line limited and soma chromosomes of Acricotopus lucidus (Diptera, Chironomidae)

The germ-line limited chromosomes (Ks) [K being derived from ‘Keimbahn’ (Bauer, 1970)] of Acricotopus lucidus were studied in gonial and differential mitosis. After C-banding the soma chromosomes (Ss) are stained only at their centromeric regions whereas the Ks exhibit centromeric, intercalary and terminal heterochromatin. By X-raying sperms it was attempted to transfer K sections on or into Ss in order to bring finally S-linked K sections to polytenization in the salivary glands, and to obtain more knowledge about the structure of Ks. Seven F1-larvae were detected with K-S-rearrangements: four with insertions of heterochromatic segments, two with insertions of sections with S-homologous banding pattern and one with a translocated K chromosome part, which consists of S-homologous euchromatic sections as well as of an intercalary and a terminal heterochromatic segment. The present results strongly suggest that the Ks of A. lucidus are derived from the Ss by rearrangements and by formation and accumulation of repetitive sequences.

The kinetic behaviour of the cranial neural epithelium during neurulation in the rat

The kinetic behaviour of the cranial neuroepithelial cells of rat embryos during neurulation is described. Serial transverse sections of 4-, 8-, 12- and 16-somite-stage embryos show that differential mitosis does not play a part in the mechanisms responsible for effecting cranial neural tube closure. A constant cell number is found in the midbrain/hindbrain neural epithelium during all four stages; the mitotic spindle axes are oriented parallel to the long axis of the embryo, so that increase in cell number occurs in this direction only. Growth is only expressed by an expansion in the volume of the forebrain, which projects rostral to the notochordal tip. [3H]thymidine studies (using an in vitro culture technique) show no significant variation in the cell cycle time between the forebrain and the midbrain/anterior hindbrain neural epithelium. It is suggested that the neural epithelium is a fluid structure whose overall shape is strictly controlled while the cells within it flow towards and into the rapidly expanding forebrain.

Cinemicrographic study of cell proliferation pattern and interdivision times of human keratinocytes in primary culture.

The stratified squamous epithelium of the skin, the epidermis, is a renewing cell population. In order for epidermis to remain the same size, each dividing basal cell must produce on an average, one daughter cell that will remain as a germinative cell and another that will perform terminal differentiation (differential mitosis). In order to investigate cell kinetics in the epidermis, the information obtained from in vivo study is limited, and offers only indirect evidence for the determination of cell cycle time and cell proliferation pattern. Keratinocytes in cell culture are unique in formation of a multilayered cellular sheet in which the keratinocytes form a structure resembling the epidermis in vivo, and keratinize at the top. In the early days of the primary culture of human keratinocytes, when the cells proliferate to form a monolayer sheet, direct access to cell proliferation pattern and measurement of interdivision time can be done using techniques of time-lapse cinemicrography. The primary cultures at 8-20 days of incubation were employed for cinemicrographic observation when small polygonal cells appeared in groups, when numerous mitotic figures were observed, and when stratification of the cells had not yet occurred. The appropriate field was marked and followed for up to 6 days. Photographic prints were made from the 16 mm cine film, and dendrograms were made and analyzed for pattern of cell proliferation and interdivision time. Most cells in the field divided two or three times during the period of observation. Sister-sister pairs of the second and third generations divided after approximately the same interdivision times. However, some cells have never divided. Some of the sister-sister pairs differed considerably in their interdivision times. In some cultures synchronous division was quite evident. The average interdivision time was about 26 hr in the majority of cultures, and it is suggested that the estimated long cell cycle time in vivo might be overestimated due to the existence of non-cycling cells in the germinative population.

Differential mitosis and degeneration patterns in relation to the alterations in the shape of the embryonic ectoderm of early post-implantation mouse embryos

The shape of the embryonic ectoderm of early post-implantation mouse embryos changes greatly in the period of 6.2-7.3 days post coitum. The subcellular morphology of the embryonic ectoderm remains unchanged, except in the primitive-streak region. Cell kinetics differ between ectodermal regions. These differences may be related to the changes in the shape of the ectoderm. The increase in cell number in the lateral ectoderm (the prospective surface ectoderm) exceeds that in the frontal ectoderm (the future neurectoderm). This is not due to differences in the duration of the cell cycle. It can be explained, however, by the occurence of different relative numbers of dividing and non-dividing cells. These numbers vary between the two regions. The percentage of non-dividing cells in the frontal ectoderm may reach 45, whereas in the lateral ectoderm this percentage is not higher than 15. Autoradiography in tritiated thymidine-treated embryos combined with the mitotic indices gave us all of the parameters necessary to present a model capable of clarifying the growth of the ectoderm during gastrulation, as well as the changes in the shape of the ectoderm.

Clonal segregation and positional information in late ommatidial development in Drosophila.

CELLULAR DIVERSITY in an organ might arise from differential mitoses within a prefixed, genetically determined cell lineage or through the reading of positional values by equipotent cells in a morphogenetic field1. We have studied this problem in the developing eye imaginal disc of Drosophilia, in particular in the cells giving rise to the eight photoreceptor (R) and the five secondary pigment (SP) cells of the adult ommatidium2,3. These cells lie close together within the ommatidium and are easily distinguishable with morphological criteria. We asked the following question: do R and SP cells have common precursors at the latest developmental stage or do they belong to different cell lineages?

Regions of Differential Cell Elongation and Mitosis, and Root Meristem Morphology in Different Tissues of Geotropically Stimulated Maize Root Apices

We examined cell length, mitosis, and root meristem "cuticle" in different tissues of geostimulated, red light-exposed primary roots of corn (Zea Mays, Wisconsin hybrid 64A x 22R). The examination was done at 15-minute intervals for a period of 240 minutes. Differences in cell elongation between the upper and lower sides were most prominent between 1.5 and 2.5 mm from the root meristem; the outer cortex had the greatest elongation growth, and the upper cells showed a significant increase in length compared to the lower. A differential mitosis was also found, with the lower tissue being greater. We infer that the mitotic activity is indicative of cell division, and this division occurs strictly in the first 1.5 mm of the root meristem. The combined effect of differential cell elongation and cell division results in the localization of the geotropic curvature in the 1.5- to 2.5-mm region from the root meristem. Mitosis that occurs primarily in the cortex and stele were asynchronous; the peak of cortical division preceded that of the stele. Both peaks occurred before the peak of geotropism. A densely stained layer separates the cap from the root meristem. This layer is thinner at the apex of the root meristem. The area of the thin region increased with time and peaked at 180 minutes after geostimulation, which was coincidental with the peak of the geotropic response.

Spatial distribution of DNA-synthesizing cells in colonic crypts of the guinea pig.

The ascending colon of a guinea pig injected with tritiated thymidine was cut serially, autoradiographed and stained with periodic-acid Schiff-hematoxylin. Maps of transversely sectioned crypts were prepared with the use of a microscope eye-piece projector. The number and angular positions of pulselabelled (DNA-synthesizing) cells around the circumference of transverse sections of the crypt were recorded. A method of 'statistics of the circumference' was applied in order to find the variances of angular distances between labelled cells and thereby to find the type of arrangement of DNA-synthesizinbg cells in the crypt. The spatial distribution of DNA-synthesizing cells both around the crypt circumference and along the crypt, was found to be non-random. While the pattern of non-randommess around the crypt circumference is such that the DNA-synthesizing cells tend to occupy positions in the crypt circumference at maximal distances from each other, DNA-synthesizing cells along the crypt tend to occupy positions at minimal distances from each other. DNA-synthesizing cells are arranged in the crypt in rows, each consisting of several cells and each parallel to the long axis of the crypt. Apparently the dividing cell of the crypt produces either two proliferating or two differentiating cells. No evidence of differential mitosis could be found.

Extrachromosomal DNA and the origin of oocytes in the telotrophic-meroistic ovary ofCreophilus maxillosus (L.) (Staphylinidae, Coleoptera-Polyphaga).

The development of the telotrophic ovary in the Staphylinid beetle,Creophilus maxillosus was examined. Cells, termed chordoblasts were identified in the germarium of 1-day-old pupae. Each of the chordoblasts undergoes a series of synchronous mitoses. Owing to the precise control of the cleavage plane, which is vertical to the long axis of the ovariole, each of the chordoblasts gives rise to a linear chain of sibling chordocytes. Extra DNA synthesis within each sibling string is usually limited to the most posterior chordocyte only, this being an oocyte progenitor.Divisions of the oocyte progenitor are differential mitoses in which the extra DNA material is transported preferentially towards the posterior pole of the spindle. As extra DNA synthesis and preferential segregation of this material result in gradual increase of this DNA in the nuclei of oocyte progenitors, cytokinesis of these cells becomes highly unequal, the larger of the two cells produced at each differential mitosis being as a rule the posterior cell, i.e. the oocyte progenitor of the next cell generation. As a resul of the series of differential mitoses each chordoblast gives rise to a number of nurse cells and only one definitive oocyte.It is suggested that somatic prefollicular tissue plays a decisive role in oocyte determination in the Coleopteran telotrophic ovary.

An in vitro Analogue of Early Chick Limb Bud Outgrowth

Our culture system appears to represent an in vitro analogue of early chick limb morphogenesis. Organized mesodermal cell accumulations resembling limb buds were derived from a monolayer of limb mesoderm cells when covered by limb ectoderm which included the apical ectodermal ridge (AER). The ridge retained its normal configuration when grown over a limb mesoderm monolayer and the mesoderm cells accumulated under the ridge to form a multilayered structure (10-25 cells in thickness) with the characteristic shape of a limb bud. Ectoderm which did not include the ridge failed to promote the formation of limb-like mesodermal accumulations thus the action of the ridge appears to be specific. The AER-elicited expression of mesodermal cell behaviour leading to early limb outgrowth is discussed in terms of possible morphogenetic mechanisms involved i.e. differential mitosis, cell migration, changes in cell shape and especially the adhesive properties of the cells.

Chapter 4 The Cocoonase Zymogen Cells of Silk Moths: A Model of Terminal Cell Differentiation For Specific Protein Synthesis

Publisher Summary This chapter discusses the zymogen cell differentiation for specific protein synthesis. Development of terminally differentiated cells specialized for specific protein synthesis involves the sequential and coordinated occurrence of several steps. Some of these are: differential mitosis, morphogenesis, mitosis or polyploidization, elaboration of an appropriate cytoplasm, synthesis of the specific cell product at a low and then high rate, and finally turning off or the modulation of specific synthesis. Continuous increase in specific protein synthesis during phase I and II is quite significant. If the specific gene is very active in transcription, differential stability would lead to mRNA accumulation. Thus, the stability of the mRNA can account for the marked progressive increase in translation, within phase II, without the necessity of postulating a parallel increase in the transcriptional activity. The absolute increase in specific protein synthesis during phase I is much slower, suggesting that accumulation and perhaps specific gene transcription is occurring more slowly at that time. Assuming that protein synthesis is proportional to mRNA content, the increase in protein synthesis during phase II measures the rate, at which specific mRNA accumulates in the system. This rate of accumulation is compatible with the existence of a single copy of the differentiation-specific gene per genome.