Formation of the paratomic fission zone in freshwater oligochaetes

Oligochaetes Nais communis and Pristina longiseta are capable of paratomy, i.e., asexual reproduction of the cross division type, when division proceeds across the long body axis and daughter organisms retain the maternal axes. Paratomy is represented by two forms: slow and rapid. Slow paratomy is accompanied by the formation of chains from no more than two zooids (N. communis), while rapid paratomy leads to the formation of chains from many zooids (P. longiseta). Analysis of the appearance and development of the zone of paratomy (constriction) has shown that when its cephalogenic part is formed, only head segments appear and are formed simultaneously, while in the somatogenic part, trunk segments appears successively. Dedifferentiated cells of the integumental epithelium appear to serve as a source of newly formed structures. It was shown using mathematical statistics that constriction is laid down in the zone defined by a system of coordinates formed by unknown factors. Although the constriction is laid down in the middle of a segment, its subsequent growth is not related to the transformation of adjacent areas of the segment.     

Quantitative observations on asexual reproduction of Aeolosoma viride (Annelida,Aphanoneura)

Agametic reproductive activity (via paratomy) of Aeolosoma viride was analyzed throughout the life cycle in individually reared specimens. Aeolosoma viride is organized in linear chains of 3–4 zooids; the main zooid is anterior, and the secondary zooids are positioned posterior to the main zooid in inverse order with respect to their degree of growth, the most advanced being at the posterior end, and those less advanced nearer the main zooid. On average, worms lived 66±10 d and produced 57±6 offspring. A budding area located in the sub‐terminal part of the main zooid produced chaetigers that formed the origin of the secondary zooids. A growth zone was located in the posterior end of each secondary zooids. Fission occurred between the penultimate and the last zooid of the chain. Just before fission, the growth zone of each secondary zooid became a budding area. Agametic reproduction was via multiple paratomy with linear succession of the secondary zooid and terminal fission. The structure of the chain was therefore modulated by the interaction of the processes of budding, growth, cephalic differentiation, and fission, which occurred continuously and on different timescales. Values of parameters describing paratomic activity (interval between origin of the zooids, time to produce a chaetiger, growth time of the zooids, and interval between the fission of the filial chains) are low early in an individual's life, but increase during senescence. Due to its relatively rapid lifecycle and high reproductive activity, A. viride is a convenient experimental organism for the study of agametic reproduction.     

Growth patterns during segmentation in the two polychaete annelids, Capitella sp. I and Hydroides elegans: comparisons at distinct life history stages

Many animals generate new body segments sequentially from a posterior growth zone, and this is generally thought to be the case for the annelids. Most annelids, including polychaetes, have an indirect life cycle and generate their earliest segments during larval life. We have characterized the nature of the growth zone in two polychaetes, Hydroides elegans and Capitella sp. I, during both larval and juvenile stages of segment formation by examining cell division patterns with 5-bromo-2'-deoxyuridine incorporation. Cell division patterns show commonalities between the two species, even though they have distinct body plans and life history characteristics. In both polychaetes, larval segments arise from a field of dividing cells located in lateral regions of the body, rather than from a localized posterior growth zone. Circumferential expansion of the forming segmental tissue is particularly pronounced in Capitella sp. I. Post-metamorphic segments, in contrast, originate from a classical posterior growth zone, with the exception of four posterior thoracic segments of H. elegans, which appear to arise from an area in the middle of the body, indicating plasticity of segment-generating mechanisms present in different annelid life histories. The distinct nature of larval versus juvenile growth zones in H. elegans and Capitella sp. I raises the question of the mechanistic relationship between these two growth zones. The results of this study increase our understanding of the cellular origins of segments in annelids, and serve as a basis for interpretation of molecular expression patterns associated with segment formation in polychaetes.     

The ventral nerve cord signals positional information during segment formation in an annelid (Ophryotrocha puerilis,Polychaeta)

Summary Growth and regeneration of segments were recorded in the polychaeteOphryotrocha puerilis. In one experiment the ventral nerve cords (VNCs) of the animals were cut; in the other, VNCs were left intact. VNC lesion in some specimens resulted in the outgrowth of supernumerary posterior parts from the site of operation. The characteristics of outgrowth of these supernumeraries were essentially the same as in normal specimens without double tails. After removing different numbers of caudal setigers, each of the two tails of the same double-tail monster independently regenerated different segment numbers within a given time. A simple model is proposed, allowing for these results, which states that the larval body of a polychaete consists of two regions with completely different positional values (episphere — prostomium; hyposphere — pygidium). During growth, segments with intervening positional values are intercalated. The rate of segment formation is high when there is a wide gap in positional values between pygidium and adjoining budding zone and the posteriormost segment. As this gap narrows, the growth rate slows down. During caudal regeneration, first of all a new pygidium with an adjacent proliferation zone is formed and the original positional value of the posteriormost part of the body is reestablished. Segment regeneration follows the same rules as segment growth. The results presented here also demonstrate that the VNC plays an important role, not only in segment proliferation, but also in signalling positional information to the newly formed segments.     

The specification of metameric order in the insectCallosobruchus maculatus Fabr. (Coleoptera)

Summary Mechanically dividing an insect egg into anterior and posterior fragments results in a segment gap (Sander 1976), a loss of non-terminal segments in the constricted region. By altering the stage and duration of constriction, we produced different types of egg fragments in the pea beetleCallosobruchus. The patterns formed by these fragments suggest the existence of interactions between anterior and posterior egg regions that influence segment patterning and placement. Segments in excess of the numbers expected on the basis of permanent constrictions were produced in fragments when: (1) the constriction was released before cellularization occurred and (2) in addition the complementary fragment degenerated. Apparently the degenerating fragment induced the formation of excess segments in the developing fragment. Differences in the time and extent of excess segment formation in anterior versus posterior fragments suggest an asymmetric distribution of prerequisites for segment formation. This conclusion is consistent with our finding that a partial reversal of segment sequence (double abdomen formation) can be induced only in posterior fragments by a degenerating fragment, but not in anterior fragments (see companion paper).The formation of excess segments shows that the segment gap observed after permanent separation cannot be due to non-specific damage, caused by the process of constriction as such, to the egg or to localized putative segment precursors.     

THE FINE STRUCTURE OF MITOSIS AND CELL DIVISION IN THE CHRYSOPHYCEAN ALGA OCHROMONAS DANICA1

At the ultrastructural level, cell division in Ochromonas danica exhibits several unusual features. During interphase, the basal bodies of the 2 flagella replicate and the chloroplast divides by constriction between its 2 lobes. The rhizoplast, which is a fibrous striated root attached to the basal body of the long flagellum, extends under the Golgi body to the surface of the nucleus in interphase cells. During proprophase, the Golgi body replicates, apparently by division, and a daughter rhizoplast, appears. During prophase, the 2 pairs of flagellar basal bodies, each with their accompanying rhizoplast and Golgi body, begin to separate. Three or 4 flagella are already present at this stage. At the same time, there is a proliferation of microtubules outside the nuclear envelope. Gaps then appear in the nuclear envelope, admitting the microtubules into the nucleus, where they form a spindle. A unique feature of mitosis in O. danica is that the 2 rhizoplasts form the poles of the spindle, spindle microtubules inserting directly onto the rhizoplasts. Some of the spindle microtubules extend from pole to pole; others appear to attach to the chromosomes. Kinetochores, however, are not present. The nuclear envelope breaks down, except, in the regions adjacent, to the chloroplasts; chloroplast ER remains intact throughout mitosis. At late anaphase the chromosomes come to lie against part of the chloroplast ER. This segment of the chloroplast ER appears to be incorporated as part of the reforming nuclear envelope, thus reestablishing the characteristic nuclear envelope—chloroplast ER association of the interphase cell.     

Notch/Delta signalling is not required for segment generation in the basally branching insect Gryllus bimaculatus

Arthropods and vertebrates display a segmental body organisation along all or part of the anterior-posterior axis. Whether this reflects a shared, ancestral developmental genetic mechanism for segmentation is uncertain. In vertebrates, segments are formed sequentially by a segmentation 'clock' of oscillating gene expression involving Notch pathway components. Recent studies in spiders and basal insects have suggested that segmentation in these arthropods also involves Notch-based signalling. These observations have been interpreted as evidence for a shared, ancestral gene network for insect, arthropod and bilaterian segmentation. However, because this pathway can play multiple roles in development, elucidating the specific requirements for Notch signalling is important for understanding the ancestry of segmentation. Here we show that Delta, a ligand of the Notch pathway, is not required for segment formation in the cricket Gryllus bimaculatus, which retains ancestral characteristics of arthropod embryogenesis. Segment patterning genes are expressed before Delta in abdominal segments, and Delta expression does not oscillate in the pre-segmental region or in formed segments. Instead, Delta is required for neuroectoderm and mesectoderm formation; embryos missing these tissues are developmentally delayed and show defects in segment morphology but normal segment number. Thus, what initially appear to be 'segmentation phenotypes' can in fact be due to developmental delays and cell specification errors. Our data do not support an essential or ancestral role of Notch signalling in segment generation across the arthropods, and show that the pleiotropy of the Notch pathway can confound speculation on possible segmentation mechanisms in the last common bilaterian ancestor.     

Developmental pattern of colonies in the polystyelid ascidian,Polyandrocarpa misakiensis

Summary

The growth pattern of zooids formed asexually by budding was studied in the colonial ascidian, Polyandrocarpa misakiensis. Each colony started from a blas- tozooid (the first generation) on the glass plate in two series of experiments. To evaluate the growth of colonies, lineage of all the zooids of three successive generations was traced on photographs which were taken once a week. The zooids of the first generation produced many buds from any basal margin of the zooidal body, and those of the second generation produced a small number of buds mainly from anterior parts of the zooidal body. The zooids of the second generation produced by early budding of mother zooids were clearly more prolific than those produced by late budding. Circular colonies which developed around a zooid of the first generation consisted of stratified zones of successive generations. Each zone was composed of two subzones; the outer one mainly containing early-produced zooids, and the inner one mainly containing late-produced zooids. The zooids in the marginal area of colony are early-produced ones from generation to generation. The seawater temperature may influence the growth of zooids and/or the frequency of budding.     

A study of regression and budding in Perophora viridis

1. A method has been devised for studying the regression of the zooid of Perophora into a stolon and the subsequent differentiation of a new zooid from this stolon. 2. Circulatory cells of the stolon resulting from regression will aggregate into masses larger than the minimal size necessary for differentiation of a zooid, but fail to differentiate into a zooid. 3. The cells of a zooid after staining with neutral red appear in the stolon during regression and finally come to lie in the newly formed zooid. 4. During the cycle of adult zooid to stolon to newly formed zooid, there is no evidence for cell division from studies with tritiated thymidine. 5. It is concluded that under conditions of starvation, an adult zooid furnishes all the cells for the formation of a stolon and the subsequent zooids without cell division.     

Peristaltic waves of tubicolous worms and the problem of irrigation in Sabella pavonina

A simple scheme is presented to illustrate four possible kinds of locomotory peristalsis in worm-like animals. The application of this scheme to real animals is discussed. Peristaltic waves may be of constriction or dilatation. A continuous body cavity enables the worm to regulate both speed and direction of travel by controlling the relative tonus of its body wall muscles. Thus peristaltic waves can be used to pump water without causing locomotion.
Sabella irrigates its tube by peristaltic swellings but the coelom and intestine are sub-divided by entire septa. Anatomical and morphological features which allow the shortest, widest segments forming a "piston" to slide down the tube and the narrower elongated segments to grip its walls are considered. In this way the construction of the typical body segment is given a functional explanation.
The functions of septa in annelids are discussed.     

Distribution Pattern of Cell Division Centers on the Epidermis of Stem Segments of Torenia fournieri during de Novo Bud Formation

The stem epidermis in Torenia fournieri, which has budding potentialialities, is composed of one cell layer which can be easily separated from the rest of the stem segment at different stages of bud formation. As the buds are formed directly from the epidermis, without intermediate callus formation, it is possible to observe simultaneously the cell division centers over the entire excised epidermal surface. The quantitative analysis at the 6-day stage of bud formation showed that the cell division centers do not have a random distribution on the epidermal surface. With respect to the length of the stem segment, the frequency of cell division centers increases toward the base which is also the direction of auxin transport. With respect to the width, the maximum number of division centers is observed on either side of the median zone. The median zone and the lateral zones have few division centers. An anatomical study showed that the zones with few division centers are the closest to underlying vascular tissue. A more uniform distribution of division centers can be obtained by addition of auxin to the medium.     

Conserved relationship between FtsZ and peptidoglycan in the cyanelles of Cyanophora paradoxa similar to that in bacterial cell division

Cyanelles of the biflagellate protist Cyanophora paradoxa have retained the peptidoglycan layer, which is critical for division, as indicated by the inhibitory effects of β-lactam antibiotics. An FtsZ ring is formed at the division site during cyanelle division. We used immunofluorescence microscopy to observe the process of FtsZ ring formation, which is expected to lead cyanelle division, and demonstrated that an FtsZ arc and a split FtsZ ring emerge during the early and late stages of cyanelle division, respectively. We used an anti-FtsZ antibody to observe cyanelle FtsZ rings. We observed bright, ring-shaped fluorescence of FtsZ in cyanelles. Cyanelles were kidney-shaped shortly after division. Fluorescence indicated that FtsZ did not surround the division plane at an early stage of division, but rather formed an FtsZ arc localized at the constriction site. The constriction spread around the cyanelle, which gradually became dumbbell shaped. After the envelope’s invagination, the ring split parallel to the cyanelle division plane without disappearing. Treatment of C. paradoxa cells with ampicillin, a β-lactam antibiotic, resulted in spherical cyanelles with an FtsZ arc or ring on the division plane. Transmission electron microscopy of the ampicillin-treated cyanelle envelope membrane revealed that the surface was not smooth. Thus, the inhibition of peptidoglycan synthesis by ampicillin causes the inhibition of septum formation and a marked delay in constriction development. The formation of the FtsZ arc and FtsZ ring is the earliest sign of cyanelle division, followed by constriction and septum formation.     

Postembryonic development of Antygomonas incomitata (Kinorhyncha: Cyclorhagida)

Postembryonic development in the kinorhynch species Antygomonas incomitata was examined using scanning electron microscopy. The morphology of the six juvenile stages, J‐1 to J‐6, varies at numerous details, but they can also be distinguished by a few key characters. Juvenile stage 1 by its composition of only nine trunk segments; J‐2 by the combination of possessing 10 trunk segments, but no cuspidate spines on segment 9; J‐3 by the presence of cuspidate spines on segment 9, but only one pair of cuspidate spines on segment 8; J‐4 by the combination of 10 trunk segments only, but having two pairs of cuspidate spines on segment 8; J‐5 by possessing 11 trunk segments and same spine compositions as adults but is still maintaining postmarginal spiculae; J‐6 specimens closely resemble adults and are most easily identified by their reduced trunk lengths. New segments are formed in a growth zone in the anterior part of the terminal segment. The complete number of segments is reached in J‐5. Development of cuticular head and trunk structures are described through all postembryonic stages and following developmental patterns could be outlined: the mouth cone possesses outer oral styles from J‐1, but in J‐1 to J‐3, the styles alternate in size. Scalids of the introvert are added after each molt, and scalids appear earliest in the anterior rings, whereas scalids in more posterior rings are added in older postembryonic stages. The early J‐1 stage is poor in spines and sensory spots and both structures increase in number after each molt. The complete spine composition is reached in J‐4, whereas new sensory spots appear after all molts, inclusive the final one from J‐6 to adult. Sensory spots in the paraventral positions often appear as Type 3 sensory spots but are through development transformed to Type 2. This transformation happens earliest on the anterior segments. J. Morphol., 2010. © 2010 Wiley‐Liss, Inc.     

Purification and immunofluorescence localization of the mutant gene product of a Tetrahymena cdaA1 mutant affecting cell division

A division arrest mutant, cdaA, of Tetrahymena thermophila is known to have a ts-defect in the formation of the fission zone which determines the position of the fission plane. A protein (Mr = 85,000; pI = 4.7, designated as p85) has recently been identified in our laboratory as a possible gene product of the cdaA locus by two-dimensional gel electrophoresis and genetic experiments (Ohba et al., submitted). In the present research, we have isolated p85, prepared its antibody, and demonstrated that in wild-type cells or in cdaA cells at permissive temperature, immunofluorescence for p85 appears on the equatorial basal bodies at the predicted fission zone just before formation of the zone. In such a case, the fission zone appears to be formed just anterior to the fluorescence-associated basal bodies, and then constriction of the division furrow occurs at the zone. However, in cdaA cells at the restrictive temperature, the equatorial p85 deposit and subsequent fission zone formation and furrowing do not occur at all. Thus, we conclude that p85 plays a key role in the formation of the fission zone and in the positioning of the equatorial fission line.     

Role of peptidoglycan amidases in the development and morphology of the division septum in Escherichia coli

Escherichia coli contains multiple peptidoglycan-specific hydrolases, but their physiological purposes are poorly understood. Several mutants lacking combinations of hydrolases grow as chains of unseparated cells, indicating that these enzymes help cleave the septum to separate daughter cells after cell division. Here, we confirm previous observations that in the absence of two or more amidases, thickened and dark bands, which we term septal peptidoglycan (SP) rings, appear at division sites in isolated sacculi. The formation of SP rings depends on active cell division, and they apparently represent a cell division structure that accumulates because septal synthesis and hydrolysis are uncoupled. Even though septal constriction was incomplete, SP rings exhibited two properties of mature cell poles: they behaved as though composed of inert peptidoglycan, and they attracted the IcsA protein. Despite not being separated by a completed peptidoglycan wall, adjacent cells in these chains were often compartmentalized by the inner membrane, indicating that cytokinesis could occur in the absence of invagination of the entire cell envelope. Finally, deletion of penicillin-binding protein 5 from amidase mutants exacerbated the formation of twisted chains, producing numerous cells having septa with abnormal placements and geometries. The results suggest that the amidases are necessary for continued peptidoglycan synthesis during cell division, that their activities help create a septum having the appropriate geometry, and that they may contribute to the development of inert peptidoglycan.     

ORIGIN AND DEVELOPMENT OF POLLEN EMBRYOIDS AND POLLEN CALLUSES IN CULTURED ANTHER SEGMENTS OF HYOSCYAMUS NIGER (HENBANE)

The dedifferentiation of pollen grains of Hyoscyamus niger (henbane) into embryoids and calluses was examined by culturing identical segments of the same anther in a mineral salt-sucrose basal medium and in the basal medium supplemented with 2.0 mg/l 2,4-dichlorophenoxyacetic acid, respectively. Addition of auxin enhanced anther efficiency but did not affect the number of embryogenic pollen grains of an anther segment transformed into calluses. In anther segments cultured in the basal medium, the organogenetic part of the pollen embryoid was formed by the division of the generative cell alone, or by the division of both generative and vegetative cells. More or less similar pathways were followed by pollen grains of anther segments cultured in a medium containing auxin to form calluses. Culture of anther segments in a medium containing a high concentration of auxin (50.0 mg/l) led to a significant reduction in the yield of calluses which were formed almost entirely by the division of both generative and vegetative cells. The bearing of these observations on the role of auxin in determining the pathway of differentiation of embryogenic pollen grains in cultured anther segments is considered. The appearance of embryogenic pollen grains in close proximity to the tapetum as seen in longitudinal sections of cultured anther segments has suggested a role for a gradient of tapetal factors in embryogenic induction.     

Demarcation of the cortical division zone in dividing plant cells

Somatic cytokinesis in higher plants involves, besides the actual construction of a new cell wall, also the determination of a division zone. Several proteins have been shown to play a part in the mechanism that somatic plant cells use to control the positioning of the new cell wall. Plant cells determine the division zone at an early stage of cell division and use a transient microtubular structure, the preprophase band (PPB), during this process. The PPB is formed at the division zone, leaving behind a mark that during cytokinesis is utilized by the phragmoplast to guide the expanding cell plate toward the correct cortical insertion site. This review discusses old and new observations with regard to mechanisms implicated in the orientation of cell division and determination of a cortical division zone.     

Replication of DNA in mammalian chromosomes

DNA replication has been studied in cells (CHO) synchronized by mitotic selection from roller cultures. A study of the incorporation of ³H supplied as uridine indicates that cells cannot be blocked precisely at the beginning of the S phase, but DNA synthesis can be stopped in early S by treating with F-dU in G₁. After blockage potential initiation sites continue to increase at a linear rate for atleast 13 hours after division. Incorporation of ³H-thymidine begins at most of these sites within seconds after thymidine is supplied in the medium and incorporation continues at a linear rate for 20–24 minutes. There appears to be a pause after this interval before synthesis is resumed at about two times the initial rate. ³H-bromodeoxyuridine can be substituted for thymidine without affecting the kinetic pattern over a similar period. The increased rate is probably an increase in sites of chain growth rather than a change in rate of chain growth. A study of the labeled DNA segments by band sedimentation in a preformed NaClO₄ isokinetic gradient shows that two distinctly different sized segments can be released from the chromosomes by lysis at submelting conditions. One is the previously reported single chain segments averaging about one-half micron in length, but the other is a much larger segment (26S) which is native DNA with perhaps small regions of single chains presumably at the ends. Primarily single chain DNA is released after 1–2 minute pulse labeling, but after 2 minutes the larger segments (26S) contain most of the newly formed DNA except that attached to the chains of the major part of the template DNA which exhibits a discontinuous distribution, sedimenting far faster than either newly replicated segment. A consideration of the kinetics of formation of the 26S component indicates that is may contain the replicating fork. If this proves to be the correct interpretation the template chains would both have non-adjacent nicks preceeding the fork and also in a post-fork site at a mean distance of about 2 microns in both directions. The isolation of the growing points of DNA replication in chromosomes is now possible and the study of properties of the newly replicated regions should be greatly facilitated.     

Asexual reproduction in Microstomum lineare (Turbellaria). I. An autoradiographic and ultrastructural study

The distribution of cells preparing for proliferation during asexual reproduction by paratomy in Microstomum lineare (Turbellaria) was investigated using the technique of exogenous tritiated thymidine ([³H]T) labelling and routine electron microscopy. The subsequent fates of labelled cells in developing and mature zooids were followed by fixing tissues at various intervals. The only labelled cells are mesenchymal and gastrodermal neoblasts, occurring along the whole worm without any special growth zones or axial gradient. Organ primordia develop as a result of locally dividing as well as of migrating cells.     

Mitosis, Cytokinesis and Colony Formation in Pediastrum boryanum

Uninucleate cells of Pediastrum become multinucleate by a seriesof synchronous mitoses. Mitotic nuclei are enclosed by a perinuclearenvelope of endoplasmic reticulum. Cytoplasmic cleavage of themultinucleate cells leads to the production of uninucleate,biflagellate zoospores (zooids) which are subsequently releasedinto a lenticular vesicle through a rupture in the outer layerof the parental cell wall. Within the vesicle, presumably derivedfrom part of the inner layer of parental wall, the zooids swarmactively before aggregating in a planar array. Bands of microtubulesunderlie the plasmalemma of the zooids which, when the zooidsaggregate, are usually coplanar with the newly formed colony.The role of microtubules in patterned colony formation and inthe development of the characteristic horns on peripheral cellsof colonies of Pediastrum is discussed.