Posterior Daughter Cell Research Articles

Development of surface pattern during division in Paramecium: III. Study of stomatogenesis in the wild type using antitubulin antibodies and confocal microscopy

Summary Stomatogenesis, during the vegetative division of Paramecium, is a complex morphogenetic process involving massive proliferation of basal bodies and their progressive patterning to generate a new oral apparatus in the immediate vicinity of the old one. This new oral apparatus will be inherited by the posterior daughter cell while the old one remains in the anterior daughter cell. We have carried out a detailed description of the whole process, using new antibodies and confocal microscopy, allowing visualization of all basal bodies and a number of other cytoskeletal elements, thus completing our previous description of morphogenesis. The following inferences could be made: 1) The new oral structures are exclusively formed from the anarchic field by a two step wave of basal body duplication. These two steps strictly parallel, slightly in advance, those we have previously detailed at the level of the “somatic” cortex, suggesting *) that a common set of biochemical cascades is involved, **) that the OA may be triggering these cascades. 2) The initial proliferation of basal bodies occurs in the immediate vicinity of the parental “paroral kinety” which itself remains invariant and appears to dictate the progressive patterning of the oral anlage (as can be inferred from the staining intensity, distance and alignment of the basal bodies of the anarchic field with respect to the paroral kinety). This suggests that the concept of cytotaxis or structural guidance can be extended to the genesis of an elaborate set of basal bodies. 3) The number of basal bodies within the anarchic field is determined during this formation of the new oral apparatus. Thus, the size and possibly part of the pattern of the new OA is determined in one generation, with storage of morphogenetic potentialities used in the next generation. In this process, a slight cortical rotation around the cell axis is effected at each division. 4) The whole morphogenetic assemblage up to the level of the whole cell displays an overall left-right asymmetry which is progressively built up upon the basic asymmetry of the basal body.

The influence of fission line expression on the number and positioning of oral primordia in the cdaA1 mutant of Tetrahymena thermophila.

During cytokinesis, furrowing creates new boundaries for daughter cells. Following a shift to a restrictive temperature, cells of the temperature-sensitive cell-division-arrest (cdaA1) mutant of Tetrahymena thermophila complete development of the oral apparatus for the prospective posterior daughter cell before becoming arrested in cytokinesis. When maintained under weak restrictive conditions (35 degrees C), some of the chains were arrested prior to the start of fission line formation (D-shaped chains), whereas others manifested rudimentary unilateral furrowing on the ventral side (B-shaped chains). In their second cell cycle following the temperature shift, the D-shaped chains usually formed only one oral primordium, at a position highly correlated with the length of the entire chain. The B-shaped chains always produced two separate oral primordia, located at irregular positions anterior and posterior to the division furrow, often close to the posterior oral apparatus produced during the first cycle. These results suggest that the formation of the fission line sets a reference boundary to assess the number of oral primordia and influence their position, that appear during subsequent morphogenetic episodes. They also indicate that, during cell division cycles, pre-existing oral apparatuses do not strongly inhibit the formation of new oral apparatuses in their close vicinity.

Cytoplasmic streaming direction reverses in dividing Paramecium bursaria

Using the interference-contrast videomicroscopy the speed of cytoplasmic streaming was measured during the sequence of division stages in thigmotactically settled specimens of Paramecium bursaria. The speed of cytoplasmic flow gradually decreased during the first stages of binary fission and movement became indistinguishable at stage D(3). Almost at the same time cytoplasm started to move in the opposite direction, pushing or pulling the dividing micronucleus into the prospective posterior daughter cell and eventually stopped at stage D(5)-D(6). Further cell division events proceeded without detectable movement of cytoplasmic components. Cytoplasmic streaming in the normal interphase route was gradually restored in daughter cells about 30-40 min after cell separation. During the whole period of binary fission phagocytosis was arrested. Transportation and participation in the positioning of prospective micronuclei in daughter cells seems to be the main function of cytoplasmic streaming activity in cell division of Paramecium bursaria. The possible relationship between the stages of cytoskeleton transitions and the kinetics of cytoplasmic streaming associated with cell divison is discussed.

Asymmetrical cell division in Blepharisma japonicum: Difference between daughter cells in mating-type expression

In cell division of high-frequency-selfers in the ciliate Blepharisma japonicum, daughter cells are different in mating-type expression. The anterior daughter cell is mating type I. The posterior daughter cell is mating type II at first and then changes to mating type I after about 24 h. The anteroposterior polarity of predivision cells appears to correlate with the asymmetrical cell division. This work introduces a unicellular organism about the size of microscopic metazoa as a model system for the study of asymmetrical cell division, which is particularly important in developmental processes.

Vergleichende morphogenetische untersuchung von 13 arten der gattung Euplotes (ciliophora, hypotrichida)

Summary The morphogenesis of 13 species of the genus Euplotes (E. aediculatus, E. charon, E. crassus, E. daidaleos, E. eurystomus, E. minuta, E. octocarinatus, E. patella, E. raikovi, E. rariseta, E. vannus, E. finki and E. woodruffi) was investigated in the expectation to use morphogenetic criteria for discrimination of species. Six stages of the division process which occur during cortical and nuclear fission were analysed and compared. The morphogenetic events are very similar in all examinated species. Thus, they can not be used to discriminate species. The morphogenesis starts with a proliferation of the basal bodies for the oral primordium closely behind the adoral zone of membranelles of the parental cell. From the developing oral primordium evolve the adoral membranelles and the paroral membrane of the opisthe. The frontoventral and transverse cirri differentiate from anterior and posterior sets each of 5 kinetosomal streaks (Anlagen I–V). Within ciliary outgrowth in cirrotype-9-species a 3:3:3:2:2 pattern of cirral fields is found; in cirrotype-10-species the new cirri develop in the ratio 3:3:3:3:2 always from left to right across the fields of the anterior and posterior sets. The development of the dorsal kineties proceeds according to type 7 and shows no special features. New left caudal cirri for the anterior and posterior daughter cell generate from small linear arrangements of basal bodies on the ventral surface between the developing membranellar band and the left cell margin. On the other hand, the new right caudal cirri develop within the two last kineties on the dorsal surface: for the proter at a level just anterior to the prospective fission line and for the opisthe at the posterior ends of the kineties. The parental adoral membranelles and the paroral membrane are not dedifferentiated during the whole process of morphogenesis, but the parental cirri are completely resorbed.

Stomatogenesis in cyrtophorid ciliates: II. Chilodonella cyprini (Moroff, 1902): The kinetofragment as an anlagen-complex

Non-dividers of Chilodonella cyprini possess two circum-oral kineties and one preoral kinety. These oral kineties are inverted kineties and consist of dikinetids. The cytostome is accompanied by a large microtubular organelle, the cyrtos. The latter consists of an outer ring of nematodesmata and an inner ring of cytopharyngeal microtubular lamellae. In stomatogenesis five somatic kineties are involved. Each of them produces one kinetofragment. These kinetofragments are anlagen-complexes for the oral kineties and the cyrtos. The somatic monokinetid is the elementary unit of stomatogenesis. In front of almost all somatic kinetosomes of a kinetofragment a new kinetosome is assembled, at an angle of 40° to the kinety axis. The somatic cilium disappears and the somatic mother kinetosome transforms into the non-ciliated kinetosome of the oral kinetosomal pair. The newly synthesized anterior daughter kinetosome bears the oral cilium and becomes the ciliated kinetosome of the oral kinetosomal pair. At the anterior end of each of these oral kineties anlagen there are two to four non-ciliated single somatic kinetosomes. Their triplets are covered with electron-dense material. Together with their subkinetal microtubules each of these non-ciliated single somatic kinetosomes forms one nematodesmata anlage. The postciliary microtubules of a part of the non-ciliated kinetosomes of the paired kinetosomes in the oral kineties anlagen and of the non-ciliated single kinetosomes increase in number and align to form one cytopharyngeal lamella anlage. Subsequently all components of the anlagen-complexes separate. Seen from outside the cell the five oral kineties anlagen perform an anti-clockwise circular movement of approximately 180° around the center of the new oral field of the posterior daughter cell, and rearrange to form the circum-oral kineties and the preoral kinety. In the nematodesmata anlagen the nonciliated kinetosome is resorbed. The electron-dense material covering the triplets transforms into the matrix plate of the nematodesma and the subkinetal microtubules increase in number. In the center of the new oral field the nematodesmata anlagen (subkinetal microtubules) and the cytopharyngeal tube lamella anlagen (postciliary microtubules) form the new cyrtos. The anlagen-complexes develop at the anterior ends of the somatic kineties of the posterior daughter cell; the Cyrtophorida show the telokinetal type of stomatogenesis.

Stomatogenesis in Cyrtophorid Ciliates: I. Trithigmostoma steini (Blochmann, 1895): From Somatic Kineties to Oral Kineties

Non-dividers of Trithigmostoma steini possess two or three circum-oral kineties and one preoral kinety. The orientation of these oral kineties is inverted relative to the somatic kineties. The oral kineties consist of dikinetids, the somatic kineties consist of monokinetids. In stomatogenesis the middle postoral kinety becomes the stomatogenic kinety. To produce a posterior daughter cell with three circum-oral kineties and one preoral kinety, six somatic kineties (the stomatogenic kinety included) generate one kinetofragment each. These kinetofragments (named 1 to 6 from right to left) perform an anti-clockwise circular movement (as seen from outside the cell) of about 180° around the center of the oral field anlage. As they move they follow three concentric paths: on the outer circle, kinetofragment 6 migrates first followed by kinetofragments 5 and 4, together they form the outer circum-oral kinety anlage; on the middle circle, kinetofragment 3 moves parallel to kinetofragment 4, forming the middle circum-oral kinety anlage; on the inner circle, kinetofragment 2 moves parallel to kinetofragment 3, forming the inner circum-oral kinety anlage; kinetofragment 1 (produced from the stomatogenic kinety) is the last to move and it becomes the new preoral kinety. The ultrastructural differentiation of somatic fragments to new oral kineties terminates before the start of the circular movement. The somatic monokinetid is the elementary unit of stomatogenesis. The somatic kinetosome transforms into the non-ciliated kinetosome of the oral dikinetid. The somatic cilium disappears and also all somatic associated fibrils, except the postciliary microtubules, which increase in number from four up to seven (in circum-oral kineties) or up to nine (in the preoral kinety). An anterior daughter kinetosome is synthesized in front of each somatic kinetosome, at an angle of 40° to the kinety axis. This new kinetosome becomes the ciliated kinetosome of the oral dikinetid. It bears the oral cilium and a dense transverse fibril at triplet number 3. The oral dikinetids anlagen show the same polarity as the somatic monokinetids did before. The inverted orientation of the oral kineties is due to the circular movement of the oral kineties anlagen after their differentiation.

Division and growth kinetics of the division mutant Conical of Tetrahymena: A contribution to regulation of generation time

The recessive mutant conical of Tetrahymena thermophila is characterized by unequal cytoplasmic division resulting in a large anterior (proter) and a small posterior (opisthe) daughter cell with similar macronuclear DNA contents. Opisthes have long and proters short generation times. This gives the opisthes more time to accumulate cell mass, thereby reducing differences in size between sister cells. Growth rates as determined by cytophotometry do not contribute to the reduction and eventual elimination of differences between sisters but rather should increase them, since small cells accumulate mass at a slower rate than large ones. By tracing consecutive generations it is shown that differences between sister cells in generation time as well as in cell size require more than one generation to be regressed to the mean of the whole population. These findings are incompatible with the probabilistic mode of regulation of generation time.

Cortical control of macronuclear positioning in the ciliate Stentor

AbstractIn the ciliate Stentor, the chain macronucleus has a fixed position within the cell; it is located just beneath the region of narrow cortical striping to the right of the gullet (nuclear site). Stentor therefore lends itself well to experiments on control of nuclear positioning. I have studied this problem by microsurgically producing various abnormalities of nuclear positioning and determining how these are corrected. Breaks in the nuclear chain were repaired by segregation of broken parts into the two daughter cells at division. When a second nucleus was transplanted into a stentor, the abnormality was eventually corrected because the two parallel chains tended to move during oral development (regeneration or division) so that one was anterior to the other. The two nuclei were then segregated into the anterior and posterior daughter cells at division. When a doublet stentor, which has two nuclear sites and two macronuclei, was deprived of one nucleus, it acquired a second one because the remaining nucleus was attracted to both nuclear sites when it elongated during oral development. This phenomenon first produced doublets with a single nuclear chain sharing the two nuclear sites at opposite ends of the cell, a break in the chain then presumably created two nuclei. These latter observations indicate that one mechanism by which the nucleus is positioned involves its attraction toward the nuclear site, a differentiated region of cell cortex, during oral development. Other experiments have shown that the nuclear site also holds the nucleus firmly in place.

Dependence of oral development and cleavage on cell size in the ciliate Stentor

AbstractIn Stentor, the two main cytoplasmic events of cell division are development of the oral primordium that gives rise to the oral apparatus of the posterior daughter cell and cleavage. The present work examines directly the dependence of these two events on the size of the cell body. Cell body size was reduced by various amounts with a microneedle during early stages of cell division and the effects on oral development and cleavage observed. The oral primordium was always resorbed and cleavage never occurred when the cell body was reduced to one quarter of its original size at Stage 3 of cell division; appropriate controls showed that this result was not caused by operation injury. Progressively smaller reductions in cell size yielded correspondingly lower percentages of inhibited cells. These data suggest that the mechanisms underlying the dependence of oral development and cleavage on cell size do not involve accumulation of regulator substances up to a critical concentration during interphase. The fact that oral development can be reversed by reducing cell body size supports the hypothesis that oral development and cell division are initiated when the oral apparatus becomes disproportionately small in relation to cell body size as a result of interphase growth. The size‐dependence of cleavage is probably best understood as a mechanism for regulating cell size in Stentor.

Formation and fate of extranuclear chromatin bodies in Tetrahymena.

The fate of extranuclear chromatin bodies (ECBs) formed by exclusion of macronuclear material at the time of karyokinesis was followed quantitatively in Tetrahymena pyriformis strain GL-I. In a logarithmic growth phase culture, 51% of the dividing cells produced one (43%) or more (8%) ECBs. Most of these gradually disappear before the next cell division, but approximately 13% are retained and carried into subsequent cell cycles. The random distribution of ECBs into anterior or posterior daughter cells, their staining and morphological characteristics, and their rapid loss in cells in starvation medium, all indicate that ECBs play no more of a role in cellular activity than that of an internally produced food vacuole.

Stomatogenic Events Accompanying Binary Fission in Blepharisma*

Stomatogenesis was studied in the heterotrich ciliate Blepharisma japonicum stained with protargol. During binary fission not only is a new oral apparatus made for the posterior daughter, but the already existing oral apparatus of the parent cell is reorganized, i.e., partially disassembled and then subsequently reassembled to provide a functional feeding apparatus for the anterior daughter cell. These morphogenetic events, requiring 2 1/2 to 3 hr, are complete by the time the anterior and posterior daughters separate. In preparation for division, an oral anlage is formed by the rapid proliferation of kinetosomes along 4-5 stomatogenic kinetics directly subtending the cytostome. This field of randomly oriented kinetosomes ultimately gives rise to the feeding apparatus of the posterior daughter cell. Early in division, the oral anlage separates into 2 longitudinal fields of kinetosomes: one is destined to give rise to the undulating membrane and the other forms the adoral zone of membranelles. Shorly after the anlage is established posterior to the cytostome, reorganization of the existing functional mouth is initiated. The morphologic changes associated with this dedifferentiation-redifferentiation sequence lead to the formation of an oral apparatus for the anterior daughter and cannot be distinguished from those characteristically seen during physiologic reorganization.

Further evidence for cortical control over replication of the macronucleus and basal bodies in Stentor

This paper presents further evidence that the cortex controls macronuclear replication and basal body production during the cell cycle of Stentor. At the onset of cell division, basal body production occurs on the ventral side of the cell to form an oral primordium; this structure develops slowly into the oral apparatus destined for the posterior daughter cell. Meanwhile, a series of morphological changes in the macronucleus (coalescence, elongation, nodulation) doubles the number of nodes in preparation for division. When a cell undergoing oral development is grafted to a morphostatic cell of equal size, oral development is usually induced in the morphostatic component and the two members of the graft complex eventually become synchronized with respect to macronuclear morphology. However, primordium induction and nuclear synchronization usually do not occur when the 2 members of the graft complex are separated by cortical discontinuities which heal gradually, even though the graft components demonstrably share a common endoplasm throughout the experiment. These results suggest that the cell surface may control the replication of organelles such as the macronucleus and basal bodies which are normally kept “in step” with the cell cycle in such a way that they are not lost or reproduced too frequently.

X-Ray-Induced Delay of Cell Division in Synchronized Tetrahymena pyriformis

X-Irradiation delays division of heat-synchronized Tetrahymena pyriformis (a ciliate protozoon) if exposure occurs prior to a critical time after the end of the synchronizing treatment (EST). Exposure after this critical time does not significantly delay the first synchronized division but does delay the second. The time of the transition from a state where the cells are delayed to one where they are not is somewhat dose dependent: within limits, the larger the exposure, the later is the transition time. The division delay response is always correlated with a resorption of the oral primordium of the presumptive posterior daughter cell. The amount of time for resorption varies with the time after EST at which the cells are irradiated and is dose dependent. The data are consistent with the hypothesis that irradiation produces damage during all parts of the synchronized cell cycle, but that, depending on the dose and the time of irradiation after EST, the damage may or may not trigger a response that leads t...

The Common Occurrence of Micronuclear Variation During Binary Fission in an Unusual Race of Paramecium caudatum

Unstable micronuclear behavior was investigated in an unusual race of Paramecium caudatum. Lines were begun, after isolating single vegetative specimens, to yield mass cultures. After study of living specimens in the precision micro‐compression chamber and permanent preparations stained with Feulgen and iron‐hematoxylin, most paramecia revealed a single micronucleus but many were observed with two micronuclei and fewer with more than two micronuclei—occasionally three, rarely four. Specimens measured 225 microns in length; the micronucleus about seven microns. Instead of normal cyclosis, the protoplasm exhibited great fluidity. Generally two contractile vacuoles were present but specimens with three vacuoles, even four, were not uncommon.The method of producing amicronucleate specimens and those with variable numbers of micronuclei was discovered by studying large numbers of actively dividing paramecia. Instead of a micronucleus passing to each daughter at the end of fission as in normal animals, the entire late telophase stage of the dividing micronucleus is passed to the anterior or posterior daughter cell at the time of constriction of the cell. The result is that one daughter is bimicronucleate and the other daughter amicronucleate. Bimicronucleate and amicronucleate animals are capable of dividing normally. In bimicronucleate specimens, division stages show two micronuclei in late telophase stages that result in four micronuclei, two passing to each daughter cell. However, aberrations may again occur in fission to produce variations in number of micronuclei. The irregularly shaped, atypical, macronucleus may extrude chromatin or may undergo complete fragmentation (hemixis, type D). Paramecia and macronuclei may be parasitized by a microorganism which may cause this derangement of normal micronuclear events. This possibility of induction of micronuclear variation in Paramecium by foreign microorganisms may be of considerable evolutionary significance in the genus.