Cell volume growth after cell cycle block with chemotherapeutic agents.

The effects of various chemotherapeutic agents on the volume of Chinese hamster V79 fibroblasts and murine lymphoma L5178Y cells were studied by electronic volume spectroscopy. Cells arrested in the division cycle by a chemotherapeutic block continued to grow in volume resulting in abnormally large cells unable to reduce their volume by cell division. This was observed in cells treated with colcemid, vinblastine, excess thymidine, hydroxyurea, ARA-C, 5-fluorouracil, actinomycin-D and bleomycin, but not with puromycin or cycloheximide. Increase in cell volume of blocked cells was correlated with a decrease in cell survival as measured by clonogenic ability. The results suggest the possibility of volume spectroscopy for a rapid in vitro test to determine tumor sensitivity to chemotherapeutic agents and the in vivo monitoring of response to chemotherapy. Mechanisms for increased cell kill by a second agent acting selectively on enlarged cells are considered.     

DIEL IN SITU PICOPHYTOPLANKTON CELL DEATH CYCLES COUPLED WITH CELL DIVISION1

The diel variability in picophytoplankton cell death was analyzed by quantifying the proportion of dead cyanobacteria Prochlorococcus and Synechococcus cells along several in situ diel cycles in the open Mediterranean Sea. During the diel cycle, total cell abundance varied on average 2.8 ± 0.6 and 2.6 ± 0.4 times for Synechococcus and Prochlorococcus populations, respectively. Increasing percentages of dead cells of Prochlorococcus and Synechococcus were observed during the course of the day reaching the highest values around dusk and decreasing as the night progressed, indicating a clear pattern of diel variation in the cell mortality of both cyanobacteria. Diel cycles of cell division were also monitored. The maximum percentage of dead cells (Max % DC) and the G2 + M phase of the cell division occurred within a period of 2 h for Synechoccoccus and 4.5 h for Prochlorococcus, and the lowest fraction of dead cells occurred at early morning, when the maximum number of cells in G1 phase were also observed. The G1 maximum corresponded with the maximal increase in newly divided cells (minimum % dead cells), and the subsequent exposure of healthy daughter cells to environmental stresses during the day resulted in the progressive increase in dying cells, with the loss of these cells from the population when cell division takes place. The discovery of diel patterns in cell death observed revealed the intense dynamics of picocyanobacterial populations in nature.     

CELL DIVISION AND CELL ELONGATION IN LEAF DEVELOPMENT OF XANTHIUM PENSYLVANICUM

Maksymowych, Roman. (Villanova U., Villanova, Pa.) Cell division and cell elongation in leaf development of Xanthium pensylvanicum. Amer. Jour. Bot. 50(9) : 891–901. Illus. 1963.—Cell division in different parts of the lamina and cell enlargement of the upper epidermis and palisade mesophyll were studied in vertical and horizontal planes during the entire period of growth. The leaf plastochron index (L.P.I.) was used for designation of developmental stages of the leaf. From cell-length data and measurements of cell area the absolute rates of elongation (dX/dpl) and relative rates of elongation (dlnX/dpl) were calculated. The increase in number of cells in the early plastochrons is exponential and cell division stops at about L.P.I. 3.0. Divisions cease first at the tip and last in the basal lobes of the leaf, indicating a basipetal trend of this process. Cells are elongating while division is in progress, though this elongation proceeds at low rates and for a limited time. Palisade cells elongate in the vertical plane at higher rates and at least 1 plastochron sooner than the upper epidermis. The latter cells, however, expand in area with higher absolute and relative rates, and about 2 plastochrons in advance of the palisade mesophyll. The rates are not constant during the whole period of development but are represented by the bell-shaped curves with maximal peaks around L.P.I. 3.0 for the middle portion of the lamina. The increase in volume of the 2 types of cells stops around L.P.I. 5.0, or shortly after. In addition to unequal durations of cellular enlargement, both tissues expand at differential rates, which for the upper epidermis is high in the horizontal plane but low in the vertical plane, while the opposite is true for the palisade mesophyll. It is suggested that palisades and spongy mesophyll are separated and intercellular spaces formed during the course of development because of the greater rate of expansion in area of the upper epidermis.     

Unbalanced cell growth and increased protein synthesis induced by chemotherapeutic agents

Unbalanced cell growth as manifested by an increase in cellular volume and in cellular dry mass following exposure to a variety of chemotherapeutic agents has been shown for neoplastic cells in vitro and human leukemic cells in vivo. The purpose of the present investigation was to test the hypothesis that unbalanced cell growth results from a disassociation of cell growth and cell division due to the blocking effect of chemotherapeutic agents. Monolayer cultures of CHO fibroblasts were studied in terms of their response to two chemotherapeutic agents that differ significantly in their mode of action, adriamycin and chlorambucil. Following exposure to these drugs, cell volume increased at a rate of from 1% to 4% per h; the total cell protein increased at a rate of from 4% to 7% per h. These changes were observed in both log and stationary phase cultures. Thus exposure to adriamycin and chlorambucil was followed by a more rapid rate of protein synthesis relative to the rate of degradation, resulting in larger cells with more protein whether or not the cells were actively in the division cycle. This is inconsistent with the hypothesis that unbalanced growth results simply from a disassociation of the cell division cycle from cell growth. These observations suggest that a final common pathway in the mode of action of chemotherapeutic agents may be the induction of unscheduled protein synthesis resulting in unbalanced cell growth.     

MEASUREMENT OF MICROALGAL CELL VOLUME BY FLOW CYTOMETRY

Single cell analysis by flow cytometry is a powerful tool that has been employed to identify many different characteristics of phytoplankton populations. Cell volume is an important physiological component of many cellular processes. We have used a Coulter EPICS XL flow cytometer to measure cell volume in the spheroid dinoflagellate Amphidinium operculatum as a function of forward scatter. Cell volume measurements of this alga were quantified as equivalent spherical diameters from a standard curve obtained with latex beads of known diameter. This parameter was used to monitor cell diameter throughout the cell division cycle. In log phase cultures, A. operculatum showed increasing cell volumes throughout the light phase and a maximum cell volume concurrent with the onset of cell division late in the light phase. The maximum equivalent spherical diameter measured 14 μm, while the minimum equivalent spherical diameter was 10 μm that occurred late in the dark phase. Stationary phase cultures of A. operculatum did not exhibit oscillating cell volumes throughout the diel cycle. Chemical inhibition of the cell cycle using 100 μM olomoucine diminished cell volume changes during the light phase. These results suggest a coupling of size control to the cell division cycle.     

COBALT EFFECTS ON CELL DIVISION AND CALCIUM UPTAKE IN THE COCCOLITHOPHORID CRICOSPHAERA CARTERAE (HAPTOPHYCEAE)1

Cobalt was found to produce effects on cell division, cell protein, volume, calcium uptake, and ultrastructure of the calcifying alga Cricosphaera carterae (Braarud & Fager.) Braarud. The rate of cell division increased slightly as the added Co concentration of the medium was increased from 0–10 μM. At higher added Co concentrations, the rate of division decreased approximately linearly with concentration until division was blocked almost completely at 100 μM Co. Inhibition of division was reversible after 2 days in Co. Protein content was elevated in Co-treated nondividing cells but the rate of protein synthesis was markedly reduced. Cell volume also increased progressively with Co concentration, and after 72 h in 200 μM Co, the volume was 3.2 times that in culture medium (0.09 μM Co). Co had a dual effect on calcification as indicated by the uptake of ⁴⁵Ca. After 48 h treatment, Ca uptake had increased 53% in 100 μM and decreased 40% in 200 μM Co as compared with cells in 0.09 μM Co. Ultrastructurally, Co caused enlargement of the cell vacuole and the appearance of membrane-bound vacuoles containing electron dense bodies.     

DNA SYNTHESIS,CELL DIVISION,AND CELL DIFFERENTIATION DURING LEAF DEVELOPMENT OF XANTHIUM PENNSYLVANICUM

Leaf growth consists of two basic processes, cell division and cell enlargement. DNA synthesis is an integral part of cell division and can be studied with autoradiographic techniques and incorporation of some labeled precursor. Studies were made on the synthesis of nuclear DNA through incorporation of ³H-thymidine in various parts of the lamina during the entire course of leaf development of Xanthium pennsylvanicum. The time course analysis of DNA synthesis was correlated with cell division and rates of cell enlargement. Significant differences in ³H-thymidine incorporation were found in various parts of the lamina. Cell division and DNA synthesis were highest in the early stages of development. Since no ³H-thymidine was incorporated after cessation of cell division (LPI 2.8) in the leaf lamina, it appears that DNA synthesis is not needed for enlargement and differentiation of Xanthium cells. Rates of cell enlargement were negligible in the early development and reached their maximum after cessation of mitoses, between plastochron ages (LPI) 3 and 4. Cells matured between LPI's 5 and 6. Enzymatic activity was correlated with cell division and cell differentiation at various stages of leaf development.     

GROWTH AND CELL CYCLE OF TWO CLOSELY RELATED RED TIDE-FORMING DINOFLAGELLATES: GYMNODINIUM NAGASAKIENSE AND G. CF. NAGASAKIENSE1

Small-sized vegetative cells were found to co-occur with normal-sized cells in populations of the European bloom-forming dinoflagellate Gymnodinium cf. nagasakiense Takayama et Adachi, currently known as Gyrodinium aureolum Hulburt, but not in populations of the closely related Japanese species Gymnodiniumn agasakiense. We examined how cell size differentiation may influence growth and cell cycle progression under a 12:12-h light: dark cycle in the European taxon, as compared to the Japanese one. Cell number and volume and chlorophyll red fluorescence in both species varied widely during the photocycle. These variations generally appeared to be related lo the division period, which occurred at night, as indicated by the variations of the fraction of binucleated cells (mitotic index) as well as the distribution of cellular DNA content. “Small” cells of G. cf. nagasakiense divided mainly during the first part of the dark period, although a second minor peak of dividing cells could occur shortly before light onset. In contrast, “large” cells displayed a sharp division peak that occurred 9 h after the beginning of the dark period. The lower degree of synchrony of “small” cells could be a consequence of their faster growth. Alternatively, these data may suggest that cell division is lightly controlled by an endogenous clock in “large” cells and much more loosely controlled in “small” cells. Cells of the Japanese species, which were morphologically similar to “large” cells of the European taxon, displayed an intermediate growth pattern between the two cell types of G. cf. nagasakiense, with a division period that extended to most of the dark period.     

THE DEPENDENCE OF CELL DIVISION IN CHLORELLA ON TEMPERATURE AND LIGHT INTENSITY

The effects of temperature and light on cell division were studied in synchronized suspensions of the high-temperature strain Chlorella 7–11–05. It was found that the time for incipient cell division, the progress in the process after it started, and the number of cells produced are influenced by temperature and light intensity. Within limits, cell division is generally favored by the increase in temperature. The increase in light intensity first favors cell division then, after the optimal light intensity is attained, a further increase in light intensity inhibits cell division. Observations are discussed in connection with the findings of other investigators. The limitations of cell division by temperature and light intensity are considered to be separate from the effects of these factors on growth.     

ROLE OF ASYMMETRIC CELL DIVISION IN PTERIDOPHYTE DIFFERENTIATION. II. EFFECT OF CA2+ ON ASYMMETRIC CELL DIVISION,RHIZOID ELONGATION,AND ANTHERIDIUM DIFFERENTIATION IN VITTARIA GEMMAE

Gametophytes of Vittaria graminifolia reproduce vegetatively by means of gemmae. Each gemma consists of a linear array of six cells: four body cells and a knob-shaped terminal cell at each end. When gemmae are shed from the gametophyte onto Knop's mineral medium, the two terminal cells do not divide, but elongate to form primary rhizoids. The body cells undergo asymmetric cell division, and the smaller daughter cells differentiate into either secondary rhizoids or prothalli. When gibberellic acid is included in the medium, antheridia are formed as a result of asymmetric cell division instead of vegetative structures. We studied the effect of Ca²⁺ on asymmetric cell division, rhizoid elongation, and antheridium formation in gemmae cultured on Knop's mineral medium and variations of Knop's medium. Ca²⁺ inhibited the onset of cell division and rhizoid elongation, but was required for differentiation of antheridia. Treatments which lowered the Ca²⁺ content of gemmae (EGTA and dilute HCl extraction, culture on verapamil-containing and Ca²⁺-deficient medium) caused an early onset of cell division and rhizoid elongation. The stimulation of growth was most pronounced when gemmae were deprived of Ca²⁺ during the first 24 hr of culture. The proportion of cell divisions which differentiated into antheridia in response to GA was greatly reduced when the Ca²⁺ status of gemmae was lowered with verapamil and Ca²⁺-EGTA buffers.     

CELL DIVISION IN THE DINOFLAGELLATE GAMBIERDISCUS TOXICUS IS PHASED TO THE DIURNAL CYCLE AND ACCOMPANIED BY ACTIVATION OF THE CELL CYCLE REGULATORY PROTEIN,CDC2 KINASE1

The cell division cycle in several pelagic dinoflagellate species has been shown to be phased with the diurnal cycle, suggesting that their cell cycle may be regulated by a circadian clock. In this study, we examined the cell cycle of an epibenthic dinoflagellate, Gambierdiscus toxicus Adachi and Fukuyo (Dinophyceae), and found that cell division was similarly phased to the diurnal cycle. Cell division occurred during a 3-h window beginning 6 h after the onset of the dark phase. Cell cycle progression in higher eukaryotes is regulated by a cell cycle regulatory protein complex consisting of cyclin and the cyclin-dependent kinase CDC2. In this report, we identified a CDC2-like kinase in G. toxicus that displays activity in vitro against a known substrate of CDC2 kinase, histone H1. As in higher eukaryotes, CDC2 kinase was expressed constitutively in G. toxicus throughout the cell cycle, but it was activated only late in the dark phase, concurrent with the presence of mitotic cells. These results indicate that cell division in G. toxicus is regulated by molecular controls similar to those found in higher eukaryotes.     

CHANGES IN CELL COMPOSITION AND LIPID METABOLISM MEDIATED BY SODIUM AND NITROGEN AVAILABILITY IN THE MARINE DIATOM PHAEODACTYLUM TRICORNUTUM (BACILLARIOPHYCEAE)1

The effects of nitrogen starvation in the presence or absence of sodium in the culture medium were monitored in batch cultures of the marine diatom Phaeodactylum tricornutum Bohlin. During nitrogen starvation in the presence of sodium, cell nitrogen and chlorophyll a decreased, mainly as a consequence of continued cell division. These decreases were accompanied by decreases in the rates of photosynthesis and respiration. There was no change in either cell volume or carbohydrate, but both carbon and lipid increased. During nitrogen starvation in the absence of sodium, cell division ceased. Cell nitrogen and chlorophyll a remained constant, and respiration did not decrease, but the changes in the photosynthetic rate and the lipid content per cell were similar to cultures that were nitrogen-starved in the presence of sodium. The carbon-to-nitrogen ratio increased in both cultures. Nitrogen, in the form of nitrate, and sodium were resupplied to cultures that had been preconditioned in nitrogen- and sodium-deficient medium for 5 d. Control cultures to which neither nitrate or sodium were added remained in a static state with respect to cell number, volume, and carbohydrate but showed slight increases in lipid. Cells in cultures to which 10 mM nitrate alone was added showed a similar response to cultures where no additions were made. Cells in cultures to which 50 mM sodium alone was added divided for 2 d, with concomitant small decreases in all measured constituents. Cell division resumed in cultures to which both sodium and nitrate were added. The lipid content fell dramatically in these cells and was correlated to metabolic oxidation via measured increases in the activity of the glyoxylate cycle enzyme, isocitrate lyase. We conclude that lipids are stored as a function of decreased growth rate and are metabolized to a small extent when cell division resumes. However, much higher rates of metabolism occur if cell division resumes in the presence of a nitrogen source.     

A COMPARATIVE STUDY OF CELL DIVISION PATTERNS DURING GERMINATION OF SPORES OF ANEMIA,LYGODIUM AND MOHRIA (SCHIZAEACEAE)

Cell division patterns during germination of spores of Anemia (A. hirsuta, A. munchii, A. phyllitidis), Lygodium (L. circinatum, L. flexuosum, L. japonicum, L. salicifolium) and Mohria caffrorum have been examined by light microscopy of glycol methacrylate embedded materials. Spores of all species in a genus exhibited a constant pattern of division under different conditions of germination. In spores of species of Anemia, following an asymmetrical division, the proximal cell differentiated into the protonemal cell while the distal cell divided to produce the rhizoid. A similar division sequence was found in spores of Mohria caffrorum, but the fate of cells formed was reversed. In Lygodium spores, a proximal cell formed by an initial division of the spore cut off a protonemal cell, a rhizoid and a wedge-shaped cell by walls parallel to the polar axis. Our results contradict earlier observations on cell division sequence during germination of spores of these genera based on whole mount preparations.     

THE EFFECT OF AUXIN AND GUANINE ON CELL EXPANSION AND CELL DIVISION IN THE GAMETOPHYTE OF THE FERN,ONOCLEA SENSIBILIS

Miller , J. H. (Yale U., New Haven, Conn.) The effect of auxin and guanine on cell expansion and cell division in the gametophyte of the fern, Onoclea sensibilis. Amer. Jour. Bot. 48(9): 816–819. Illus. 1961.—Auxin and guanine promote cell expansion in 0. sensibilis gametophytes. The optimum concentration of auxin for total expansion is 10^−-5 M, but the optimum for elongation is 10^−-6 M. Above this concentration the cells expanded isodiametrically. Guanine is active at higher concentrations than auxin. Increasing concentrations of auxin progressively inhibit red light-induced cell division, while guanine has no effect on cell division. Neither kinetin nor adenine promotes cell expansion or cell division.     

SMALL CELL AND INTERMEDIATE CELL FORMATION IN SPECIES OF DINOPHYSIS (DINOPHYCEAE, DINOPHYSIALES)

Observations of two distinct size classes with similar shape in natural populations of Dinophysis Ehrenberg were first reported by Jorgensen in 1923 and intermediate forms exhibiting a continuum between the typical vegetative cell and a putative small cell by Wood in 1954. Focused attention on Dinophysis spp. associated with diarrhetic shellfish intoxications in the last decade has provided new examples of small cells in the genus, sometimes with contours dissimilar from the corresponding vegetative cells; dimorphic individuals; and large/small cell couplets. This work was based on in situ observations during intensive sampling for cell cycle studies of Dinophysis acuminata Claparéde et Lachmann, Dinophysis acuta Ehrenberg, Dinophysis caudata Saville-Kent, and Dinophysis tripos Gourret; on laboratory incubations of D. acuminata; and on a thorough search of documented information on morphological variability of Dinophysis spp. During in situ division, most dividing cells exhibit a normal longitudinal fission, but some (1%–10%) undergo a “depauperating” fission, leading to pairs of dimorphic cells with dissimilar moieties. After separation and sulcal list regeneration, these dimorphic cells become D. skagii Paulsen, D. dens Pavillard, D. diegensis Kofoid, and D. diegensis Kofoid var. curvata-like individuals, which can also be observed forming couplets D. acuminata/D. skagii, D. acuta/D. dens, and D. caudata/D. diegensis attached by their ventral margins. Small cells can grow again to large size, as shown in laboratory incubations of D. acuminata, thus partly explaining observations of thecal intercalary bands, and intermediate forms. The sexual nature of the small cells will not be unequivocally demonstrated until controlled germination of the alleged cyst forms is achieved, and some intermediate forms may correspond to undescribed stages after cyst germination. These observations suggest common patterns in the life cycle of Dinophysis spp. Intraspecific morphological variability of Dinophysis spp. in a given geographic area can largely be attributed to small cell formation, as a response to changing environmental conditions, and may be a part of the sexual cycle of these species. Small cells seem to be able to enlarge, leading to intermediate cell and further vegetative cell formation as part of a three-looped life history pattern in Dinophysis.     

COMPARATIVE PHYSIOLOGICAL STUDY OF MARINE DIATOMS AND DINOFLAGELLATES IN RELATION TO IRRADIANCE AND CELL SIZE. I. GROWTH UNDER CONTINUOUS LIGHT1,2

Cell division rates and chlorophyll a and protein contents for ten diatom and dinoflagellate species were measured. Species were chosen to include a wide range of cell size in terms of both cell volume and cell protein: from 0.004 ng protein/cell for a small Chaetoceros sp. to 2.2 ng protein/cell for Prorocentrum micans Ehrenberg. Experiments were conducted in batch or semi-continuous cultures at 21 C under continuous illumination from 8–256 μEin ^.m^-2'^.s^-1. Light saturation of cell division occurred at 32–80 μEin m^-1 s^-1 for all species, with no observable difference between the two phylogenetic groups. When the light-saturated cell division rates were plotted against cell size as protein/cell, the diatoms and dinoflagellates fell on two separate lines with the diatoms having higher rates. Chl a /protein ratios (μg/μg) decreased with increasing irradiance. The diatoms had higher chl a per unit protein. The relationship between cell division rate and the chl a/protein ratio is discussed.     

DIURNAL CELL DIVISION REGULATED BY GATING THE G1/S TRANSITION IN ENTEROMORPHA COMPRESSA (CHLOROPHYTA)1

The cell‐cycle progression of Enteromorpha compressa (L.) Nees (=Ulva compressa L.) was diurnally regulated by gating the G₁/S transition. When the gate was open, the cells were able to divide if they had attained a sufficient size. However, the cells were not able to divide while the gate was closed, even if the cells had attained sufficient size. The diurnal rhythm of cell division immediately disappeared when the thalli were transferred to continuous light or darkness. When the thalli were transferred to a shifted photoperiod, the rhythm of cell division immediately and accurately synchronized with the shifted photoperiod. These data support a gating‐system model regulated by light:dark (L:D) cycles rather than an endogenous circadian clock. A dark phase of 6 h or longer was essential for gate closing, and a light phase of 14 h was required to renew cell division after a dark phase of >6 h.     

INFLUENCE OF CELL VOLUME ON THE GROWTH AND SIZE REDUCTION OF MARINE AND ESTUARINE DIATOMS1

The maximal growth rate (μ_max) of 19 marine and estuarine diatoms decreased with increasing cell volume (V). The relationship between log μ_max (Y) and log V (X) was calculated. Statistical analyses showed that the slope of the equation was not significantly different from those obtained by other researchers and that the 95% confidence intervals of mean μ_max at cell volumes of 10³–10⁵μm³ were not significantly different from those cited in most studies. A new regression line for diatoms was calculated as follows: log μ_max= 0.47–0.14 log V; r =–0.69. The rate of size reduction per generation of the 19 diatom species ranged from 0.03 to 0.87 μm per generation. The rate increased with increasing cell length and cell volume and with decreasing maximum division rate. Statistical analyses showed that the rate was closely related to the cell volume and to the reciprocal of the growth rate. The relationships between maximal growth rate and cell volume and between rate of size reduction and cell volume showed that a diatom with a large volume had a smaller maximal growth rate and a larger rate of size reduction than a diatom with a small volume. The estimates using the equation for the regression line between the rate of size reduction and the reciprocal of maximum division rate indicated that a diatom with a high maximum division rate would need more generation equivalents for a certain size reduction than a diatom with a low maximum division rate, but the periods required for reduction would be approximately equal irrespective of maximum division rate.     

Correlation of cell division and cell expansion to potato microtuber growth in vitro

The sink capacity of plant storage organs influences crop economic yield and relates to the number and volume of their cells. To obtain a better understanding of their contributions to the growth of potato microtubers produced in vitro, the number and volume of the cells in the tuber tissues were measured as tubers grew. Two potato cultivars, E-Potato 1 and Mira were employed and the results showed that cortex, perimedulla and pith tissue contributed for about 30, over 65 and up to 3% to the volume of the mature microtuber, respectively. The number of cells and cell volume increased simultaneously as the microtubers grew and the relationships could be described by a power function, Y = aW ^b. However, the rate of cell division was greater than the rate of cell expansion and the former contributed more than the latter to the increase in tuber size. The rate of cell division was greatest in the cortex and least in the pith, but, because the perimedulla forms the largest part of the tuber, cell division in this tissue was particularly important. The regulation of cell division to improve the production of usable microtubers is discussed.     

CELL DIVISION IN BULBOCHAETE. II. HAIR CELL FORMATION1

Most vegetative cells of Bulbochaete, and all those of Oedogonium, possess an apical, circular discontinuity in the structure of their secondary wall. Rupture of the wall at this precise site permits expansion of the ring during cell division and release of the zoospore following zoosporogenesis. Certain cells of Bulbochaete (always the apical daughter cell of a division pair) lack this type of discontinuity. Instead, the apical wall is thinned out on one side, so that the cell bulges asymmetrically. In the middle of the bulge is a wall discontinuity which extends only part way around the cell. The wall will rupture here, too, for zoospore release, but if a cell having such a wall, divides, it invariably does so asymmetrically, with one pole of the spindle located in the bulge. Cytokinesis then cuts off a small, colorless daughter cell. The wall ruptures at the discontinuity, and this daughter cell emerges through the slit and differentiates into a hair. The creation of hairs in such cells commences with the deposition of a pad of primary wall lining the bulge. Golgi bodies are involved in its secretion, but not in that of a secondary wall layer which forms next in the premitotic cell and covers the primary wall. The cell becomes polarized; the nucleus migrates toward this region as the chloroplast moves aside. After the asymmetric mitosis, a curved phycoplast cuts off the hair cell nucleus and prevents the chloroplast from moving back into the future hair, whose cytoplasm soon loses much of its affinity for heavy metal stains. Following rupture of the parental wall, the phycoplast moves some distance past the limits of the newly deposited secondary wall layer and then forms a cross wall under the hair. The secondary wall of the hair is not continuous with the secondary wall structure of the parental cell; the circular discontinuity that arises around the base of the bulging parental wall is then perpetuated and accentuated as the hair's secondary wall thickens. This wall weakening becomes the dislocation that will predetermine the site of the ring and consequently the direction of cell expansion in the next normal division of the cell subtending the hair. Abnormal ring formation and the creation of terminal twin hairs have also been examined. The lip of the growing hair contains a characteristic organization of membranes and other components which may be related to the organization of the hair's numerous longitudinally oriented microtubules. These results are discussed in terms of the morphology of the wall in the Oedogionales generally. The creation of the special wall morphology that leads to hair cell formation is considered to be ontogenetically related to a similar wall morphology that is involved in formation of the fertilization pore of the oogonium.