PHOSPHATASE ACTIVITY AND CELLULAR DIFFERENTIATION IN PHLEUM ROOT MERISTEM

Using 9 different organic phosphate substrates as alternatives in a standardized 5′-nucleotidase histochemical test system, enzyme activity patterns were recorded for timothy grass root epidermis. At least 4 different phosphatases were distinguished on the bases of substrate specificity, reaction rate, tissue distribution, and response to inhibitors. Except with adenosine-3′-monophosphate, all activities were restricted to the 300-mμ-long root tip meristem. These enzyme activities were associated with the earliest phases of differentiation of the epidermal hair and hairless cell initials. The distribution of activities was not associated with the same cell type in each part of the meristem. Little activity was found with most substrates in the undifferentiated cells of the 0-100μ zone; alternating active hairless and inactive hair cell initials predominated in the 100-200μ segment; and active hair–inactive hairless sister cells formed the principal pattern in the 200-300μ segment of the meristem. The data showed that a particular enzyme activity was associated with a specific cell type only in relation to that cell's position along the differentiation gradient of the entire tissue. But, within a meristem segment, a specific cell type might act differently from its neighbors, depending on its mitotic capacity. This complex of physiological dependence and independence of a cell type on tissue ontogeny was cited as a characteristic of the phenomenon of cellular differentiation superimposed on tissue differentiation gradients.     

NUCLEOLAR SIZE DIFFERENCES IN THE GRASS ROOT EPIDERMIS

Root tips of the festucoid grass, Festuca arundinacea, and 2 panicoid species, Chloris gayana and Panicum virgatum, were processed using 2 different staining techniques. Measurements of nucleolar size were taken on epidermal and cortical cells. Trichoblasts and hair cells of Festuca were found to contain much larger nucleoli than those in hairless initials or hairless cells. Significant nucleolar size differences between hair and hairless cells were also found in the 2 panicoid species. In contrast to Festuca, this difference between the 2 cell types was not as pronounced, and overlapping in nucleolar size occurred between adjacent hair and hairless cells. The cortex was composed of rows of cells in which nucleolar size simply decreased with cell distance from the apex. The significance of the observed nucleolar differences among cell types of the root tip is discussed briefly in relation to systematics, enzyme activity patterns, and differentiation.     

COMPARATIVE ENZYME DIFFERENTIATION IN GRASS ROOTS. I. ACID PHOSPHATASE

Avers , Charlotte J. (U. Miami, Coral Gables, Fla.), and Robert B. Grimm . Comparative enzyme differentiation in grass roots. I. Acid phosphatase. Amer. Jour. Bot. 46(3) : 190-193. Illus. 1959.—There is a correlation between the pattern of acid phosphatase activity and the particular morphogenetic pattern in the root epidermis of festucoid and panicoid grasses. Four festucoid species all showed intensified enzyme activity in trichoblasts and loss of activity in hairless cell initials prior to the maturation of these cells. The 3 panicoid grasses showed no phosphatase-inactive cells during epidermal development. The festucoid epidermis contains alternating long and short cells which differentiate into hairless and hair cells respectively. The panicoid type shows no such cellular pattern and any epidermal cell seems capable of producing a root hair. Treatment of Phleum roots with 10^-4 M coumarin caused a foreshortening of the growth zones and a concurrent apical shift in differential acid phosphatase activity. This response was interpreted as further evidence of a direct correlation between the morphogenetic and enzymatic differentiations in the root epidermis.     

Control of nucleolar growth during interphase in higher plant meristem cells

Summary The evolution of nuclear and nucleolar sizes throughout interphase have been studied in synchronous caffeine-labeled binucleate cells of onion root meristems by using silver impregnation and stereological methods over semithin sections. Nucleus and nucleolus grow independently, since nucleolus enlarges at its fastest rate in G 1, while nucleus grows mostly in two periods: onset of replication and G 2. Nucleolar size in the cycle seems to be a genecontrolled function, hardly affected by protein synthesis inhibition. Hence, there is a biphasic response to cycloheximide (CHM) in the fast growing nucleoli of both early and late G 1 with an initial stimulation later counterbalanced by a depressed rate, so that nucleolar size in S was similar to control shortly afterwards the start of the CHM treatment. The initial enlargement under CHM was due to an increase of all nucleolar structural components, i.e., fibrillar, granular, vacuolar, and lacunar regions. No cycloheximide effect whatsoever was detected in S and G 2 nucleoli.Abbreviations CHM cycloheximide - F fibrillar component - G granular component - L lacunae - V vacuoles - VN nuclear volume - VNu nucleolar volume - VvNu volume density of the nucleoli     

ORIGIN,DEVELOPMENT, AND GROWTH OF DIFFERENTIATING TRICHOBLASTS IN ELODEA CANADENSIS

Root hairs of Elodea canadensis develop only from cells which undergo a particular series of developmental steps. These cells, the trichoblasts, are formed as the smaller, proximal product of an asymmetric division, and immediately enter a prolonged phase of synthesis. Histochemical tests show that large amounts of RNA and protein accumulate in the vastly enlarged nucleolus and cytoplasm, while histone increases in the enlarging nucleus. Cytophotometry shows that DNA in the nucleus reaches polyploid levels. Throughout the synthetic phase, almost to the point of root hair initiation at 9.5 mm proximal to the meristem initials, vacuolation is delayed and the trichoblasts elongate less extensively. All results suggest that this synthesis is the type which normally follows cell division, but is greatly enhanced in the trichoblast. In contrast, the initially larger atrichoblasts only accumulate RNA, DNA, and protein in the region from 1 mm to 2 mm proximal to the meristem tip, and they then enter a phase of extensive vacuolation and elongation.     

Relation between the nuclei and the nucleoli during cell differentiation in roots of Vicia faba volumetric and cytochemical analysis

Summary The behaviour of the nuclei and the nucleoli of roots of Vicia faba during cell differentiation was studied quantitatively. The relations between these cell constituents and the polyploidy was analysed. The study was made on isolated nuclei and nucleoli and on plastic sections. A method for the isolation of nuclei and nucleoli of secondary roots fixed in formol was modified and another developed for material fixed in ethanol/acetic acid mixture. The volumetric investigation showed that the nuclear volume increases while the nucleolar decreases during cell differentiation. The mean number of nucleoli decreases. In Vicia faba there is no relation between the ploidy and the volume of nuclei and nucleoli; the protein synthesis rate has an influence on the size of these organelles. Quantitative investigation has shown the proportionality of dry weight, DNA, total protein, histone, protein-bound lysine and arginine content of the nuclei and their ploidy. The same experiments made on nucleoli showed linear relation between their content and volume. The concentration of analysed substances in constant in nucleoli.     

Cell-to-cell movement of the CAPRICE protein in Arabidopsis root epidermal cell differentiation

CAPRICE (CPC), a small, R3-type Myb-like protein, is a positive regulator of root hair development in Arabidopsis. Cell-to-cell movement of CPC is important for the differentiation of epidermal cells into trichoblasts (root hair cells). CPC is transported from atrichoblasts (hairless cells), where it is expressed, to trichoblasts, and generally accumulates in their nuclei. Using truncated versions of CPC fused to GFP, we identified a signal domain that is necessary and sufficient for CPC cell-to-cell movement. This domain includes the N-terminal region and a part of the Myb domain. Amino acid substitution experiments indicated that W76 and M78 in the Myb domain are critical for targeted transport, and that W76 is crucial for the nuclear accumulation of CPC:GFP. To evaluate the tissue-specificity of CPC movement, CPC:GFP was expressed in the stele using the SHR promoter and in trichoblasts using the EGL3 promoter. CPC:GFP was able to move from trichoblasts to atrichoblasts but could not exit from the stele, suggesting the involvement of tissue-specific regulatory factors in the intercellular movement of CPC. Analyses with a secretion inhibitor, Brefeldin A, and with an rhd3 mutant defective in the secretion process in root epidermis suggested that intercellular CPC movement is mediated through plasmodesmata. Furthermore, the fusion of CPC to tandem-GFPs defined the capability of CPC to increase the size exclusion limit of plasmodesmata.     

Relations between nucleolar morphometric parameters and pre-rRNA synthesis in animal and plant cells

In order to study if there are differences between cells of the same tissue with one and two nucleoli, nuclear and nucleolar volume, density of tritiated uridine incorporation, amount of DNA per nucleus and intensity of cytoplasmic basophilia were measured in mononucleolated and binucleolated rat epithelial endometrial cells, in onion root meristematic cells and in chick embryo matrix cells of the central nervous system, neuroblasts and neurons. No significant differences in nuclear volume, density of tritiated uridine incorporation and amount of DNA per nucleus were found between cells of the same type with diverse numbers of nucleoli. Binucleolated endometrial cells, matrix cells, and root meristematic cells have biphasic distributions of nucleolar volumes. One peak of this distribution roughly coincides with the nucleolar volume of mononucleolated cells, the other peak corresponds almost to double the volume. As the density of uridine incorporation is the same irrespective of the nucleolar number and volume, the cells with larger nucleolar volumes have higher pre-rRNA synthesis. These cells also have higher amounts of ribosomes in the cytoplasm, as revealed by the photometric study of basophilia. It is concluded that in this population of cells the ribosomal production is regulated to a higher steady equilibrium than in the general population. This difference is not due to polyploidism or to the increased DNA content of G2 phase cells. Binucleolated neuroblasts and neurons have nucleolar volumes similar to those of mononucleolated ones.     

Cytodifferentiation of the chick pancreas. 3. The content and synthesis of ribonucleic acid and proteins in developing acinar cells

The content and synthesis of ribonucleic acid (RNA) and protein was studied by microphotometry and autoradiography in the developing pancreatic acinar cells of White Leghorn chick embryos. These findings were correlated with previously reported changes in ultrastructural components. Shortly before or concomitant with zymogen granulation, RNA synthesis increased, in association with increases in the amount of nucleolar and cytoplasmic protein. The cytoplasmic fraction was transitory, whereas the accumulated nucleolar protein was maintained and was soon followed by an increase in nucleolar RNA. Concomitantly, a decrease in chromosomal RNA was observed, with the total amount of nuclear RNA staying constant. When zymogen first appeared, nucleoli were greatly enlarged due to large amounts of RNA and protein; total cellular RNA and protein had decreased slightly, in association with a decrease in cell volume. Subsequent development presented smaller nucleoli with decreased amounts of RNA and protein. Total cellular RNA increased due to its accumulation in the cytoplasm, probably as ribosomes. The accumulation of zymogen and the enlargement of other cellular structures contributed to an increase in total cellular protein. Prior to hatching, total cell RNA and protein decreased in amount, probably due to a reduction in cell volume through cell division.     

Ulfrastructure of the histological zones in growing vegetative buds of Salix spp.

Ultrastructural differentiation in the shoot apex of growing vegetative buds of Salix was studied, and some micrographs analysed morphometrically. The distribution of inorganic phospahte (P;) was analysed cytochemically. A distinct histological zona–tion was observed in the apex. The relative volumes of nuclei and plastids were significantly higher in the central tunica zone than in the peripheral one. The corpus differed from the central tunica zone by significantly lower volume density of nuclei and higher of vacuoles and mitochondria. During differentiation of the rib meristem vacuole volume increased significantly, while the relative volumes of nuclei, mitochondria, nucleoli, and heterochromatin decreased. It was not possible to decide whether the vacuoles originate from ER or GERL. Morphogenesis of chloroplasts with large starch grains and grana from proplastids was evident in the rib meristem; dedifferentiation to S–plastids was found in the protophloem. Prolamellar bodies were observed in the procambium plastids. The protophloem was characterized by P–protein and spiny vesicles. P_i was found in the nucleoli of most epidermis cells, several procambium cells, and a few chlorencyma cells, but never in the tunica of the growing apical and developing lateral buds. P_i also occurred in some plasmalemma–somes and occasionally in the walls in connection with plasmodesmata.     

Comparative Enzyme Differentiation in Grass Roots: II PEROXIDASE

Results of histochemical tests for peroxidase activity in sevenspecies of grasses have been reported. The root epidermis ofthe festucoid grasses is characterized by rows of alternatingshorter hair and longer hairless cells which can be recognizedthroughout their development. Peroxidase activity occurred inall the growing cells, but intensified reactions were observedin the hair cell initials in the bases portion of the elongationzone. The panicoid species have a root epidermis in which anycell seems capable of producing a root hair, and in these speciesall cells in the growing regions showed equal peroxidase activity.The close correlation between the differentiations of enzymesand cell types implies that physiological changes occur longbefore the morphological maturation of the tissue.     

Two ways to skin a plant: The analysis of root and shoot epidermal development in Arabidopsis

The post-embryonic architecture of higher plants is derived from the activity of two meristems that are formed in the embryo: the shoot meristem and the root meristem. The epidermis of the shoot is derived from the outermost layer of cells covering the shoot meristem through repeated anticlinal divisions. By contrast, the epidermis of the root is derived from an internal ring of cells, located at the centre of the root meristem, by a precise series of both periclinal and anticlinal divisions. Each epidermis has an independent origin. In Arabidopsis the mature shoot epidermis is composed of a small number of cell types: hair cells (trichomes), stomatal guard cells and other epidermal cells. In shoots, hairs take the form of branched trichomes that are surrounded at their base by a ring of accessory cells in a sheet of epidermal cells. The root epidermis is composed of two cell types: trichoblasts that form root hair cells and atrichoblasts that form non-hair cells. Mutations affecting both the patterning and the morphogenesis of cells in both shoot and root epidermis have recently been described. Most of these mutations affect development in a single epidermis, but at least one, ttg, is involved in development in both epidermal systems.     

IMPLICATIONS OF NUCLEOLAR DIFFERENCES IN THE ROOT EPIDERMIS AMONG SEVERAL GRASS SPECIES

The nucleolar sizes of root epidermal cells were determined in 10 species of the Gramineae. Festucoid and panicoid roots presented significantly larger nucleoli in hair-producing, as opposed to hairless, cells. The magnitude of the nucleolar size differences between cell types seemed to be species specific and was not related to the type of epidermal cellular pattern. There seemed to be some suggestion that meristematic nucleoli of festucoid species may generally be larger than those of panicoid types. There was also an indication that festucoid grasses might be characterized by a high percent of multinucleolate epidermal cells. In contrast, cells of panicoid roots contained a single nucleolus in most instances. Possible implications of these observations were briefly discussed in terms of development and systematics.     

Different nucleolar antigen expression in resting and proliferating human lymphocytes as studied by fluorescence microscopy and flow cytometry

Abstract. The aim of this study was to investigate the use of a nucleolar antigen to discriminate between proliferating and resting cells. Antinucleolar antibodies (Si87) were obtained from a scleroderma patient. The specificity of immunostaining was verified and morphological changes in nucleoli were monitored using a fluorescence microscope. Fluorescence of propidium iodide-stained DNA and nucleolar immunofluorescence were measured by flow cytometry.
Following phytohaemagglutinin stimulation the number of nucleoli of normal human peripheral blood lymphocytes increased about 3-fold, accompanied by enlargement of nucleolar size. Simultaneously a mean increase in total immunofluorescence per cell by a factor of three was detected. The method developed and applied here allows a discrimination between resting and proliferating human lumphocytes on the basis of their nucleolar antigen content.     

Different nucleolar antigen expression in resting and proliferating human lymphocytes as studied by fluorescence microscopy and flow cytometry

The aim of this study was to investigate the use of a nucleolar antigen to discriminate between proliferating and resting cells. Antinucleolar antibodies (Si87) were obtained from a scleroderma patient. The specificity of immunostaining was verified and morphological changes in nucleoli were monitored using a fluorescence microscope. Fluorescence of propidium iodide-stained DNA and nucleolar immunofluorescence were measured by flow cytometry. Following phytohaemagglutinin stimulation the number of nucleoli of normal human peripheral blood lymphocytes increased about 3-fold, accompanied by enlargement of nucleolar size. Simultaneously a mean increase in total immunofluorescence per cell by a factor of three was detected. The method developed and applied here allows a discrimination between resting and proliferating human lymphocytes on the basis of their nucleolar antigen content.     

Variations in the Size of Mitotic Cells in the Root Meristem

Variations in the length of mitotic and interphase cells were analyzed in various tissues of wheat roots and in the cortex of maize roots. Reliable differences were shown in the length of mitotic cells in individual file clones of cells of the same tissue. The mean lengths of dividing cells in different roots differed to a lesser extent than those of different files in the same tissue of one root. Within the file, the length of the sister simultaneously dividing cells differed the least, while the difference of lengths of the neighbor simultaneously dividing nonsister cells was bigger. The mean length of interphase cells in any file was always less than that of mitotic cells by a factor of 1.45. This ratio was almost invariable for files and tissues in both the plants we studied and corresponded to that of an exponentially growing cell population. In addition, a very small number of cells were found (less than 1%) in meristems, which are longer than the mitotic cells. The length of these cells exceeded those of mitotic cells by less than twice. The origin of such cells is discussed. The length of mitotic cells near the quiescent center is more variable than in the middle of the meristem in the cortex of both plants. In the meristem basal part, the mitotic cells were no longer than those in the middle of the meristem but there were no small dividing cells. In the wheat epidermis, the cells are differentiated into trichoblasts and atrichoblasts and, therefore, the length of the dividing cells is highly variable. The cell length is essential for their transition to mitosis for all studied proliferating meristem cells.     

Variations in the size of mitotic cells in the root meristem

Variations in the length of mitotic and interphase cells were analyzed in various tissues of wheat roots and in the cortex of maize roots. Reliable differences were shown in the length of mitotic cells in individual files-clones of cells of the same tissue. The mean lengths of dividing cells in different roots differed to a lesser extent than those of different files in the same tissue of one root. Within the file, the length of sister simultaneously dividing cells differed the least, while the difference of lengths of neighbor simultaneously dividing nonsister cells was bigger. The mean length of interphase cells in any file was always less than that of mitotic cells by a factor of 1.45. This ratio was almost invariable for files and tissues in both plants we studied and corresponded to that of an exponentially growing cell population. In addition, a very small number of cells were found (less than 1%) in meristems, which are longer than the mitotic cells. The length of these cells exceeded those of mitotic cells by less than twice. The origin of such cells is discussed. The length of mitotic cells near the quiescent center is more variable than in the middle of the meristem in the cortex of both plants. In the meristem basal part, the mitotic cells were no longer than those in the middle of the meristem but there were no small dividing cells. In the wheat epidermis, the cells are differentiated into trichoblasts and atrichoblasts and, therefore, the length of dividing cells is highly variable. The cell length is essential for their transition to mitosis for all studied proliferating meristem cells.     

Inhibition of RNA Synthesis and the Relationship between Nucleolar and Mitotic Cycles in Zea mays Root Meristems

The relationship between nucleolar and mitotic cycles has beendetermined after treatment of root apices of Zea mays with ethidiumbromide. In the meristematic regions of the stele the two cyclesare not much displaced in relation to each other except fora delay in the onset of the disorganization phase. A few nucleolipersist into metaphase and a few nuclei undergo an amitoticdivision. In the cap initials the drug greatly delays the onsetof disorganization of the nucleolus, which normally occurs beforeprophase in this region. It also delays the completion of reorganizationso that fully organized nucleoli are no longer available duringthe last half of telophase. In the quiescent centre the onsetof disorganization and the end of reorganization of the nucleoliare also delayed in relation to mitosis. There is no evidencefor a delay in the onset of reorganization in any region ofthe meristem. Some cells form multiple micronucleoli and this aberrant behaviouroccurs more often in the cap initials than elsewhere as doesamitotic division.     

Role of a positive regulator of root hair development,CAPRICE, in Arabidopsis root epidermal cell differentiation

In Arabidopsis, root hairs are formed only from a set of epidermal cells named trichoblasts or hair-forming cells. Previous studies showed CAPRICE (CPC) promotes differentiation of hair-forming cells by controlling a negative regulator, GLABRA2 (GL2), which is preferentially expressed in hairless cells. Here, we show that CPC is also predominantly expressed in the hairless cells, but not in the neighboring hair-forming cells, and that CPC protein moves to the hair-forming cells and represses the GL2 expression. We also show that the N terminus of bHLH protein interacts with CPC and is responsible for the GL2 expression. We propose a model in which CPC plays a key role in the fate-determination of hair-forming cells.     

盾叶薯蓣根状茎的发育解剖学和组织化学研究

盾叶薯蓣根状茎顶端的生长点由鳞片包被,其衍生细胞分化为原表皮、基本分生组织和散生的原形成层束,以后分化为表皮、基本组织和散生的维管束构成的初生结构。根状茎顶端下方的原表皮内存在初生增厚分生组织,其细胞不断向内分裂和其衍生细胞的体积增大使根状茎能够迅速增粗。分化完成的根状茎主要由周皮、基本组织和散生的维管束构成。周皮由木栓层、木栓形成层和栓内层组成;基本组织由薄壁细胞组成;维管束属于有限维管束。薯蓣皂甙主要存在于基本组织薄壁细胞中。原分生组织和原形成层不含薯蓣皂甙,维管束的木质部和韧皮部中的韧皮纤维也无薯蓣皂甙的分布,韧皮部的生活细胞和维管束鞘细胞有薯蓣皂甙的积累。近顶端的基本分生组织细胞内薯蓣皂甙不形成液滴,随着细胞分裂逐渐停止,细胞内开始形成含薯蓣皂甙的液滴,反映皂甙是在成熟细胞内积累。其中,有小型维管束分布的基本组织中薯蓣皂甙的积累与分布最丰富,两年生根状茎中薯蓣皂甙的含量比一年生的高。