Ploidy Level Of Cells Research Articles

Endoreduplication in plant organogenesis: a mean to boost fruit growth.

Endoreduplication is the major source of somatic endopolyploidy in higher plants, leading to variation in cell ploidy levels due to iterative rounds of DNA synthesis in the absence of mitosis. Despite its ubiquitous occurrence in many plant organs, tissues and cells, the physiological meaning of endoreduplication is not fully understood, although several roles during plant development have been proposed, mostly related to cell growth, cell differentiation and specialization via transcriptional and metabolic reprogramming. Here we review the recent advances in the knowledge of the molecular mechanisms and cellular characteristics of endoreduplicated cells, and provide an overview of the multi-scale effects of endoreduplication on supporting growth in plant development. Finally, the effects of endoreduplication in fruit development are discussed, since during fruit organogenesis, endoreduplication is highly prominent where it acts as a morphogenetic factor supporting rapid fruit growth as illustrated with case of the fleshy fruit model tomato (Solanum lycopersicum).

BIO-CATHARANTIN EFFECTS ON PHENOTYPIC TRAITS AND CHROMOSOME NUMBER OF SHALLOTS (ALLIUM CEPA L. VAR. ASCALONICUM 'TAJUK')

The seasonal production of Allium cepa var. ascalonicum causes a rise in its demand during the off-season. Consumers mostly prefer onion cultivars like the 'Super Philip', because of their high productivity, large and round bulbs, shiny appearance, and less spicy taste. In plant breeding, polyploidy induction through mutagens is a technique often used to produce shallot cultivars of better quality. Bio-Catharantin from the leaf extract of Catharanthus roseus L. is used as a polyploid induction agent instead of colchicine. The latest study aimed to determine the effect of Bio-Catharantin concentration (0.2% and 0.4%) on phenotypic traits (plant height, bulb mass, and the number of bulbs), and the chromosome number to determine the minimum concentration that could cause polyploidization in shallots. The research was conducted from December 2020 to February 2021 in a greenhouse in Madurejo, Prambanan, and the Laboratory of Genetics and Breeding, Faculty of Biology, Gadjah Mada University, Indonesia. Bio-Catharantin concentration did not affect plant height which was comparable with the control. Both treatments caused an increase in bulb mass up to 37.7 and 41.76 g at the concentrations of 0.2% and 0.4%, respectively, compared with the control (31.47 g). The number of bulbs increased up to 10.6 and 9.8 g for 0.2% and 0.4% concentrations, respectively, compared with 8.8 in the control. The ploidy level of cells was increased from 2n (16) to 3n (24) at 2% and 4n (32) at 4% Bio-Catharantin.

Cytokinesis regulators as potential diagnostic and therapeutic biomarkers for human hepatocellular carcinoma.

Cytokinesis, the final step of mitosis, is critical for maintaining the ploidy level of cells. Cytokinesis is a complex, highly regulated process and its failure can lead to genetic instability and apoptosis, contributing to the development of cancer. Human hepatocellular carcinoma is often accompanied by a high frequency of aneuploidy and the DNA ploidy pattern observed in human hepatocellular carcinoma results mostly from impairments in cytokinesis. Many key regulators of cytokinesis are abnormally expressed in human hepatocellular carcinoma, and their expression levels are often correlated with patient prognosis. Moreover, preclinical studies have demonstrated that the inhibition of key cytokinesis regulators can suppress the growth of human hepatocellular carcinoma. Here, we provide an overview of the current understanding of the signaling networks regulating cytokinesis, the key cytokinesis regulators involved in the initiation and development of human hepatocellular carcinoma, and their applications as potential diagnostic and therapeutic biomarkers.

Comprehensive observation of trichome development in Micro-tom tomato

Trichomes help plants defend against various stresses by formation of a protective barrier. Although the unicellular trichomes in <i>Arabidopsis</i> have been well studied, little is known of the development of multicellular trichomes in tomato. With seven morphologically and functionally different types, tomato trichomes are more complex than the unicellular trichomes in <i>Arabidopsis</i>. Here, we comprehensively analyzed the spatiotemporal pattern of tomato trichome development. Our observations show that the gradual reduction of type I/IV trichomes during leaf development is accompanied by an increase of other trichome types. We further examined the gland development of type I/IV trichomes by constructing a marker reporter, pSlAT2: GUS-YFP which is specifically expressed in the top cell of the trichomes. Interestingly, we found the multicellular base of trichomes is initiated as early as the two-cell stage. Through tracking a nuclear marker, H2B-GFP in tomato trichomes, we found the correlation between stalk cell length and nuclear size, suggesting that cell ploidy level could affect trichome cell size. Finally, we found type VII glandular trichomes exhibit two morphologically distinct forms, with the majority covered by a wax-like substance, which could help to form extra storage space for secretory metabolites. Our results provide the basis for further mechanistic study of trichome development in tomato.

Measuring Cell Ploidy Level in Arabidopsis thaliana by Flow Cytometry.

Cell ploidy levels are regulated by developmental and environmental factors and they also impact the outcome of plant microbe interactions. Here we describe a simple and quick procedure to measure cell ploidy levels in Arabidopsis thaliana leaves by flow cytometry. Cell nuclei are isolated by filtering tissue homogenates from chopped plant tissues. DNA in the nuclei is stained by propidium iodide and the fluorescence emitted from the DNA of each nucleus is read by using a flow cytometer. Distribution of ploidy levels within the plant tissues can be calculated based on the distribution of fluorescence signals. Multiple samples can be prepared and analyzed within the same day.

Enhancement of paclitaxel content in induced tetraploid Corylus avellana L. cell suspension culture with regulating the expression of genes in paclitaxel biosynthetic pathway

During recent years, hazel cell suspension culture has been significantly considered as a new important source of paclitaxel. Artificial polyploidy alters different characters in plants which results in amplifying the secondary metabolites in medicinal plants. In this paper, the effects of tetraploidy induction on paclitaxel content and gene expression in hazel cell suspension culture were investigated. Various concentrations of colchicine and duration of exposure in solid and liquid media were examined and the ploidy level of cells was determined using flow cytometric analysis. The tetraploid cells were obtained from 0.2% colchicine in the exposure time of 5 and 6 days in solid medium and 0.3% colchicine in the exposure time of 3 and 4 days in liquid medium. Tetraploid cells were employed to prepare cell suspension. 3 µM of phenylalanine and 0.05 mM of vanadyl sulfate were added to both tetraploid and diploid (control) suspension to elicit paclitaxel induction. High performance liquid chromatography analysis demonstrated that the tetraploid cell suspensions produced paclitaxel of about [9.88 µg g−1 (DW)] 1.7-fold compared with diploid cells [5.74 µg g−1 (DW)]. The application of phenylalanine and vanadyl sulfate increased the concentration of paclitaxel in both diploid and tetraploid cells. Moreover, qRT-PCR analysis showed that the expression of GGPPS gene was significantly increased and PAL gene expression was altered after tetraploidization.

Cellular changes during Medicago truncatula hypocotyl growth depend on temperature and genotype

Hypocotyl growth is a key characteristic for plant emergence, influenced by environmental conditions, particularly temperature, and varying among genotypes. Cellular changes in Medicago truncatula hypocotyl were characterized to study the impact of the environment on heterotrophic growth and analyze differences between genotypes. The number and length of epidermal cells, ploidy levels, and sugar contents were measured in hypocotyls grown in the dark at 20°C and 10°C using two genotypes with contrasting maximum hypocotyl length. Hypocotyl elongation in the dark was due to cell elongation and not to an increase in cell number. A marked increase in cell ploidy level was observed just after germination and until mid elongation of the hypocotyl under all treatments. Larger ploidy levels were also observed in the genotype with the shorter hypocotyl and in cold conditions, but they were associated with larger cells. The increase in ploidy level and in cell volume was concomitant with a marked increase in glucose and fructose contents in the hypocotyl. Finally, differences in hypocotyl length were mainly due to different number of epidermal cells in the seed embryo, shown as a key characteristic of genotypic differences, whereas temperature during hypocotyl growth affected cell volume.

Light-Dependent Regulation ofDEL1Is Determined by the Antagonistic Action of E2Fb and E2Fc

Endoreduplication represents a variation on the cell cycle in which multiple rounds of DNA replication occur without subsequent chromosome separation and cytokinesis, thereby increasing the cellular DNA content. It is known that the DNA ploidy level of cells is controlled by external stimuli such as light; however, limited knowledge is available on how environmental signals regulate the endoreduplication cycle at the molecular level. Previously, we had demonstrated that the conversion from a mitotic cell cycle into an endoreduplication cycle is controlled by the atypical E2F transcription factor, DP-E2F-LIKE1 (DEL1), that represses the endocycle onset. Here, the Arabidopsis (Arabidopsis thaliana) DEL1 gene was identified as a transcriptional target of the classical E2Fb and E2Fc transcription factors that antagonistically control its transcript levels through competition for a single E2F cis-acting binding site. In accordance with the reported opposite effects of light on the protein levels of E2Fb and E2Fc, DEL1 transcription depended on the light regime. Strikingly, modified DEL1 expression levels uncoupled the link between light and endoreduplication in hypocotyls, implying that DEL1 acts as a regulatory connection between endocycle control and the photomorphogenic response.

Morphofunctional characterization and ploidy levels of cells in the bivalve digestive gland, with special reference to somatic polyploidy

This study investigates the histological organization and ploidy levels of the digestive gland cells in 29 species of marine, brackish water, and freshwater bivalves belonging to 5 subclasses. In all species studied, the digestive gland epithelium consists of two types of differentiated cells: digestive and basophilic. The nuclei of digestive cells contain the diploid quantity of DNA. Basophilic (secretory) cells often remain diploid also; however, in a number of species all or some of the cells showed an increase in the quantity of DNA per nucleus up to 4c. Tetraploidy of basophilic cells in several species of the subclass Anomalodesmata seems to be due to carnivory. In other species, no apparent correlation was found between selective polyploidy of basophilic cells and environmental conditions or biology traits of the bivalves. Additionally, there was no relationship between the occurrence of polyploid cells and the lifespan of the investigated species. In the bivalve mollusks, somatic polyploidy appears to be an adaptation; it is neither a tissue growth strategy nor a component of the cytodifferentiation program.

The nuclear DNA mass variability in embryo root cells of sugarbeet

Cytophotometrical study was conducted on the nuclear DNA mass variability in the embryo root cells of two inbred lines of sugar beet, with different tendency to the agamospermic mode of seed reproduction. An average DNA mass in nuclei of the line SOAN-41 (which has a tendency to agamospermy) was more than 2.5 times larger than the one of the line SOAN-210 (which has a tendency to gamospermy). Higher values of the DNA mass in the cells’ nuclei of the line SOAN-41 correlate with the disturbances of DNA distribution during the mitotic divisions. For the line SOAN-41 in telophase the daughter nuclei had different DNA mass in 80% cases, while there were only about 20% of such differences for the line SOAN-210. The results obtained do not contradict the known correlation and mutual dependence of the ploidy level of cells’ genome and the plants’ tendency to the apomictic mode of seed reproduction (agamospermy).

Localisation and interaction of the protein components of the yeast 2 mu circle plasmid partitioning system suggest a mechanism for plasmid inheritance.

Replicating plasmids are highly unstable in yeast, because they are retained in mother cells. The 2 mu circle plasmid overcomes this maternal inheritance bias by using a partitioning system that involves the plasmid encoded proteins Rep1p and Rep2p, and the cis-acting locus STB. It is thus widely exploited as a cloning vehicle in yeast. However, little is known about the cellular or molecular mechanisms by which effective partitioning is achieved, and models of both free diffusion and plasmid localisation have been proposed. Here we show that Rep1p and Rep2p proteins interact to form homo- and hetero-complexes in vitro. In vivo, Rep1p and Rep2p are shown to be nuclear proteins, exhibiting sub-nuclear concentration in distinct foci. The number of foci appears constant regardless of plasmid copy number and cell ploidy level. Before cell division, the number of foci increases, and we observe approximately equal allocation of foci to mother and daughter cell nuclei. We show that whereas Rep2p expressed alone is found exclusively in the nucleus, Rep1p requires the presence of Rep2p for effective nuclear localisation. High levels of 2 mu plasmid induce a multiple-budded elongated cell phenotype, which we show can be phenocopied by overexpression of both REP1 and REP2 together but not alone. Taken together, these results suggest that Rep1p and Rep2p interact in vivo, and occupy defined nuclear sites that are allocated to both mother and daughter nuclei during division. We propose a model for 2 mum plasmid partitioning based on these results, involving the association of plasmid DNA with specific, segregated subnuclear sites.

Variability of cardiomyocyte DNA content, ploidy level and nuclear number in mammalian hearts.

DNA content, ploidy level, cell size and nuclear number were investigated in 54 mammalian hearts from nine species. DNA content was determined biochemically and ploidy level of cells was studied by the means of Feulgen cytophotometry. Nuclear number was calculated by a new method, while cell size was determined by using ocular micrometry. In most mammals diploid cell nuclei predominate. Higher ploidy levels were found in the human and the pig hearts. The total amount of DNA correlated with the myocardial weight. Eight million heart muscle cell nuclei were found in mice (myocardial weight 160 mg), and 2600 million heart muscle cell nuclei in the human heart (myocardial weight 210 g), but in the hearts of horses up to 35000 million heart muscle cell nuclei (myocardial weight 3.4 kg) were found. The number of heart muscle and connective tissue cell nuclei was correlated with myocardial weight. The ratio of connective tissue cell nuclei to heart muscle cell nuclei was between 2:1 and 3:1. In cardiac growth this ratio shifted towards connective tissue cell nuclei. Increased heart weight corresponds to an increase in cell size. Diameter between 11 microns and 18 microns may be an optimum for heart muscle cells of mammals.

Callus growth, tumour development and polyploidization in the tetraploid potato cultivar bintje

Various combinations of exogenously applied growth regulators supported callus growth to different extents in cultures of leaf and tuber explants of the tetraploid potato cv. Bintje ( Solanum tuberosum L.). Various strains of Agrobacterium tumefaciens differing in induction of endogenous hormonal activities, induced tumours of varying habitus and frequency on tuber dics. The ploidy levels of cells from tissues were determined by means of flow cytometry. In the original explants, varying degrees of polyploidization were observed. Moreover, upon ageing of explanted leaf tissues, rapid endopolyploidization occurred. The DNA histograms of cell populations of calli or tumours were similar, irrespective of hormonal regime and explant source. Within 1 month of growth, a stable pattern of polyploidization was reached, the DNA histograms resembling the patterns found in tuber tissues and in aged leaf explants, cultured in the absence of plant growth regulatory substances. The factors influencing genetic stability, are attributed to predominant genotype effects.

Adenocarcinoma and mixed carcinoma of the uterine cervix. II. Prognostic value of nuclear DNA analysis.

The prognosis of the glandular neoplasms of the uterine cervix is related to the clinical stage and, to a lesser extent, to the histologic type, growth pattern, and degree of differentiation of the neoplasms. To determine further the prognostic significance of the stem cell ploidy levels as determined by nuclear DNA quantitation, the authors separated the tumors into low ploidy (less than 3N) and high ploidy (greater than 3N) groups. Of the clinical Stage I and II neoplasms, low ploidy tumors had a better prognosis than high ploidy tumors of comparable stage. Low ploidy tumors had a better prognosis than high ploidy tumors irrespective of the degree of differentiation. Mixed carcinomas had a poorer prognosis than pure adenocarcinomas of comparable clinical stage. This might be explained by the greater proportion of high ploidy stem cells in mixed carcinomas than in pure adenocarcinomas. Although advanced clinical Stage III and IV neoplasms had a poor outlook regardless of the DNA ploidy level, there was a proportional increase of high ploidy tumors with increasing clinical stage. These findings suggest that cervical granular neoplasms having high ploidy stem cell lines are biologically more aggressive than those with low ploidy stem cell lines.

CHANGES IN CHROMOSOME NUMBER IN MICROSPORE CALLUS OF RICE DURING SUCCESSIVE SUBCULTURES

Forty-six callus cultures of rice (Oryza sativa L., 2n = 24), each presumably originating from a single microspore, were established and maintained on a medium containing 2,4-D. At the end of the first transfer 24% of the cultures were nonhaploid consisting of only diploid or polyploid cells, or of cells of two ploidy levels. Nuclear fusion and endomitosis occurring during the initial stages of in vitro microspore development were postulated to account for the formation of nonhaploid callus. Seventeen cultures were studied cytologically through 19 transfers. Only in one tetraploid and one hexaploid callus did the ploidy levels of cells remain unchanged during culture. Chromosome numbers in 13 cultures fell into a geometric series. Since no diplo- and quadruplochromosomes were observed, it was inferred that endomitosis rather than endoreduplication was responsible for the changes. In spite of the tendency for chromosome doubling, the proportions of cells of different ploidy levels were fixed in the 13 cultures at later transfers. Haploid cells were eliminated from all cultures. Diploid cells became predominant in eight cultures and tetraploid cells in five, suggesting a selection for either cell type. Triploids appeared in two cultures which initially did not contain this type of cell. Limited cytological information indicated that triploid cells might have originated from tetraploid cells through reductional grouping of chromosomes accompanied by multipolar formation.

Cytological studies on the mosaic silkworms induced by low temperature treatment

Mosaic silkworms were induced when the hybrid eggs of two strains with different egg color and larval markings were exposed to low temperature. Cytological studies were conducted to find out the relation between mosaic larval pattern and ploidy in reproductive cells along with demonstration of chromosomes in the mosaic embryos. — Mosaic eggs eith the characters of both the father and the hybrid were two types, one with large serosa nuclei (LN-mosaic) and the other with small serosa nuclei (SN-mosaic). The cytological studies demonstrated that LN-mosaic individuals were 2n/4n, while SN-mosaic ones were n/2n. In both types of silkworms, cell ploidy level in nuclei of synkaryon origin was two times that of androgenic ones. — From the results obtained in the present studies as well as in the previous studies, a possible mechanism of induction of mosaicism in silkworms by cold shock is contemplated.

Simultaneous staining of ribonucleic and deoxyribonucleic acids in unfixed cells using acridine orange in a flow cytofluorometric system.

Simultaneous staining of deoxyribonucleic (DNA) and ribonucleic acid (RNA) in nonfixed, but permeable, cells is described. Cells are made permeable by treatment with non-ionic detergent at low pH. RNA is denatured prior to, or during staining, by exposure of cells to chelating agents to ensure that DNA (native) and RNA (dentured) may be stained differentially with the metachromatic dye, acridine orange. The fluorescence of individual cells is measured in a flow cytofluorometer. A comparison between various staining procedures employing acridine orange or other intercalating dyes in unfixed cells is discussed in terms of staining specificity, cell permeability and preservation. Evidence is provided that acridine orange staining of unfixed cells may be used as a simple, fast means of obtaining information on cell ploidy levels and cell cycle status from DNA measurements (green fluorescence), and cell transcriptional activity from RNA staining (red fluorescence), in human and murine cells lines, peripheral blood and bone marrow specimens from patients with leukemia and mitogenically (phytohemagglutinin) or antigenically (mixed lymphocyte culture) stimulated human peripheral blood cultures. Exposure of cells to detergent at low pH as an alternative to cell fixation or hypotonic treatment is proposed as a fast, convenient method of making cells permeable to dyes.