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Cell differentiation by non-identical copies

Cell differentiation by non-identical copies


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There are in principle two ways that two cells with the same mother cell can differentiate/specialize/diverge:

  1. Because they are not perfect identical copies (= different internal influencing factors). I explictly don't mean "non-identical on the DNA level", but on the level of other cellular components!

  2. Because they are exposed to different external influencing factors (chemical, mechanical).

Can it be said, which of these two reasons of cell differentiation is the more common, frequent, resp. "important"? Or does it completely depend on the (type of mother) cell and some general circumstances?

If internal factors (= non-identical copies) are an important source of differentiation, I have two (optional) questions:

  1. How does the mother cell manage to cleave into two non-identical copies in a rather fine-tuned and determinstic way? Or is any kind of asymmetry enough?

  2. Which is the first cell (starting with the zygote) that cleaves into two non-indentical cells, different enough to give rise to differentiation? (From this answer I believe to have learned that this might happen already after 1-4 divisions.)


I found a partial answer to my and this question at Quora:

What regulates initial cell differentiation?

Ian Driver says:

In many organisms the initial cell has mRNA (or protein) partitioned in one half of the zygote. One daughter cell inherits most of the mRNA or protein and goes on to activate or repress genes in that cell, but not in the other. This has been visualized and well studied in C. elegans: Asymmetric cell division and axis formation in the embryo.


This question is kind of bad. There are common internal influencing factors that are unrelated to improper replication, most commonly:

Methylation. Addition of methyl groups to DNA regions inhibits RNA transcription from that region.

Imprinting. Can't recall exactly how this works but it's another mechanism for enabling/inactivating genes.

Histone structure (wrapping/unwrapping). DNA coiled tightly around histones is more unlikely to be transcribed.

External influencing factors include cell signaling, substrate detection, and large-scale endocrine signaling. Most important are growth factors and growth inhibitors (which let the cells know if they need to replicate quickly (repairing damage/large-scale growth) or slow down replication (overcrowding). Other external factors could include metabolism (produce these digestive enzymes when exposed to these foods) or endocrine signaling.

Non-identical copies at the DNA level (mutation) are thankfully relatively rare in practice, particularly among higher-order species with longer lifespans.


Generation and characterization of an immortalized human mesenchymal stromal cell line

Human mesenchymal stromal cells (hMSCs) show great potential for clinical and experimental use due to their capacity to self-renew and differentiate into multiple mesenchymal lineages. However, disadvantages of primary cultures of hMSCs are the limited in vitro lifespan, and the variable properties of cells from different donors and over time in culture. In this article, we describe the generation of a telomerase-immortalized nontumorigenic human bone marrow-derived stromal mesenchymal cell line, and its detailed characterization after long-term culturing (up to 155 population doublings). The resulting cell line, iMSC#3, maintained a fibroblast-like phenotype comparable to early passages of primary hMSCs, and showed no major differences from hMSCs regarding surface marker expression. Furthermore, iMSC#3 had a normal karyotype, and high-resolution array comparative genomic hybridization confirmed normal copy numbers. The gene expression profiles of immortalized and primary hMSCs were also similar, whereas the corresponding DNA methylation profiles were more diverse. The cells also had proliferation characteristics comparable to primary hMSCs and maintained the capacity to differentiate into osteoblasts and adipocytes. A detailed characterization of the mRNA and microRNA transcriptomes during adipocyte differentiation also showed that the iMSC#3 recapitulates this process at the molecular level. In summary, the immortalized mesenchymal cells represent a valuable model system that can be used for studies of candidate genes and their role in differentiation or oncogenic transformation, and basic studies of mesenchymal biology.

Figures

Analyses of morphology, proliferation rates,…

Analyses of morphology, proliferation rates, surface markers, and ectopic expression of TERT .…

Analyses of genome-wide gene expression…

Analyses of genome-wide gene expression and DNA methylation of iMSC#3 and primary hMSCs.…

Osteoblast differentiation of iMSC#3. (A)…

Osteoblast differentiation of iMSC#3. (A) iMSC#3 cells was differentiated for 28 days before…

Adipocyte differentiation of iMSC#3. (A)…

Adipocyte differentiation of iMSC#3. (A) iMSC#3 cells were differentiated for 21 days before…

High-throughput sequencing of miRNAs during…

High-throughput sequencing of miRNAs during adipogenesis of iMSC#3. (A) Twenty-five most up- and…


Cell differentiation by non-identical copies - Biology

Unit 1: The organisation of the human body 1 2 3 4 5

3. Cell variety and differentiation

The human body has enormous amount of cells. The exactly number is unknown, but it is estimated that it is closed to 70 millions of millions.

Although all cells of our body have a similar structure, they are non identical.

There are differences among them about shape, size, functions, etc.

It seems that exist almost 250 kinds of cells in the human body.

Characteristics of each cell type depend on their function, the tissue that they form part and the cells around it.

When a person develops and grows, cells divide and rise in number. In addition these cells change and take a particular characteristics about shape, size, organelles, etc., which make them especially able to do a particular function. This process is called cellular differentiation or specialisation.

READING ACTIVITIES

After reading the text, copy and answer the following questions into your notebook:

Remember: you must make complete sentences.

3.1. What is the reason for the enormous variety of cellular types

in the human body?

3.2. Is it possible to see a human ovule with the naked eye?


Roles of DNA and RNA in Cell Differentiation

Dexoyribonucleic Acid, or DNA, controls the way cells function. It also determines what type of specialized cells will be made. Stem cells are cells that have the ability to become any type of specialized cell in the body. After an egg cell and sperm cell unite to begin forming a new organism, all of the DNA in each cell of that organism will be virtually identical. If every part of the DNA in each cell is the same, then how do cells become different types of cells? Let’s look more closely at DNA to find out.

DNA is wound tightly into chromosomes. Different regions of the chromosome code for every different function and cell type. Not all sections of a chromosome are turned on, or expressed, at the same time. Only the regions that are needed to perform a specific function are expressed in each cell. These regions are often depicted as bands or stripes on a drawing of a chromosome. These bands are called genes, and whether or not a gene is expressed determines what type of cell will be created. For example, genes that are expressed (turned on) in a nerve cell are different from the genes that are expressed in a muscle cell. Both cells have the same DNA, but expressing different genes generates different cell types.

This process by which information from a gene is used to make the structures of a cell is called gene expression. Since RNA translates and transcribes the DNA code into proteins (the structures of a cell), it also plays a role in cell differentiation.


Directed differentiation of regulatory T cells from naive T cells and prevention of their inflammation-mediated instability using small molecules

E. Hajizadeh-Saffar, Department of Regenerative Medicine, Cell Science Research Center, Royan Institute for Stem Cell Biology and Technology, ACECR, Tehran, Iran.

H. Baharvand, Department of Stem Cells and Developmental Biology, Cell Science Research Center, Royan Institute for Stem Cell Biology and Technology, ACECR, Tehran, Iran.

Department of Regenerative Medicine, Cell Science Research Center, Royan Institute for Stem Cell Biology and Technology, ACECR, Tehran, Iran

Correspondence: B. Negahdari, Department of Medical Biotechnology, School of Advanced Technologies in Medicine, Tehran University of Medical Sciences, Tehran, Iran

E. Hajizadeh-Saffar, Department of Regenerative Medicine, Cell Science Research Center, Royan Institute for Stem Cell Biology and Technology, ACECR, Tehran, Iran.

H. Baharvand, Department of Stem Cells and Developmental Biology, Cell Science Research Center, Royan Institute for Stem Cell Biology and Technology, ACECR, Tehran, Iran.

Columbia Center for Translational Immunology, Department of Medicine, Columbia University College of Physicians and Surgeons, New York, NY, USA

Department of Stem Cells and Developmental Biology, Cell Science Research Center, Royan Institute for Stem Cell Biology and Technology, ACECR, Tehran, Iran

Department of Regenerative Medicine, Cell Science Research Center, Royan Institute for Stem Cell Biology and Technology, ACECR, Tehran, Iran

Department of Stem Cells and Developmental Biology, Cell Science Research Center, Royan Institute for Stem Cell Biology and Technology, ACECR, Tehran, Iran

Department of Developmental Biology, University of Science and Culture, Tehran, Iran

Correspondence: B. Negahdari, Department of Medical Biotechnology, School of Advanced Technologies in Medicine, Tehran University of Medical Sciences, Tehran, Iran

E. Hajizadeh-Saffar, Department of Regenerative Medicine, Cell Science Research Center, Royan Institute for Stem Cell Biology and Technology, ACECR, Tehran, Iran.

H. Baharvand, Department of Stem Cells and Developmental Biology, Cell Science Research Center, Royan Institute for Stem Cell Biology and Technology, ACECR, Tehran, Iran.

Department of Medical Biotechnology, School of Advanced Technologies in Medicine, Tehran University of Medical Sciences, Tehran, Iran

Clinical Microbiology Research Center, Ilam University of Medical Sciences, Ilam, Iran

Department of Medical Biotechnology, School of Advanced Technologies in Medicine, Tehran University of Medical Sciences, Tehran, Iran

Correspondence: B. Negahdari, Department of Medical Biotechnology, School of Advanced Technologies in Medicine, Tehran University of Medical Sciences, Tehran, Iran

E. Hajizadeh-Saffar, Department of Regenerative Medicine, Cell Science Research Center, Royan Institute for Stem Cell Biology and Technology, ACECR, Tehran, Iran.

H. Baharvand, Department of Stem Cells and Developmental Biology, Cell Science Research Center, Royan Institute for Stem Cell Biology and Technology, ACECR, Tehran, Iran.

Department of Regenerative Medicine, Cell Science Research Center, Royan Institute for Stem Cell Biology and Technology, ACECR, Tehran, Iran

Correspondence: B. Negahdari, Department of Medical Biotechnology, School of Advanced Technologies in Medicine, Tehran University of Medical Sciences, Tehran, Iran

E. Hajizadeh-Saffar, Department of Regenerative Medicine, Cell Science Research Center, Royan Institute for Stem Cell Biology and Technology, ACECR, Tehran, Iran.

H. Baharvand, Department of Stem Cells and Developmental Biology, Cell Science Research Center, Royan Institute for Stem Cell Biology and Technology, ACECR, Tehran, Iran.

Columbia Center for Translational Immunology, Department of Medicine, Columbia University College of Physicians and Surgeons, New York, NY, USA

Department of Stem Cells and Developmental Biology, Cell Science Research Center, Royan Institute for Stem Cell Biology and Technology, ACECR, Tehran, Iran

Department of Regenerative Medicine, Cell Science Research Center, Royan Institute for Stem Cell Biology and Technology, ACECR, Tehran, Iran

Department of Stem Cells and Developmental Biology, Cell Science Research Center, Royan Institute for Stem Cell Biology and Technology, ACECR, Tehran, Iran

Department of Developmental Biology, University of Science and Culture, Tehran, Iran

Correspondence: B. Negahdari, Department of Medical Biotechnology, School of Advanced Technologies in Medicine, Tehran University of Medical Sciences, Tehran, Iran

E. Hajizadeh-Saffar, Department of Regenerative Medicine, Cell Science Research Center, Royan Institute for Stem Cell Biology and Technology, ACECR, Tehran, Iran.

H. Baharvand, Department of Stem Cells and Developmental Biology, Cell Science Research Center, Royan Institute for Stem Cell Biology and Technology, ACECR, Tehran, Iran.

Summary

Regulatory T (Treg) cell therapy is a promising approach for immune tolerance induction in autoimmunity conditions and cell/organ transplantations. Insufficient isolation yields and impurity during downstream processes and Treg instability after adoptive transfer in inflammatory conditions are major limitations to Treg therapy, and indicate the importance of seeking a valid, reliable method for de-novo generation of Tregs. In this research, we evaluated Treg-like cells obtained from different Treg differentiation protocols in terms of their yield, purity and activity. Differentiation was performed on naive CD4 + cells and a naive CD4 + /Treg co-culture by using three different protocols – ectopic expression of forkhead box protein P3 (E-FoxP3), soluble transforming growth factor β (S-TGF) and small molecules [N-acetyl puromycin and SR1555 (N-Ac/SR)]. The results showed that a high yield of a homogeneous population of Treg-like cells could be achieved by the N-Ac/SR method under a T helper type 17 (Th17)-polarizing condition, particularly interleukin (IL)-6 and TGF-β, when compared with the E-FoxP3 and S-TGF methods. Surprisingly, SR completely inhibited the differentiation of IL-17-producing cells and facilitated Treg generation in the inflammatory condition and had highly suppressive activity against T cell proliferation without Treg-specific demethylase region (TSDR) demethylation. For the first time, to our knowledge, we report the generation of efficient, pure Treg-like cells by using small molecules during in-vitro inflammatory conditions. Our results suggested that the N-Ac/SR method has several advantages for Treg generation when compared with the other methods, including a higher purity of Tregs, easier procedure, superior suppressive activity during the inflammatory condition and decreased cost.

Fig. S1. Different levels of forkhead box protein P3 (FoxP3) + cells in different concentrations of IL-2. The abundance of FoxP3 + cells after naïve T cell activation for 1:3 cells/beads at 100 and 200 IU/ml IL-2 as analyzed by flow cytometry.

Fig. S2. Optimization of efficient transduction method. Double transduction was performed in order to obtain a high transduction rate. Different levels of forkhead box protein P3 (FoxP3) + cells in different concentrations of IL-2. The abundance of FoxP3 + cells after naïve T cell activation for 1:3 cells/beads at 100 and 200 IU/ml IL-2 as analyzed by flow cytometry.

Fig. S3. Comparison of percentages of Treg-like cell in E-FoxP3 and control, control vector groups.

Fig. S4. Regulatory T (Treg) cell generation from naïve T cells via a combination of small molecules. (a) The percentage of the regulatory T (Treg) cell phenotype as determined by flow cytometry. (b) Comparison of suppressive activities between the N-acetyl puromycin and SR1555 (N-Ac/SR) group and the control group. Conventional T (Tconv) cells were the negative control.

Fig. S5. Representative plots of suppressive activity.

Fig. S6. CD4 + CD25 + FoxP3 + regulatory T (Treg) cell generation during naïve/Treg cell co-culture. The level of FoxP3 + cells was similar in the soluble TGF-β (S-TGF) and N-acetyl puromycin and SR1555 (N-Ac/SR) groups. The combination of small molecules considerably increased forkhead box protein P3 (FoxP3) + levels compared with the control group.

Please note: The publisher is not responsible for the content or functionality of any supporting information supplied by the authors. Any queries (other than missing content) should be directed to the corresponding author for the article.


The direct and indirect regulation of follicular T helper cell differentiation in inflammation and cancer

Key Laboratory of Cell Proliferation and Regulation Biology, Ministry of Education, Institute of Cell Biology, College of Life Sciences, Beijing Normal University, No. 19, Xinjiekouwai Street, Haidian District, Beijing, China

Key Laboratory of Cell Proliferation and Regulation Biology, Ministry of Education, Institute of Cell Biology, College of Life Sciences, Beijing Normal University, No. 19, Xinjiekouwai Street, Haidian District, Beijing, China

Key Laboratory of Cell Proliferation and Regulation Biology, Ministry of Education, Institute of Cell Biology, College of Life Sciences, Beijing Normal University, No. 19, Xinjiekouwai Street, Haidian District, Beijing, China

Immunochina Pharmaceuticals Co., Ltd., No. 80, Xingshikou Road, Haidian District, Beijing, China

Key Laboratory of Cell Proliferation and Regulation Biology, Ministry of Education, Institute of Cell Biology, College of Life Sciences, Beijing Normal University, No. 19, Xinjiekouwai Street, Haidian District, Beijing, China

Key Laboratory of Cell Proliferation and Regulation Biology, Ministry of Education, Institute of Cell Biology, College of Life Sciences, Beijing Normal University, No. 19, Xinjiekouwai Street, Haidian District, Beijing, China

Key Laboratory of Cell Proliferation and Regulation Biology, Ministry of Education, Institute of Cell Biology, College of Life Sciences, Beijing Normal University, No. 19, Xinjiekouwai Street, Haidian District, Beijing, China

State Key Laboratory of Pathogen and Biosecurity, Beijing Institute of Microbiology and Epidemiology, No. 20, East Street, Fengtai District, Beijing, China

Correspondence Yujing Bi , State Key Laboratory of Pathogen and Biosecurity, Beijing Institute of Microbiology and Epidemiology, Beijing, 100071, China.

Guangwei Liu, Key Laboratory of Cell Proliferation and Regulation Biology, Ministry of Education, Institute of Cell Biology, College of Life Sciences, Beijing Normal University, Beijing, 100875, China.

Key Laboratory of Cell Proliferation and Regulation Biology, Ministry of Education, Institute of Cell Biology, College of Life Sciences, Beijing Normal University, No. 19, Xinjiekouwai Street, Haidian District, Beijing, China

Correspondence Yujing Bi , State Key Laboratory of Pathogen and Biosecurity, Beijing Institute of Microbiology and Epidemiology, Beijing, 100071, China.

Guangwei Liu, Key Laboratory of Cell Proliferation and Regulation Biology, Ministry of Education, Institute of Cell Biology, College of Life Sciences, Beijing Normal University, Beijing, 100875, China.

Key Laboratory of Cell Proliferation and Regulation Biology, Ministry of Education, Institute of Cell Biology, College of Life Sciences, Beijing Normal University, No. 19, Xinjiekouwai Street, Haidian District, Beijing, China

Key Laboratory of Cell Proliferation and Regulation Biology, Ministry of Education, Institute of Cell Biology, College of Life Sciences, Beijing Normal University, No. 19, Xinjiekouwai Street, Haidian District, Beijing, China

Key Laboratory of Cell Proliferation and Regulation Biology, Ministry of Education, Institute of Cell Biology, College of Life Sciences, Beijing Normal University, No. 19, Xinjiekouwai Street, Haidian District, Beijing, China

Immunochina Pharmaceuticals Co., Ltd., No. 80, Xingshikou Road, Haidian District, Beijing, China

Key Laboratory of Cell Proliferation and Regulation Biology, Ministry of Education, Institute of Cell Biology, College of Life Sciences, Beijing Normal University, No. 19, Xinjiekouwai Street, Haidian District, Beijing, China

Key Laboratory of Cell Proliferation and Regulation Biology, Ministry of Education, Institute of Cell Biology, College of Life Sciences, Beijing Normal University, No. 19, Xinjiekouwai Street, Haidian District, Beijing, China

Key Laboratory of Cell Proliferation and Regulation Biology, Ministry of Education, Institute of Cell Biology, College of Life Sciences, Beijing Normal University, No. 19, Xinjiekouwai Street, Haidian District, Beijing, China

State Key Laboratory of Pathogen and Biosecurity, Beijing Institute of Microbiology and Epidemiology, No. 20, East Street, Fengtai District, Beijing, China

Correspondence Yujing Bi , State Key Laboratory of Pathogen and Biosecurity, Beijing Institute of Microbiology and Epidemiology, Beijing, 100071, China.

Guangwei Liu, Key Laboratory of Cell Proliferation and Regulation Biology, Ministry of Education, Institute of Cell Biology, College of Life Sciences, Beijing Normal University, Beijing, 100875, China.

Key Laboratory of Cell Proliferation and Regulation Biology, Ministry of Education, Institute of Cell Biology, College of Life Sciences, Beijing Normal University, No. 19, Xinjiekouwai Street, Haidian District, Beijing, China

Correspondence Yujing Bi , State Key Laboratory of Pathogen and Biosecurity, Beijing Institute of Microbiology and Epidemiology, Beijing, 100071, China.

Guangwei Liu, Key Laboratory of Cell Proliferation and Regulation Biology, Ministry of Education, Institute of Cell Biology, College of Life Sciences, Beijing Normal University, Beijing, 100875, China.

Yejin Cao, Lin Dong, Ying He, and Xuelian Hu contributed equally to this work as co-first author.

Abstract

Follicular T helper (Tfh) cells play important roles in facilitating B-cell differentiation and inducing the antibody response in humoral immunity and immune-associated inflammatory diseases, including infections, autoimmune diseases, and cancers. However, Tfh cell differentiation is mainly achieved through self-directed differentiation regulation and the indirect regulation mechanism of antigen-presenting cells (APCs). During the direct intrinsic differentiation of naïve CD4 + T cells into Tfh cells, Bcl-6, as the characteristic transcription factor, plays the core role of transcriptional regulation. APCs indirectly drive Tfh cell differentiation mainly by changing cytokine secretion mechanisms. Altered metabolic signaling is also critically involved in Tfh cell differentiation. This review summarizes the recent progress in understanding the direct and indirect regulatory signals and metabolic mechanisms of Tfh cell differentiation and function in immune-associated diseases.


Revisiting Mitochondrial Function and Metabolism in Pluripotent Stem Cells: Where Do We Stand in Neurological Diseases?

Pluripotent stem cells (PSCs) are powerful cellular tools that can generate all the different cell types of the body, and thus overcome the often limited access to human disease tissues this becomes highly relevant when aiming to investigate cellular (dys)function in diseases affecting the central nervous system. Recent studies have demonstrated that PSC and differentiated cells show altered mitochondrial function and metabolic profiles and production of reactive oxygen species. This raises an emerging paradigm about the role of mitochondria in stem cell biology and urges the need to identify mitochondrial pathways involved in these processes. In this respect, this review focuses on the metabolic profile of PSC and how mitochondrial function can influence the reprogramming and differentiation processes. Indeed, both embryonic stem cells (ESCs) and induced pluripotent stem cells (iPSCs) favor the glycolytic pathway as a major source of energy production over oxidative phosphorylation. PSC mitochondria are characterized by a spherical shape, low copy number of mitochondrial DNA, and a hyperpolarized state. Indeed, mitochondria appear to have a crucial role in reprogramming iPSC, in the maintenance of a pluripotent state, and in differentiation. Moreover, an increase in mitochondrial oxidative phosphorylation has to occur for differentiation to succeed. Therefore, in vitro differentiation of neural stem cells (NSCs) into neurons can be compromised if those mechanisms are impaired. Future research should shed light on how mitochondrial impairment occurring in pre differentiation neural stages (e.g., in NSC or premature neurons) may contribute for the etiopathogenesis of neurodevelopmental and neurological disorders.

Keywords: Embryonic stem cells Energy metabolism Glycolysis Induced pluripotent stem (iPS) cells Mitochondria Neurodegenerative diseases Neuropsychiatric disorders Pluripotent stem cells.


Abstract

The roles of cyclins and their catalytic partners, the cyclin-dependent kinases (CDKs), as core components of the machinery that drives cell cycle progression are well established. Increasing evidence indicates that mammalian cyclins and CDKs also carry out important functions in other cellular processes, such as transcription, DNA damage repair, control of cell death, differentiation, the immune response and metabolism. Some of these non-canonical functions are performed by cyclins or CDKs, independently of their respective cell cycle partners, suggesting that there was a substantial divergence in the functions of these proteins during evolution.


Contents

Plant species representing all major land plant groups have been shown to be capable of producing callus in tissue culture. [2] [3] [4] [5] [6] [7] [8] [9] [10] [11] [12] A callus cell culture is usually sustained on gel medium. Callus induction medium consists of agar and a mixture of macronutrients and micronutrients for the given cell type. There are several types of basal salt mixtures used in plant tissue culture, but most notably modified Murashige and Skoog medium, [13] White's medium, [14] and woody plant medium. [15] Vitamins are also provided to enhance growth such as Gamborg B5 vitamins. [16] For plant cells, enrichment with nitrogen, phosphorus, and potassium is especially important. Plant callus is usually derived from somatic tissues. The tissues used to initiate callus formation depends on plant species and which tissues are available for explant culture. The cells that give rise to callus and somatic embryos usually undergo rapid division or are partially undifferentiated such as meristematic tissue. In alfalfa, Medicago truncatula, however callus and somatic embryos are derived from mesophyll cells that undergo dedifferentiation. [17] Plant hormones are used to initiate callus growth.

Specific auxin to cytokinin ratios in plant tissue culture medium give rise to an unorganized growing and dividing mass of callus cells. Callus cultures are often broadly classified as being either compact or friable. Friable calluses fall apart easily, and can be used to generate cell suspension cultures. Callus can directly undergo direct organogenesis and/or embryogenesis where the cells will form an entirely new plant. This process is known as callus culture. [ citation needed ]

Callus can brown and die during culture, mainly due to oxidation of phenolic compounds. In Jatropha curcas callus cells, small organized callus cells became disorganized and varied in size after browning occurred. [18] Browning has also been associated with oxidation and phenolic compounds in both explant tissues and explant secretions. [19] In rice, presumably, a condition which is favorable for scutellar callus induction induces necrosis too. [20]

Callus cells are not necessarily genetically homogeneous because a callus is often made from structural tissue, not individual cells. [ clarification needed ] Nevertheless, callus cells are often considered similar enough for standard scientific analysis to be performed as if on a single subject. For example, an experiment may have half a callus undergo a treatment as the experimental group, while the other half undergoes a similar but non-active treatment as the control group.

Plant calluses derived from many different cell types can differentiate into a whole plant, a process called regeneration, through addition of plant hormones to the culture medium. This ability is known as totipotency. Regeneration of a whole plant from a single cell allows transgenics researchers to obtain whole plants which have a copy of the transgene in every cell. Regeneration of a whole plant that has some genetically transformed cells and some untransformed cells yields a chimera. In general, chimeras are not useful for genetic research or agricultural applications.

Genes can be inserted into callus cells using biolistic bombardment, also known as a gene gun, or Agrobacterium tumefaciens. Cells that receive the gene of interest can then be recovered into whole plants using a combination of plant hormones. The whole plants that are recovered can be used to experimentally determine gene function(s), or to enhance crop plant traits for modern agriculture.

Callus is of particular use in micropropagation where it can be used to grow genetically identical copies of plants with desirable characteristics.

Henri-Louis Duhamel du Monceau investigated wound-healing responses in elm trees, and was the first to report formation of callus on live plants. [21]

In 1908, E. F. Simon was able to induce callus from poplar stems that also produced roots and buds. [22] The first reports of callus induction in vitro came from three independent researchers in 1939. [23] P. White induced callus derived from tumor-developing procambial tissues of hybrid Nicotiana glauca that did not require hormone supplementation. [14] Gautheret and Nobecourt were able to maintain callus cultures of carrot using auxin hormone additions. [ citation needed ]


Contents

Endoreplicating cell types that have been studied extensively in model organisms

Organism Cell type Biological function Citation
fly larval tissues (incl. salivary glands) secretion, embryogenesis [6]
fly ovarian follicle, nurse cells nourishment, protection of oocytes [7]
rodent megakaryocyte platelet formation [8]
rodent hepatocyte regeneration [9]
rodent trophoblast giant cell placental development, nourishment of embryo [10]
plant trichome defense from herbivory, homeostasis [11]
plant leaf epidermal cell leaf size, structure [12]
plant endosperm nourishment of embryo [13]
nematode hypodermis secretion, body size [14]
nematode intestine unknown [15]

Endoreplication, endomitosis and polytenization are three somewhat different processes resulting in polyploidization of a cell in a regulated manner. In endoreplication cells skip M phase completely, resulting in a mononucleated polyploid cell. Endomitosis is a type of cell cycle variation where mitosis is initiated, but some of the processes are not completed. Depending on how far the cell progresses through mitosis, this will give rise to a mononucleated or binucleated polyploid cell. Polytenization arises with under- or overamplification of some genomic regions, creating polytene chromosomes. [3] [4]

Based on the wide array of cell types in which endoreplication occurs, a variety of hypotheses have been generated to explain the functional importance of this phenomenon. [1] [2] Unfortunately, experimental evidence to support these conclusions is somewhat limited:

Cell/organism size Edit

Cell ploidy often correlates with cell size, [12] [14] and in some instances, disruption of endoreplication results in diminished cell and tissue size [16] suggesting that endoreplication may serve as a mechanism for tissue growth. Relative to mitosis, endoreplication does not require cytoskeletal rearrangement or the production of new cell membrane and it often occurs in cells that have already differentiated. As such it may represent an energetically efficient alternative to cell proliferation among differentiated cell types that can no longer afford to undergo mitosis. [17] While evidence establishing a connection between ploidy and tissue size is prevalent in the literature, contrary examples also exist. [18]

Cell differentiation Edit

In developing plant tissues the transition from mitosis to endoreplication often coincides with cell differentiation and morphogenesis. [18] However it remains to be determined whether endoreplication and polypoidy contribute to cell differentiation or vice versa. Targeted inhibition of endoreplication in trichome progenitors results in the production of multicellular trichomes that exhibit relatively normal morphology, but ultimately dedifferentiate and undergo absorption into the leaf epidermis. [19] This result suggests that endoreplication and polyploidy may be required for the maintenance of cell identity.

Oogenesis and embryonic development Edit

Endoreplication is commonly observed in cells responsible for the nourishment and protection of oocytes and embryos. It has been suggested that increased gene copy number might allow for the mass production of proteins required to meet the metabolic demands of embryogenesis and early development. [1] Consistent with this notion, mutation of the Myc oncogene in Drosophila follicle cells results in reduced endoreplication and abortive oogenesis. [20] However, reduction of endoreplication in maize endosperm has limited effect on the accumulation of starch and storage proteins, suggesting that the nutritional requirements of the developing embryo may involve the nucleotides that comprise the polyploid genome rather than the proteins it encodes. [21]

Buffering the genome Edit

Another hypothesis is that endoreplication buffers against DNA damage and mutation because it provides extra copies of important genes. [1] However, this notion is purely speculative and there is limited evidence to the contrary. For example, analysis of polyploid yeast strains suggests that they are more sensitive to radiation than diploid strains. [22]

Stress response Edit

Research in plants suggests that endoreplication may also play a role in modulating stress responses. By manipulating expression of E2fe (a repressor of endocycling in plants), researchers were able to demonstrate that increased cell ploidy lessens the negative impact of drought stress on leaf size. [23] Given that the sessile lifestyle of plants necessitates a capacity to adapt to environmental conditions, it is appealing to speculate that widespread polyploidization contributes to their developmental plasticity

The best-studied example of a mitosis-to-endocycle transition occurs in Drosophila follicle cells and is activated by Notch signaling. [24] Entry into endocycles involves modulation of mitotic and S-phase cyclin-dependent kinase (CDK) activity. [25] Inhibition of M-phase CDK activity is accomplished via transcriptional activation of Cdh/fzr and repression of the G2-M regulator string/cdc25. [25] [26] Cdh/fzr is responsible for activation of the anaphase-promoting complex (APC) and subsequent proteolysis of the mitotic cyclins. String/cdc25 is a phosphatase that stimulates mitotic cyclin-CDK complex activity. Upregulation of S-phase CDK activity is accomplished via transcriptional repression of the inhibitory kinase dacapo. Together, these changes allow for the circumvention of mitotic entry, progression through G1, and entry into S-phase. The induction of endomitosis in mammalian megakaryocytes involves activation of the c-mpl receptor by the thrombopoietin (TPO) cytokine and is mediated by ERK1/2 signaling. [27] As with Drosophila follicle cells, endoreplication in megakaryocytes results from activation of S-phase cyclin-CDK complexes and inhibition of mitotic cyclin-CDK activity. [28] [29]

Entry into S-phase during endoreplication (and mitosis) is regulated through the formation of a prereplicative complex (pre-RC) at replication origins, followed by recruitment and activation of the DNA replication machinery. In the context of endoreplication these events are facilitated by an oscillation in cyclin E-Cdk2 activity. Cyclin E-Cdk2 activity drives the recruitment and activation of the replication machinery, [30] but it also inhibits pre-RC formation, [31] presumably to ensure that only one round of replication occurs per cycle. Failure to maintain control over pre-RC formation at replication origins results in a phenomenon known as “rereplication” which is common in cancer cells. [2] The mechanism by which cyclin E-Cdk2 inhibits pre-RC formation involves downregulation of APC-Cdh1-mediated proteolysis and accumulation of the protein Geminin, which is responsible for sequestration of the pre-RC component Cdt1. [32] [33]

Oscillations in Cyclin E-Cdk2 activity are modulated via transcriptional and post-transcriptional mechanisms. Expression of cyclin E is activated by E2F transcription factors that were shown to be required for endoreplication. [34] [35] [36] Recent work suggests that observed oscillations in E2F and cyclin E protein levels result from a negative-feedback loop involving Cul4-dependent ubiquitination and degradation of E2F. [37] Post-transcriptional regulation of cyclin E-Cdk2 activity involves Ago/Fbw7-mediated proteolytic degradation of cyclin E [38] [39] and direct inhibition by factors such as Dacapo and p57. [40] [41] True endomitosis in the anther tapetum of the liliaceous plant Eremurus is described. The nuclear membrane does not disappear, but during metaphase the chromosomes are condensed, often considerably more than in normal mitosis. When the pollen mother cells (PMCs) go through the last premeiotic mitosis, the tapetal cells have one diploid nucleus which divides while the cell remains undivided. The two diploid nuclei may undergo an endomitosis and the resulting tetraploid nuclei a second endomitosis. An alternative pathway is an ordinary mitosis-again without cell division instead of one of the endomitotic cycles. The cytological picture in the tapetum is further complicated by restitution in anaphase and fusion of metaphase and anaphase groups during mitosis, processes which could give rise to cells with one, two, or three nuclei, instead of the expected two or four. No sign of the so-called "inhibited" mitosis is seen in these tapetal cells. When the PMCs are in leptotene-zygotene, very few tapetal nuclei are in endomitosis. When the PMCs have reached diplotene, almost 100% of cells which are not in interphase show an endomitotic stage.

Polyploidy and aneuploidy are common phenomena in cancer cells. [42] Given that oncogenesis and endoreplication likely involve subversion of common cell cycle regulatory mechanisms, a thorough understanding of endoreplication may provide important insights for cancer biology.

The unisexual salamanders (genus Ambystoma) are the oldest known unisexual vertebrate lineage, having arisen about 5 million years ago. [43] In these polyploid unisexual females, an extra premeiotic endomitotic replication of the genome, doubles the number of chromosomes. [44] As a result, the mature eggs that are produced subsequent to the two meiotic divisions have the same ploidy as the somatic cells of the adult female salamander. Synapsis and recombination during meiotic prophase I in these unisexual females is thought to ordinarily occur between identical sister chromosomes and occasionally between homologous chromosomes. Thus little, if any, genetic variation is produced. Recombination between homeologous chromosomes occurs rarely, if at all. [44] Since production of genetic variation is weak, at best, it is unlikely to provide a benefit sufficient to account for the maintenance of meiosis for millions of years. Perhaps the efficient recombinational repair of DNA damages at each generation provided by meiosis has been a sufficient advantage to maintain meiosis. [ citation needed ]


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