23 Sep 2021

Cytodifferentiation and Tissue Culture

Anjali Singh, MS

As a content and community manager, I leverage my expertise in plant biotechnology, passion for tissue culture, and writing skills to create compelling articles, simplifying intricate scientific concepts, and address your inquiries. As a dedicated science communicator, I strive to spark curiosity and foster a love for science in my audience.

Table of Contents

Introduction

Cytodifferentiation is defined as the development of specialized cells from unspecialized ones through the stimulation of specific nutritional and hormonal factors. In tissue culture, the differentiation of callus or single cells in suspension culture is the result of dedifferentiation of a few cells due to the twin phenomenon of cytoquiscence and cyto-senescence.

The well-studied example of cytodifferentiation is the formation of the vascular system composed of the tracheary elements and the phloem elements. They are formed from the meristematic cells of the stem apex, root tip, and vascular cambium.

This article will cover the basics of cytodifferentiation and the factors that affect the phenomenon in plant cells.

Cytodifferentiation in Tracheary Elements

Cytodifferentiation of tracheary elements is an irreversible process of cell specialization. In this case, hollow dead cells form the tracheary elements due to the loss of nuclei and cell contents at maturity. In the final differentiation stage of tracheary elements, the significant wall thickening is accompanied by specific biochemical changes that can be used as marker proteins for studying the early stages of this event.

Tissue culture has been an amazing tool to study the mechanism of cytodifferentiation and cellular changes during the phenomenon through in vitro studies on xylogenesis, xylem-like cell differentiation, and phloem differentiation. All these studies involve biosynthesis and modification of the cell wall and cell contents.

The formation of tracheary elements can be induced under in vitro conditions using several cell types including phloem parenchyma, cortex in roots, pith parenchyma in shoots, tuber parenchyma, and the mesophyll and epidermis in leaves.

Under in-vitro conditions, its differentiation and development pass through four stages that include:

Acquiring competence by the target cell to initiate cytodifferentiation.
Cell enlargement
Secondary wall formation
Lignification and autolysis of cell contents including the nucleus and selective dissolution of the wall.

Tissue Culture as a Tool to Study Cytodifferentiation

Tissue culture or cell culture is an excellent tool to study cytodifferentiation. It’s because:

Only a simple variation in the media composition, as changes in plant hormones, can regulate in vitro differentiation of tracheary elements.
Through tissue culture, homogenous cells and tissues can be easily obtained to initiate cultures and scale up to produce large quantities of biomass for biochemical studies.
The tissue culture system is the best fit for learning intracellular signal transduction pathways causing changes in gene expression.

Several cell culture systems have been developed to study the changes that occur during vascular differentiation. For example, Zinnia elegans cell cultures have been used to study the sequence of events leading to tracheary element differentiation.

Similarly, French beans have been useful to study the biochemical changes associated with secondary wall synthesis in cells, which develop to resemble fiber cells.

Factors affecting in vitro vascular tissue differentiation

Growth regulators

Auxin and Cytokinin:

Exogenous auxin was believed to induce trachea element induction in the majority of species. But, the optimum concentration of the PGR varies from species to species. Furthermore, the endogenous level of auxin can affect the process of differentiation.

For example, tracheary element differentiation was observed in habituated and crown gall cells without the addition of any amount of exogenous auxin.

In some cultures of plant species, like cultures of carrot and tobacco, the tracheary element differentiation is induced by using the combination of exogenous auxin and cytokinin hormones. However, in some species, like Zinnia, it is also observed that the cytokinin is required only during a brief period of initiating differentiation stage, whereas, auxin was required until the last stage of differentiation of tracheary elements.

In some other species, like Helianthus, cytokinin was required for a longer period (for two days), compared to auxin (for only one day). These hormones are involved in the activation of some molecular pathways for the initiation of differentiation or are considered to be the expression of some specific genes.

Gibberellic acid, Abscisic acid, and Ethylene:

It has been observed that in some plants gibberellin induced tracheary elements differentiation with auxin and cytokinin, in some it inhibited the repose, and in some others, it delayed the auxin cytokinin induced tracheary element differentiation response.

Scientists conclude that the endogenous level of gibberellins might be responsible for the variant differentiation responses of gibberellin in different plants. And, it appears that gibberellic acid does not directly function in the in vitro induction of tracheary elements.

Other factors

Sucrose:

In Phaseolus vulgaris, different differentiation responses in plants were recorded in presence of sucrose, in combination with auxins. For example,

1% sucrose + low amount of auxin: Only a few xylem elements were formed.
2% sucrose + low amount of auxin: Better differentiation of xylem with little or no formation of phloem.
2.5–3 % sucrose: Both, xylem and phloem differentiated.
4% sucrose: Only the phloem was formed

Calcium:

In Zinnia, it was observed that removal of calcium or addition of calcium blockers resulted in inhibition of tracheary element differentiation when added at the beginning of the culture.

Physical and physiological factors:

Factors like temperature provided to the plant cultures, age of plants, wound stress, light intensity and duration, and mechanism of cell isolation are some other measures that decide the differentiation of tracheary elements in different plant species.

Tissue culture has been an excellent tool for scientists to study plants’ developmental responses and growth mechanisms.

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Happy Culturing!