Explain the process of mitosis in a tissue culture for normal cells.

Mitosis is a process of cell division that results in the production of two cells with identical genetic heritage.

prophase

Prophase (2n-4c) is the first phase of cell division (and the longest, about 90% of dividing time) in mitosis.

1. The centrosomes, consisting of two centrioles (nine triplets of microtubules), each migrate to a pole of the cell.

The microtubule cytoskeleton forms the mitotic spindle (division spindle) that connects the two centrosomes.

2. The chromatin condenses enchromosome, that is to say, into two sister chromatids produced by the replication of the starting chromatid and which contain identical genetic information.

3. The nuclear membrane disintegrates by phosphorylation of the lamins of the nuclear lamina.

4. Kinetochores are formed at the centromere level.

This third part of prophase is called prometaphase and can be defined as an independent phase by some authors.

metaphase

The metaphase (2n-4c) is the phase where the chromosomes are placed on both sides of the equatorial plane to form the equatorial plate.

During this phase especially, but also already in prophase, the chromatin is especially present in compacted form (heterochromatin), resistant form especially around the centromeres: this makes it possible to increase the resistance of the chromosomes to avoid their rupture during their separation during the anaphase.

On the other hand, some regions are poorly compacted (euchromatin).

These gene promoter regions were active in the cell prior to entry into mitosis.

This process called "bookmarking" (gene or mitotic bookmarking, literally: mark page), is an epigenetic mechanism that allows to transmit to the cells cells the "memory" of the active genes before the entry into mitosis. Let's not forget that transcription stops during mitosis.

anaphase

Anaphase (2n-4c) is the phase where sister chromatids separate and migrate to opposite poles of the cell.

It is the sister chromatids that are separated, but not the chromatid pairs as in meiosis.

telophase

Telophase (2n-1c) is the phase in which the nuclear and cellular envelopes appear, leading to the appearance of two cells with 2n chromosomes and each with a chromatide.

During this period, or sometimes, according to the authors, in a last period, called cytokinesis or cytodiérèse, appears a division in a plane perpendicular to the axis of the mitotic spindle which separates the cell in two. This contractile ring is formed of actin and myosin.

All other organelles reform as nucleoli and chromatids decondensate to reform chromatin.

Regulation of the cell cycle

To ensure, on the one hand, the immutable order of the succession of the four phases of the cycle (regulation of the cycle), and on the other hand, the obtaining of two exactly identical daughter cells (DNA monitoring), the cell has highly sophisticated control systems. In the first case, (cycle regulation), it is mainly cyclin-dependent kinases, the Cdk, which intervene. In the second case, other molecules intervene in different mechanisms of cycle monitoring to inhibit the Cdk of the cycle regulation and stop the cycle, if the previous step is not completed, or if a "repair" is necessary .

* The regulation of the succession of the four phases of the cell cycle

The different phases of the cycle take place according to the immutable order mentioned above and it is to ensure the maintenance of this sequence that Cdks that regulate the cell cycle intervene. There are several; they intervene throughout the cycle in a specific order: in phase G1 and for the transition G1-S, that is to say for the triggering of the replication of the DNA, in phase S for the continuation of the replication , in the G2 phase and for the G2-M transition, that is to say for the triggering of the mitosis and for the execution of the mitosis. Cdk act either on the proteins that allow the realization of the events of the cycle (their function is then to cause the events of the cycle), or on the protein Rb, (their function then being to allow the progression of the cycle).

The normal succession of the different phases can take place only if the different Cdk intervening during the different phases are present and active at the opportune moments.

* Cycle monitoring mechanisms.

The monitoring mechanisms are in addition to the regulation of the succession of the four phases of the cycle by the Cdk. They allow the monitoring of fundamental aspects such as the state of the DNA molecules before, during and after their replication (DDCP = DNA Damage Checkpoint), the total completion of the replication before the entry into mitosis (RCP = Replication Checkpoint ) and the correct positioning of all the chromosomes on the plate.

this protocol describes a detailed procedure for intracellular flow cytometry staining.

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