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Checkpoints in Cell Cycle

Introduction

Cell cycle checkpoints are specific genetic or biochemical signals that monitor different phases of cell division and arrest progression through the cell cycle if a problem is detected.

What is the Cell Cycle?

The cell cycle (also known as the mitotic or division cycle) describes the series of events in a cell leading to its division. It is divided into four phases- gap 1 (G1), DNA synthesis (S phase), gap 2 (G2), and Mitosis (M stage) phase.

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What are the 4 Stages of the Cell Cycle?

The four stages of the cell cycle include;

1. Gap 1 (G1) phase of cell cycle

An image comparing Eukaryotic and Prokaryotic cells

The G 1 phase of the cell cycle is the preparation phase. During this period, eukaryotic cells grow in size, synthesize DNA and replicate organelles.

2. DNA synthesis (S phase)

DNA phase of the cell cycle is the so-called S phase. During this stage, DNA is synthesized, chromosomes are replicated, and a process known as chromatin condensation occurs during which the DNA wraps around histone proteins forming nucleosome structures.

During replication of DNA, the cell checks for any damage to ensure that only undamaged copies go onto daughter cells. If there is any damage in DNA, the cell repairs and replicates it before entering mitosis.

If there is no defect, the cell enters into the cell cycle’s gap 2 (G2) phase.

3. Gap 2(G2) phase of cell cycle

In G 2 phase, several activities occur, including protein synthesis, replication of organelles. The cell also grows in size until it reaches a critical mass, which triggers the transition into the mitosis (M) phase. During this stage, cells undergo further growth and division of cytoplasmic organelles.

If there is any damage in DNA, the cell repairs and replicates it before entering mitosis.

If there is no defect, the cell enters into the M phase of the cell cycle.

4. The Mitosis (M) phase of the cell cycle  

Mitosis- Photo credit- istockphoto

Mitosis is also called division or M-phase compared to G1 and G2 phases, which are identified as a preparation phase for a new round of mitotic divisions. Mitosis is a cell division phase in which a parent cell divides into two daughter cells genetically identical to each other.

As the mitotic of the cell cycle, the M phase is responsible for the segregation of chromosomes. During this time, chromosomes condense and attach to microtubules. The cell then splits into two new daughter cells by a process called cytokinesis.

The cell cycle is controlled by a series of checkpoints that monitor the status of important events in each phase. If an abnormality is detected, the cell cycle process must be stopped until appropriate repairs are made to ensure genetic integrity and stable growth. If the problem cannot be resolved, the organism dies. The checkpoints keep the cell’s DNA from becoming damaged and help repair the damage that may have already occurred.

DNA Synthesis (S phase) Phase of Cell Cycle

The S phase of the cell cycle is the time when DNA synthesis takes place. During this phase, cells double in size, and each chromosome is duplicated into two identical sister chromatids that are physically attached. One set of chromatids remains with the original cell, and the other set goes into each daughter cell.

The Transition from G1 phase to S phase

The G 1 phase of the cell cycle ends with the replication of some DNA and the period between these intermediate points to the end of the S phase is an important checkpoint in the cell cycle. The transition from G1 to S phase occurs via two-step mechanisms.

1. Cell cycle checkpoints

Checkpoints are genetic or biochemical signals that monitor different phases of cell division and help repair errors that may occur. There are five cell cycle checkpoints, which refer to specific stages of the process.

At the G1/S checkpoint, cells will pass on to enter the S phase or be sent back into the G phase if they do not have all their necessary components. Any abnormality discovered at this checkpoint is usually repaired before entry into the next phase.

Checkpoints in G2 (Gap 2) and M phase also exist, but defects discovered at those points are more difficult to repair. Defects may go undetected by these checkpoints and allow cells to progress into mitosis with incomplete or faulty DNA. If undiscovered, this can lead to a defective division of cells and the formation of tumors.

2. Checkpoint in DNA replication

The cell’s ability to repair itself is directly related to its level of DNA synthesis. The checkpoints are often observed at the time when double-stranded DNA is synthesized and the final phase. In this stage, if there is any defect in the genome, cells repair their DNA.

In the first phase of the cell cycle (G1), DNA synthesis is a check point. In other phases, during progression through the G2 phase or S phase, if an abnormality is detected, cells are copied and sent to repair centers for treatment before dividing into two daughter cells. Most defects discovered at these checkpoints can be corrected before the new cells divide.

The checkpoints that are incompletely understood play a role in cell death or permanent cell arrest, which occurs when damage is not repaired or detected at the right time.

Results of defective checkpoints can lead to apoptosis or so-called programmed cell death, which refers to the process whereby normal cellular activities are reined in when no longer needed. In this process, the cell uses its ability to detect defects and die by committing suicide or self-destructing for the organism to renew itself.

Cell cycle checkpoints are important because they protect against cancer cells that would arise if DNA synthesis were not monitored closely enough. Defects discovered at these checkpoints may be repaired before cells divide, and if not, they can lead to cell death. This helps protect against cancer that could arise from undiscovered genetic disorders.

How Does Checkpoints Work?

Checkpoints exist to ensure the accurate completion of such processes like DNA replication, chromosome segregation by mitosis, and cytokinesis. These processes are monitored by different biochemical signals that activate or inhibit key cell cycle regulatory proteins if activated. For example, suppose the DNA synthesis inhibitor, p21 has been activated by replication stress. In that case, it will inhibit CDK2, which inhibits cyclin-dependent kinases and prevents cell cycle progression until the appropriate repair of the damage occurs.

Checkpoints not only control when events occur but also how they occur. The process is controlled at three levels – initiation, elongation and termination. If an event is not initiated properly, then it may not occur at all, or it may be halted before completion. Checkpoints also ensure that events do not proceed with unsynchronized rapidity where they are supposed to happen at specific times.

Checkpoint controls in eukaryotes depend on the cyclin-dependent kinases (CDKs) that are present in the cell. Cyclins ensure that the CDKs are activated or deactivated at specific times during the cell cycle and become attached to them.

The phosphorylation of a protein, p34cdc2, by one of these cyclin-dependent kinases ensures that DNA replication can occur. Some of the checkpoint controls in bacteria include termination factors and endonucleases that stop DNA replication after a specific period or when the cell has reached a certain size.

These checkpoints ensure that each step in the cell cycle is completed successfully to balance the new growth and stability of the genome. If these cell-cycle events do not occur properly, the cell cycle may be prematurely halted, or inappropriate events may occur, preventing the cell cycle. The normal image and a DNA damaged image is represented below:

DNA damage during replication prevents cells from progressing through the apoptotic phase of the cell cycle and leads to DNA repair mechanisms which result in loss of genetic information more often than not. DNA repair mechanisms are complex and have evolved to keep the genome stable during reproduction, cell division and embryogenesis.

Checkpoint controls also act as a safeguard against abnormal, uncontrolled growth that may pose a threat to the body in certain circumstances such as cancer and lead to the transformation of normal cells into malignant tumors.

In cancerous cells, the normal signals that control cell division have been disrupted, and the asymmetry between the mother and daughter cells has been lost. The result is uncontrolled growth where the daughter cells continue to divide in an unregulated fashion resulting in cellular growth that can invade other tissues within the body through local invasion or metastasis.

DNA damage can be due to a variety of sources. UV radiations, chemical radiations, and certain viruses have been associated with DNA damage in humans.

To combat this problem, cells have complex mechanisms that prevent the degradation of genetic information and genomic instability. The following events need to occur for the cell cycle process to continue after the DNA damage has occurred and be repaired:

Events That Need To Occur For the Cell Cycle Process to Continue After the DNA Damage

The mechanism to ensure that these events occur is a complex process and involves different classes of proteins.

INITIATION OF REPLICATION OR CHECKPOINT CONTROLS – The first step in the cell cycle after mitosis is to ensure that replication occurs properly by ensuring that the integrity of the DNA is maintained. This happens without any errors or damage to the genetic information that would cause mutation and changes in the cell’s phenotype.

It is important to note that these checkpoints begin with a “cell-cycle dependent kinase inhibitor”, which prevents uncontrolled replication, especially when there is DNA damage. These inhibitors ensure that certain cyclin-dependent kinases are not activated until DNA replication begins. This ensures that the genetic material is protected and no mistakes occur during DNA synthesis.

PROCESSING OF INITIATION FACTORS – Another set of checkpoint controls ensures that any enzyme proteins required to create a replication fork do not begin their reaction too early or too late to prevent errors in the genome.

This ensures that the replication fork is created at the correct time and point during DNA synthesis. In certain eukaryotic cells, this process occurs as there are specific proteins released by the endonucleases to prevent premature replication of genetic information.

ONSET OF DNA SYNTHESIS – The third group of checkpoint controls ensure that replication begins at the correct time and not too late or too early to prevent the erroneous addition of nucleotides. This ensures that there are no errors in DNA synthesis and replicated sequences start with a clean slate and do not have any extra or missing genetic information from the previous cell cycle.

Checkpoint controls also ensure that DNA synthesis occurs for the duration of time required to create two identical copies of genetic information and finishes when it should. These processes ensure that each daughter cell has the same amount of genetic information as to its mother cell.

Any mistake made during the process by checkpoint controls would lead to an unequal distribution of nucleotides in chromosomes and result in an abnormal phenotype. This checkpoint controls to ensure that DNA synthesis does not continue even if there are mistakes in one of the copies. This would lead to genomic instability in the cell and possibly other abnormalities within the body.

In certain forms of DNA damage (UV light), replication needs to occur as normal because any errors made during replication leads to a decrease in genetic information and leads to a phenotype of genomic instability (e.g. skin cancer).

However, suppose DNA damage is due to certain types of radiation or chemicals. In that case, it is important that replication does not continue as any errors made during the process will lead to a cell phenotype with mistakes or mutations that would result in an abnormal phenotypic cell and possibly result in the organism’s death.

The regulation of DNA replication is a complex process involving many different proteins and cellular controls.

There are three major classes of proteins that must interact with one another to ensure the correct distribution of genetic information during the S phase, leading to viable offspring.

These classes include replication activating complexes (including origin recognition complexes), regulatory proteins, and chromosome binding proteins. The three classes of protein work together to ensure correct distribution and replication timing during the S phase.

What Happens at Each Cell Cycle Checkpoint?

Cell cycle checkpoint 1- G1 cell cycle checkpoint

In eukaryotes, cell division is monitored by a cell cycle checkpoint called the G 1 cell cycle. It takes place in the S phase; the middle stage between DNA synthesis and cell membrane reformation. Here, cancer cells bypass cell cycle checkpoints and continue cell division despite cell damage.

Cell cycle checkpoint 2 – G2 cell cycle checkpoint

After DNA synthesis (S phase), cell divide is monitored by a cell cycle checkpoint called the G2. It takes place at the end of the S phase; before cell membrane reformation. Proteins such as p53, Rb proteins and cell-cycle kinases play key roles in cell cycle checkpoints. After cell division, the cycle is checkpointed at the G0 cell cycle. This ensures that daughter cells are ready for cell division.

Cell Division –Photo credit istockphotos

Cell cycle checkpoints and regulation of cell size

Potential cancer therapeutics target cell cycle pathways. This may delay cell cycle progression leading to delay of cell division.

Cell cycle checkpoints and cell senescence

Checkpoints are cell cycle-specific. Senescent cells bypass cell cycle checkpoints to continue cell division. Cell senescence is due to premature aging caused by the shortening of telomeres, oxidation, inflammation, and oncogene expression in the checkpoints.

Cell cycle checkpoints and cell death 

There is no cell division in cell death. Cell death occurs by cell suicide (apoptosis) or cell execution (necrosis).

Cell cycle checkpoints and repair of damaged DNA

Erroneous cell division can cause daughter cells deficient in organelles, cell organelles or cell membrane.

Cell cycle checkpoints and cell differentiation

These can be distinguished from each other based on cell factors. These factors may include organelle structure, function (e.g., mitochondria) and cell size. Cell differentiation is cell cycle-specific, and cell division occurs in cell-specialized cell types.

Cell cycle checkpoints and cell migration or cell locomotion  

The cells can move from one region of the organism to another (e.g., white blood cells moving from the bloodstream into infected tissue). There is no division of cells in cell locomotion.

Cell cycle checkpoints and cell growth –The growth of cells occur by cell multiplication rather than cell division.

Cell cycle checkpoints and apoptosis – Apoptosis is cell suicide (cell execution).

 In apoptosis, cells shrink (pyknosis), condense nuclear chromatin, fragment cell organelles (apoptotic bodies), degrade cell membrane and secrete cell contents. Apoptosis is cell division-independent cell death.

What are the steps of DNA synthesis?

The steps of DNA synthesis are:

1. Occurs before cell division (in G0 phase) – DNA synthesis begins in genes within the nucleus of cells. Genes are complex structures consisting of repeating DNA sequences called exons and intervening, non-repeating DNA sequences called introns.

Splicing is the extracellular digestion of introns from messenger RNA (mRNA) to form messenger RNA (mRNA). These are molecules active in protein synthesis within the cytoplasm of cells. Cells then assemble ribosomes, which are organelles for protein synthesis in the cytoplasm of cells. After DNA synthesis begins, cell growth occurs, and cell mass increases due to replication or duplication of genetic material in the nucleus.

2. Occurs during cell division (in G1 phase) – DNA synthesis is completed within genes after DNA replication occurs and genetic material increases within the nucleus of cells. At this point in the cell cycle, DNA damage-checkpoint proteins such as p53 and Rb proteins elicit transcription of genes of repair enzymes such as DNA polymerases and proteases that remove damaged segments of DNA. During this process, cell growth ceases until DNA damage-checkpoint proteins such as p53 and Rb proteins are satisfied that cellular DNA is properly repaired.

3. Occurs during cell division (in M or mitotic phase) – If repair enzymes cannot repair cells, DNA damage-checkpoint proteins such as p53 and Rb proteins quench or impede synthesis of mRNA transcripts of genes of repair enzymes.

This results in cell division arrest (either G1 arrest or M/G2 phase cell cycle arrest), leading to apoptosis depending on the extent and persistence of DNA damage during the cell cycle. DNA damage checkpoints and cell death (cell suicide) is called apoptosis. Apoptosis occurs in the G1, G2 phase or M phase of the cell cycle.

Cell cycle checkpoints and repair of damaged DNA – Erroneous cell division can cause daughter cells to be deficient in organelles, organelles, or membranes.

Checkpoint proteins such as p53 and Rb proteins can detect DNA damage in genes of repair enzymes such as DNA polymerases. This then triggers the assembly of new repair enzymes to replace damaged DNA polymerases within the nucleus of cells during cell division (in the G1 phase).

If defective gene products or replication errors occur, checkpoint proteins block cell cycle progression until the integrity of cellular DNA is restored.

4. Occurs after cell division (in the G2 phase). The final step of DNA synthesis occurs during the S phase of the cell cycle when genetic material increases within the nucleus of cells by replicating or duplicating genetic material.

A strand of cellular DNA called the lagging strand synthesized by DNA polymerases is lost in this step. This loss of genetic material is repaired by enzymes called transposases, which can duplicate the sequence lost from the lagging strand directly onto an identical portion of the leading strand.

Transposase activity occurs at a very slow rate and facilitates rapid cell division in processes such as cytokinesis that occur in G2.

Cell cycle checkpoints and repair of damaged DNA – If transposase activity occurs too slowly to replace lost genetic material during the S phase, cell division arrest occurs until transposases are available in enough quantity to repair the loss of genetic material lagging strand of cellular DNA.

How does Cancer Affect the Cell Cycle?

Cancer affects the cell cycle by preventing cell cycle checkpoints from performing cell-repair functions. Cancer cells acquire the ability to avoid cell death (or apoptosis) by cell cycle checkpoints and block cell suicide (or cell death).

This causes cancer because cancer cell is able to replicate without being detected or removed by cell cycle checkpoints that occur in G1, G2 or M phase of cell division. The cell cycle checkpoints are unable to repair defective cell products that occur during cell division, which results in cancer formation and progression.

Cell cycle checkpoints block cell suicide or cell death – If a cancer cell acquires a mutant p53 protein, the activity of p53 is impaired, and the cell cycle checkpoint becomes defective in detecting DNA damage.

The cell cycle checkpoint is unable to repair defective cell products that occur during cell division. As a result, the cell replicates and acquires additional DNA damage without being detected or removed by cell cycle checkpoints during the G1, G2 or M phase of cell division.

Image of Cancer cells- Photo by National Cancer Institute-Unsplash

Cell cycle checkpoints block cell suicide – If cancer cells acquire mutant Rb proteins, cell cycle checkpoints in the G1, G2 or M phase of cell division can no longer detect DNA damage. This results in cell cycle arrest (either G1 arrest or M/G2 phase arrest), leading to cell suicide (apoptosis) depending on the extent and persistence of DNA damage during the cell cycle.

Cell cycle checkpoints block cell suicide – If cell cycle checkpoints become defective, cell replication can occur unchecked without being detected or removed by cell cycle checkpoints. This allows cancer cells to acquire additional DNA damage during the cell cycle. As a result, cell division may be blocked in the G1 phase, and cell suicide (apoptosis) occurs if the cell cannot be repaired with cell cycle checkpoints in G2 or M phase.

Cell cycle checkpoints block cell suicide – If cancer cells acquire mutant p53 proteins, the activity of p53 is impaired, and cell cycle checkpoints are unable to repair defective cell products that occur during cell division. This results in cell replication without being detected or removed by cell cycle checkpoints during the G1, G2 or M phase of cell division.

The cell cycle is a tightly controlled process in which cells make copies of themselves using the genetic code stored within each cell’s nucleus. Each cell does this slightly differently, making sure that our bodies are continually functioning correctly. The cell cycle is split up into four stages: G1 (the gap or growth stage), S (the synthesis stage), G2 (the gap or growth stage) and M phase, also known as mitosis. 

The link between cancer and the cell cycle is when there are over-production or under-production of certain proteins. 

The cell cycle goes through a process called the transition from G1 to S phase, and it’s during this stage, DNA replication occurs in preparation for cell division. This is the point at which cancer can form because if the genetic material isn’t copied correctly, then cancer is likely to develop. 

If there is too much of a protein being produced by the cell in the S phase, it could trigger excessive DNA replication. Alternatively, if not enough of this protein has been made, the DNA can’t be copied correctly, and cancer may result. The trick for mutations to occur lies within proteins that control cell growth. These proteins are called transcription factors. 

What Are the Steps of the Cell Division Process?

The process of cell division takes place in several steps;

– The cell grows and makes more cytoplasm, membrane, organelles, etc.

– It synthesizes DNA for the new cells from its genetic information. This is done by the replication of chromosomes.

– Each chromosome has several identical sister chromatids. A centromere connects the sister chromatids.

– The cell divides into two new cells, which have identical DNA to the parent. Each daughter cell receives one copy of all chromosomes from the parent cell and a complete set of organelles.

– In prokaryotes, after cytokinesis, the cell may remain in the G1 phase with no further progression; however, in eukaryotes, the cell then starts the G0 phase (post-mitotic) and stays there until stimulated to reenter the S phase.

How Many Checkpoints are Present in Cell Cycle?

Three main types of checkpoints have been characterized in experimental systems: (1) DNA replication, (2) chromosome alignment on the metaphase plate, and (3) spindle assembly.

DNA Replication

DNA replication is a highly conserved and precisely regulated process that contributes to genome stability. DNA synthesis is performed in several stages by several proteins complex, which all ensure that the replication occurs only once per cell cycle.

If additional rounds of replication were to occur, as in defective control by DNA damage checkpoint, genomic instability would occur due to damage caused by incompletely repaired replication intermediates.

Chromosome Alignment on the Metaphase Plate

The vertebrate cell cycle is regulated by several proteins that supervise the transition from G2 into mitosis, including the cyclin-dependent kinases (Cdks). At this point, there must be a clear separation of sister chromatids before anaphase, which requires several additional proteins (cohesins and synaptonemal complex).

Spindle Assembly

The correct positioning of chromosomes in the mitotic spindle is critical for proper cell division. Therefore, the assembly of functional spindles is carried out by multiple protein complexes that ensure that correct sister chromatid separation can occur.

Cytokinesis: Is it a Checkpoint?

Cytokinesis is the process by which cytoplasm is divided into daughter cells. The beating of the mitotic spindle and formation of the cleavage furrow are coordinated by actin filaments that grow from the spindle midzone and associated microtubules.

The first step of cytokinesis is the severing of sister kinetochores by a multiprotein complex known as the cohesin ring, composed of the major structural subunit Scc1 and other smaller proteins (Sic1, Smc3, Smc1, and Smc4). After sister chromatid separation, the cleavage furrow forms and the cell plate is assembled.

Cytokinesis is a highly regulated process that requires proteins for signaling (such as MAPK cascades), contractile actin filaments, and many other proteins involved in the septation of the plasma membrane into daughter cells such as proteases and chaperones.

However, cytokinesis may not be considered a DNA damage checkpoint because it is not due to an error in DNA replication but rather by defects in the septation machinery that can occur when other factors are impaired (for example, defective assembly of cell plate can lead to late anaphase bridges in oocytes).

Mitotic Checkpoints

DNA double-strand binds to a kinetochore. Then it opens the DNA and orientates the sister chromatids to each other. The two centromeres then join by covalent bonds (known as cohesins), forming a ring structure called a synaptonemal complex (SCC).

Mitotic Spindle Assembly Checkpoint

Spindles are assembled by the three major protein complexes: (a) a centriole-dependent outer kinetochore, which is responsible for attaching to microtubules and linking them with each other; (b) an inner spindle pole body that forms during mitosis from two proteins called Aurora B and Cdk13/cyclin; and (c) the mitotic checkpoint complex, consisting of several families of proteins that regulate spindle assembly.

A recently discovered protein called Bif-like is one such component: it associates with microtubules at kinetochores to ensure they are correctly aligned by binding to both sister chromatids simultaneously.

In addition, the spindle assembly checkpoint arrests the cell cycle at metaphase by regulating cyclin B/Cdk1 activity. This regulated proteolysis of Cdk1 prevents the massive accumulation of Aurora B (Aurora-B kinase), which activates several checkpoints dependent on the passage through mitosis.

To maintain correct chromosome segregation during cell division, the spindle assembly checkpoint must ensure that proteins (such as securin) involved in chromatin condensation are not degraded before anaphase. In addition, Kss1 prevents degradation of Cdc20 and Mad2, which allows the cyclin B-Cdk1 complex to accumulate until mid-metaphase.

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Chromosome Segregation Checkpoint:

To ensure that sister chromatids are separated properly, the mitotic spindle and centrosome proteins monitor the quality of chromosome attachment to microtubules. Proper kinetochore attachment is a prerequisite for accurate chromosome segregation in anaphase. To accomplish this, two main checkpoints ensure proper kinetochore attachment.

Coupling of the Anaphase Checkpoint to DNA Damage Signaling

Chromosomal passenger complex:

The mitotic spindle is a dynamic structure that contains three main components: microtubules, spindle pole bodies and chromosomes. The microtubule component undergoes dramatic rearrangements to segregate chromosomes into daughter cells.

The main protein kinase involved in the mitotic progression is Cdk1-cyclin B. This complex phosphorylates specific substrates (such as those involved in spindle assembly) and promotes their degradation via the 26S proteasome. However, because of its need to phosphorylate many proteins, which occurs in G1, it is not active at the start of mitosis.

Once Cdk1-cyclin B accumulates on chromatin, its activity can be regulated by phosphatase inhibitors (such as Clb2 and Aurora A). The phosphorylated substrates thus marked for destruction include proteins necessary for centrosome separation, microtubule severing, and the transition of chromosomes into anaphase (such as securin and Cdc20).

Molecular View of Some Mitosis-Related Diseases

Cytologic studies showed cell division inhibition. According to immunohistochemistry results, it was found out that villous cytotrophoblasts do not exhibit their characteristic inclusions (nuclei) within cytotrophoblast cells.

Immunohistochemistry and Western blot proved that micro-RNAs, let-7a, miR21 and miR29b are increased under hypoxic conditions.

Aneuploidy

One of the most common chromosomal abnormalities in humans is aneuploidy, which involves changing the number of chromosomes possessed by one or more cells. Aneuploidy results from errors in chromosome segregation at either meiotic prophase I (MI), or mitotic anaphase (MII). In contrast to MI, which is associated with ~10% of human conceptions and results in early pregnancy loss, aneuploidy at MII occurs in 30-50% of conceptions. Still, prenatal mortality is relatively high because most embryos are lost during the first few weeks following fertilization. This suggests that not all aneuploid embryos develop into viable fetuses.

Clinical Significance of Chromosomal Instability (CIN)

Chromosome instability (CIN) results in aneuploidy, which plays a major role in several human tumors, including most types of leukemia and solid tumors. Aneuploidy is not the only genetic change leading to the genesis of a tumor. Abnormalities may occur in the chromosomes and/or in chromosome architecture, such as deletions and translocations. Aneuploidy usually results from abnormal mitosis or meiosis.

What are the Points during the Cell Division Process at which Checkpoints can Arrest the Cell?

There are three major points during the cell division process at which checkpoints can arrest the cell.

1) G2/M Checkpoint: It is found during chromosome replication and separation of chromosomes in early mitosis. 

A cell cycle checkpoint ensures that a cell has enough proteins to complete mitosis while still completing proper DNA replication. This regression point occurs after the completion of the S phase and during the beginning of mitosis. For a cell to divide, it needs to carefully organize its DNA and ensure that all cellular machinery is present and functioning properly at the point when division occurs.

During G2/M checkpoints, Rb proteins follow cell cycle-specific modifications of histones on DNA by making sure each chromosome has been completely replicated and located on the correct side of the cell. Rb proteins also monitor the presence of microtubule-associated protein 1A/1B light chain 3 (LC3) complexes, which are used to transport cellular structures around the cell in preparation for division.

If any problems are detected during G2/M checkpoints, the cell undergoes mitotic arrest, after which it enters a form of cell cycle called anaphase lag.

2) DNA Replication Checkpoint: The replication checkpoint is involved in monitoring the progression of the S phase. It ensures that all DNA has been replicated and passed down to each daughter cell. If any problems are detected during the S phase, the cell enters a period of arrest called a DNA damage checkpoint.

3) Mitotic Checkpoints: These checkpoints are involved in the execution of mitosis, ensuring that each chromosome has been properly attached to spindle fibers before cell division occurs. A failure during any of these checkpoints can lead to anaphase lag or cytokinesis failure.

Mithorace checkpoint is a G2/M-phase cell cycle checkpoint that controls the transition from the G2 to M phase of the cell cycle in response to mitotic spindle damage.

G0 Checkpoint – It is present at the beginning of the G1 phase and monitors DNA integrity, cell size, etc. It ensures that a cell has enough time to complete DNA synthesis to enter the S phase.

G1 Checkpoint – It is present during the early growth phase of the cell cycle and ensures that all proper organelles are present and functioning and ensure no problems with DNA structure or number have occurred. 

S Checkpoint – The DNA replication checkpoint is present during DNA synthesis and ensures that the cell has undergone proper DNA replication before proceeding into mitosis.

M Checkpoint – The metaphase checkpoint is present at the beginning of mitosis and monitors the attachment of chromosomes to spindle fibers, ensuring cells have enough proteins for division. 

Anaphase Checkpoints: These checkpoints are encountered during cell division and ensure that no problems have occurred with the attachment of chromosomes to spindle fibers, such as faulty attachment or incorrect chromosomal structure. 

In eukaryotes, cell cycle regulators consist of various cyclins, CDKs (cyclin-dependent kinases), CCNEs/CCNYs (checkpoint proteins) and other regulatory molecules. 

Cyclins are important for cell cycle progression because they bind to and activate CDKs. CDKs phosphorylate the C-terminal tails of cyclins, activating them as well as other downstream targets.

Cyclin levels are tightly controlled by a family of inhibitors called ‘CDK-inhibitors’. After the binding of CDK to cyclins, the cell cycle progresses to the next stage.

In normal cells, CCNE1/CCNY (checkpoint proteins) is present during the M phase and mitotic checkpoints to arrest cells that cannot attach their chromosomes properly.

After arresting cells, CCNE1/CCNY triggers DNA damage repair or apoptosis to prevent the cell cycle from continuing until the proper attachment is achieved. Mithoracin is a compound that controls the expression of CCNE1/CCNY, inducing mitotic checkpoint defects and therefore bypassing apoptosis or DNA damage repair. 

Metaphase to Anaphase Transition in Cell Cycle

Metaphase to anaphase transition is a cell cycle stage where chromosomes are attached to spindle fibers.

During this transition, the chromosome-spindle attachment must be carefully regulated because of the very short time available for proper attachment. If any problems arise with chromosomal stability or spindles morphology, it will cause mitotic collapse. 

This cell cycle process is controlled by certain proteins, such as MAD1, MAD2, BUB1 and INCENP. MAD1/MAD2 members of the MPS4 protein family regulate spindle-kinetochore attachment during metaphase to anaphase transition.

In normal cells, MAD1 and MAD2 are expressed on kinetochores, and after cell division, they are degraded by the anaphase-promoting complex (APC). BUB1 is required for the regulation of MAD1/MAD2 degradation. INCENP regulates APC activity and prevents it from degrading MAD proteins.

A failure in metaphase to anaphase transition may lead to mitotic catastrophe, premature chromosome separation and consequently cell death. 

In humans, a mutation in the CLASP2 gene is implicated in a syndrome known as Clasp-A (Cancer-like Aniridia Spindleopathy).

Patients with this disorder have been shown to suffer from mitotic failure and have abnormal ocular development. In any cell cycle disruption, the unrepaired failed replication of chromosomes may result in aneuploidy and genetic instability, which then causes different kinds of cancer.

What are Genes?

Genes are segments of DNA that contain the instructions for building proteins in our bodies. There are many different types of proteins, each with its unique function. Proteins perform all of the tasks which keep us alive.

Genes form the blueprint from which we develop into an individual. Humans share 99% of their genetic sequence with chimpanzees, and chimpanzees are more closely related to humans than mice.

The vast majority of our DNA does not code for proteins or other vital substances, however. Instead, it plays a role in gene control mechanisms such as replication, transcription and translation. The reason why our complete set of genetic information is broken up into individual genes remains unknown. 

The human genome contains 3.2 billion base pairs, which is about one letter for every cell in the body. Each base pair is made up of a DNA molecule bonded to another; adenine (A) bonds with thymine (T), and cytosine (C) bonds with guanine (G). A double helix shape forms when A bonds with T and C binds with G. The order in which these base pairs bond together is what makes up our genetic code.

What are Gene Control Mechanisms?

Gene control mechanisms regulate the transcription of a gene to produce mRNA, which will then be translated into a protein molecule by the ribosome. Gene control mechanisms fall into two categories: negative and positive gene control. 

The first type of gene control mechanism is called negative gene control, as it prevents the production of mRNA in a cell. This method of gene regulation occurs when RNA polymerase (an enzyme responsible for regulating the transcription of genetic information) cannot access a DNA molecule due to a blockage of the DNA molecule’s 3-dimensional structure, making it unable to be transcribed.

This is how introns function: they are sections of genetic information that form introns between exons. Introns were long considered useless DNA until it was realized in 2001 that they play a role in the development and cell differentiation. 

Introns are removed from the final version of the mRNA, which is then used to produce functional proteins. This process goes unnoticed by many, as not only does it take place in a single cell; but because transcription and translation are both extremely fast processes within a human body, small mistakes such as this don’t matter too much (the cell can just correct the error itself and fix it later). 

The second type of gene control mechanism is called positive gene control, which prevents the production of mRNA. This method is used in eukaryotes (organisms with cells containing a nucleus) and occurs when RNA polymerase cannot diffuse far enough along a DNA molecule to reach the promoter sequence — the section of DNA responsible for mRNA production. This makes transcription in the area impossible, preventing mRNAs from being produced.

Tumor Suppressor Genes

Tumor suppressor genes are a type of genetics that function to prevent the uncontrolled growth of cells. Tumor suppressors act as “off” switches for other genes and stop a cell from producing certain proteins. They have been referred to as guardian angels, ensuring healthy function in our body by preventing damaged genes and cells from replicating themselves. 

Tumor suppressor genes that have been linked to breast cancer include BRCA1 and 2, p53 and the TP53 gene family (which also includes MDM2). The TP53 gene family is essential in regulating cell growth.

So far, three-quarters of all breast cancers are associated with either BRCA11 or BRCA2 (two of the most common tumor suppressor genes) because when these genes are mutated, the cell has an increased chance of either becoming cancerous or speeding up its growth cycle.

Tumor suppressors also have a use outside of breast cancer, as they have a hand in other types of cancers such as prostate cancer.

BRCA1 and BRCA2 have been linked to breast cancer since 1994, which is when a genetic link between the gene for these proteins and a high risk of developing breast and ovarian cancers was first discovered.

BRCA1 encodes a protein that helps repair damaged DNA, while BRCRA2 adds another layer of protection by preventing the spread of mutated cancer cells. Both are tumor suppressor genes. 

Not all people with BRCA1 or two gene mutations have breast and ovarian cancers, although they have a higher chance of contracting either condition than most people. Roughly 5-10% of women who get breast cancer can attribute their disease to a mutation in one of these genes.

Two specific mutations are currently known to be associated with an increased risk for breast cancer; 185delAG and 5382insC. The first of these, the 185delAG mutation, occurs when part (or all) of the gene has been deleted from a cell’s genome.

This can happen naturally, but it is more commonly due to a mistake during DNA replication. It can occur in both DNA strands (doubles). This mutation causes the gene encoding for p53 to stop working properly.

The second, 5382insC, occurs when part of a gene has been copied in reverse. Again, this could be due to a mistake in DNA replication, but it can also be inherited from a parent. The 5382insC mutation causes the function of p53 to be impaired. 

The exact mechanisms by which these two mutations lead to cancer are not yet understood. However, they both likely have the same effect; when either gene has been mutated not work properly, the risk of developing breast cancer increases to around 70-80%.

Difference between the two mutations is that 185delAG usually stops both copies (double mutation) of p53 from working; 5382insC tends to be inherited and only affects one copy (single mutation) of p53. 

The activation of p53 is closely related to the amount of time that it is active for. The longer a cell spends in G1 phase, the more likely it is to develop an additional copy of this gene (due to cell replication).

If the activation occurred early enough, then there would be two copies for each chromosome. This could potentially lead to cancer. In that case, the cell would repair damaged DNA up until it went into the S phase. If no further copies of p53 available, then cancer could develop when the cell started dividing.

It is important to note that many people with neither 185delAG nor 5382insC mutations will still develop breast and ovarian cancers. These mutations are simply two out of hundreds, perhaps thousands, of genes that can lead to cancer.

If it were not for their known link to breast and ovarian cancers, it’s unlikely that they would have been studied at all. However, some of these other genes will likely be linked to cancer in the future. 

Conclusion     

Gene mutations are what drives cancer in things like humans and chimpanzees. Mutations can occur from mistakes made during DNA replication or due to ionizing radiation. It is important for a cell to have a system that can identify mutated genes and dispose of them before they lead to cancer.

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