Micronucleus

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Micronuclei visible in boxes Micro 2.png
Micronuclei visible in boxes

A micronucleus is a small nucleus that forms whenever a chromosome or a fragment of a chromosome is not incorporated into one of the daughter nuclei during cell division. It usually is a sign of genotoxic events and chromosomal instability. Micronuclei are commonly seen in cancerous cells and may indicate genomic damage events that can increase the risk of developmental or degenerative diseases. [1]

Contents

Micronuclei form during anaphase from lagging acentric chromosomes or chromatid fragments caused by incorrectly repaired or unrepaired DNA breaks or by nondisjunction of chromosomes. This improper segregation of chromosomes may result from hypomethylation of repeat sequences present in pericentromeric DNA, irregularities in kinetochore proteins or their assembly, a dysfunctional spindle apparatus, or flawed anaphase checkpoint genes. [2] Micronuclei can contribute to genome instability by promoting a catastrophic mutational event called chromothripsis. [3] Many micronucleus assays have been developed to test for the presence of these structures and determine their frequency in cells exposed to certain chemicals or subjected to stressful conditions.

The term micronucleus may also refer to the smaller nucleus in ciliate protozoans, such as the Paramecium . In mitosis it divides by fission, and in conjugation a pair of gamete micronuclei undergo reciprocal fusion to form a zygote nucleus, which gives rise to the macronuclei and micronuclei of the individuals of the next cycle of fission. [4]

Discovery

Micronuclei in newly formed red blood cells in humans are known as Howell-Jolly bodies because these structures were first identified and described in erythrocytes by hematologists William Howell and Justin Jolly. These structures were later found to be associated with deficiencies in vitamins such as folate and B12. The relationship between formation of micronuclei and exposure to environmental factors was first reported in root tip cells exposed to ionizing radiation. Micronucleus induction by a chemical was first reported in Ehrlich ascites tumor cells treated with colchicine. [2]

Formation

Micronuclei primarily result from acentric chromosome fragments or lagging whole chromosomes that are not included in the daughter nuclei produced by mitosis because they fail to correctly attach to the spindle during the segregation of chromosomes in anaphase. These full chromosomes or chromatid fragments are eventually enclosed by a nuclear membranes and are structurally similar to conventional nuclei, albeit smaller in size. This small nucleus is referred to as a micronucleus. The formation of micronuclei can only be observed in cells undergoing nuclear division and can be clearly seen using cytochalasin B to block cytokinesis to produce a binucleated cells. [2]

Acentric chromosome fragments may arise in a variety of ways. One way is that disrepair of DNA double-strand breaks can lead to symmetrical or asymmetrical chromatid and chromosome exchanges as well as chromatid and chromosome fragments. If DNA damage exceeds the repair capacity of the cell, unrepaired double-stranded DNA breaks may also result in acentric chromosome fragments. Another way eccentric chromosome fragments may arise is when defects in genes related to homologous recombinational repair (ex: ATM, BRCA1, BRCA2, and RAD51) result in a dysfunctional error-free homologous recombinational DNA repair pathway and causes the cell to resort to the error-prone non-homologous end-joining (NHEJ) repair pathway, increasing the likelihood of incorrect repair of DNA breaks, formation of dicentric chromosomes, and acentric chromosome fragments. If enzymes in the NHEJ repair pathway are defective as well, DNA breaks may not be repaired at all. Additionally, simultaneous excision repair of damaged or inappropriate bases incorporated in DNA that are in proximity and on opposite complementary DNA strands may lead to DNA double-stranded breaks and micronucleus formation, especially if the gap-filling step of the repair pathway is not completed. [2]

Micronuclei can also form from fragmented chromosomes when nucleoplasmic bridges (NPB) are formed, stretched, and broken during telophase. [2]

Micronuclei formation may also result from chromosome malsegregation during anaphase. Hypomethylation of cytosine in centromeric and pericentromeric areas and higher-order repeats of satellite DNA in centromeric DNA can result in such chromosomal loss events. Classical satellite DNA is normally heavily methylated at cytosine residues but may become almost fully unmethylated due to ICF syndrome (Immunodeficiency, centromere instability, and facial anomalies syndrome) or after treatment by DNA methyl transferase inhibitors. Since assembly of kinetochore proteins at centromeres is affected by the methylation of cytosine and histone proteins, a reduction in heterochromatin integrity as a result of hypomethylation can interfere with microtubule attachment to chromosomes and with the sensing of tension from correct microtubule-kinetochore connections. Other possible causes of chromosome loss that could lead to micronuclei formation are defects in kinetochore and microtubule interactions, defects in mitotic spindle assembly, mitosis check point defects, abnormal centrosome amplification, and telomeric end fusions that result in dicentric chromosomes that detach from the spindle during anaphase. Micronuclei originating from chromosome loss events and acentric chromosome fragments can be distinguished using pancentromeric DNA probes. [2]

Identification

The number of micronuclei per cell can be predicted using the following formula:

AF is the number of acentric fragments and F = 0.5 - 0.5P, where P equals the probability of fragments being included in the traditional nucleus and not forming a micronucleus. [5]

One study, which used Giemsa stain to stain nuclear material, established the following criteria for identifying micronuclei:
1) diameter less than 1/3 of the primary nucleus,
2) non-retractility (excludes small stain particles),
3) color the same as or lighter than the main nucleus (excludes large stain particles),
4) location within 3 or 4 nuclear diameters of the main nucleus without touching it, and
5) no more than two associated with one primary nucleus (3 or more micronuclei are likely polymorphs or prorubicytes with nuclear fragments). [6]

Assays

The micronucleus tests provide important information about a chemical's ability to interfere with chromosome structure and function. For instance, many known human carcinogens test positive in mammalian micronucleus tests. In these tests, organisms are treated with a chemical and the resulting frequency of micronucleated cells is measured. If there is a marked increase in the number of cells with micronuclei, it can be concluded that the chemical induces structural and/or numerical chromosomal damage. Since micronucleus tests must be performed on actively dividing cells, bone marrow stem cells and the erythrocytes they produce through cell divisions are ideal candidates. These cells experience constant, rapid turnover and the lack of a true nucleus in erythrocytes makes micronuclei easily visible under a microscope. [1]

Micronucleus assay systems are very economical, require much less skill in scoring that conventional metaphase tests, and are much faster than these conventional tests. Since micronucleus assays reflect chromosomal aberrations reliably and rapidly, they are extremely useful for a quick assessment of chromosomal damage. In particular, the CBMNcyt (cytokinesis-block micronucleus cytome) assay is extremely versatile and is one of the preferred methods to measure the level of chromosomal damage and chromosomal instability in cells. The cytokinesis-block micronucleus (CBMN) assay was first developed to score micronuclei in cells that completed nuclear division by blocking them at the binucleate stage before cytokinesis. It later evolved into the CBMN 'cytome' assay to further explore cell death, cytostasis, and biomarkers of DNA damage. The major drawback of using micronucleus tests is that they cannot determine different types of chromosomal aberrations and can be influenced by the mitotic rate and proportion of cell death, skewing the results. [2]

Patterns in formation

B, C. Micronuclei in peripheral blood erythrocytes of penguins Pygoscelis papua. Micronuclei and nuclear abnormalities in peripheral blood erythrocytes of penguins Pygoscelis papua 1.JPG
B, C. Micronuclei in peripheral blood erythrocytes of penguins Pygoscelis papua.

Multiple studies have found that micronuclei frequency in women is higher than in men and that the number of micronuclei increase until around 70 years of age. Micronuclei levels ranged from 0.5 to 1.4% in men to 0.9 to 1.8% in women. Gender-related differences were mainly seen in younger age groups (<= 50 years) with an almost two-fold difference between men and women. The patterns in the number of micronuclei after 70 years of age is controversial. Some studies have shown that in individuals over 70 years of age, micronucleus frequency increases in both sexes. On the other hand, other studies have found that in the oldest age groups, micronuclei frequencies level off. The deficiency of micronuclei in some of the oldest age groups may be explained by the fact that micro nucleated cells are preferentially eliminated by apoptosis. However, higher micronuclei frequency corresponds to a decreased efficiency of DNA repair and increased genomic instability, which are typical in older subjects. Age-related increases in micronuclei frequency also correspond well with age-related increases in the hypoploidy and the age-related increase in sex chromosome loss. Alternatively, the leveling off of frequency of micronuclei in older subjects would suggest a threshold of genomic instability that cannot be crossed if the person is to survive. If this were the case, women appear to reach this threshold faster than men. [7]

Sex chromosomes contribute to the majority of chromosome loss events with increasing age. In females, the X chromosome can account for up to 72% of the observed micronuclei of which 37% appear to be lacking a functional kinetochore assembly possibly due to X chromosome inactivation. Multiple studies have shown that the frequencies of autosome-positive micronuclei in both genders and of sex chromosome-positive MN in men were similar and remained unchanged in older groups while the frequency of X-positive MN in women was higher than the average frequency of autosome-positive MN and continued to increase until the oldest age. [2]

The frequencies of chromosomal aberrations, damaged cells, and micronuclei are significantly higher in smokers than non-smokers. [8]

In normal people and many other mammals, which do not have nuclei in their red blood cells, the micronuclei are removed rapidly by the spleen. Hence high frequencies of micronuclei in human peripheral blood indicate a ruptured or absent spleen. In mice, these are not removed, which is the basis for the in vivo micronucleus test.

See also


Related Research Articles

<span class="mw-page-title-main">Centromere</span> Specialized DNA sequence of a chromosome that links a pair of sister chromatids

The centromere links a pair of sister chromatids together during cell division. This constricted region of chromosome connects the sister chromatids, creating a short arm (p) and a long arm (q) on the chromatids. During mitosis, spindle fibers attach to the centromere via the kinetochore.

<span class="mw-page-title-main">Meiosis</span> Cell division producing haploid gametes

Meiosis is a special type of cell division of germ cells and apicomplexans in sexually-reproducing organisms that produces the gametes, such as sperm or egg cells. It involves two rounds of division that ultimately result in four cells with only one copy of each chromosome (haploid). Additionally, prior to the division, genetic material from the paternal and maternal copies of each chromosome is crossed over, creating new combinations of code on each chromosome. Later on, during fertilisation, the haploid cells produced by meiosis from a male and a female will fuse to create a cell with two copies of each chromosome again, the zygote.

<span class="mw-page-title-main">Mitosis</span> Process in which chromosomes are replicated and separated into two new identical nuclei

Mitosis is a part of the cell cycle in which replicated chromosomes are separated into two new nuclei. Cell division by mitosis is an equational division which gives rise to genetically identical cells in which the total number of chromosomes is maintained. Mitosis is preceded by the S phase of interphase and is followed by telophase and cytokinesis; which divides the cytoplasm, organelles and cell membrane of one cell into two new cells containing roughly equal shares of these cellular components. The different stages of mitosis altogether define the mitotic phase of a cell cycle—the division of the mother cell into two daughter cells genetically identical to each other.

<span class="mw-page-title-main">Cell division</span> Process by which living cells divide

Cell division is the process by which a parent cell divides into two daughter cells. Cell division usually occurs as part of a larger cell cycle in which the cell grows and replicates its chromosome(s) before dividing. In eukaryotes, there are two distinct types of cell division: a vegetative division (mitosis), producing daughter cells genetically identical to the parent cell, and a cell division that produces haploid gametes for sexual reproduction (meiosis), reducing the number of chromosomes from two of each type in the diploid parent cell to one of each type in the daughter cells. Mitosis is a part of the cell cycle, in which, replicated chromosomes are separated into two new nuclei. Cell division gives rise to genetically identical cells in which the total number of chromosomes is maintained. In general, mitosis is preceded by the S stage of interphase and is followed by telophase and cytokinesis; which divides the cytoplasm, organelles, and cell membrane of one cell into two new cells containing roughly equal shares of these cellular components. The different stages of mitosis all together define the M phase of an animal cell cycle—the division of the mother cell into two genetically identical daughter cells. To ensure proper progression through the cell cycle, DNA damage is detected and repaired at various checkpoints throughout the cycle. These checkpoints can halt progression through the cell cycle by inhibiting certain cyclin-CDK complexes. Meiosis undergoes two divisions resulting in four haploid daughter cells. Homologous chromosomes are separated in the first division of meiosis, such that each daughter cell has one copy of each chromosome. These chromosomes have already been replicated and have two sister chromatids which are then separated during the second division of meiosis. Both of these cell division cycles are used in the process of sexual reproduction at some point in their life cycle. Both are believed to be present in the last eukaryotic common ancestor.

<span class="mw-page-title-main">Anaphase</span> Stage of a cell division

Anaphase is the stage of mitosis after the process of metaphase, when replicated chromosomes are split and the newly-copied chromosomes are moved to opposite poles of the cell. Chromosomes also reach their overall maximum condensation in late anaphase, to help chromosome segregation and the re-formation of the nucleus.

<span class="mw-page-title-main">Chromatid</span> One of the two identical DNA molecules making up a duplicated chromosome

A chromatid is one half of a duplicated chromosome. Before replication, one chromosome is composed of one DNA molecule. In replication, the DNA molecule is copied, and the two molecules are known as chromatids. During the later stages of cell division these chromatids separate longitudinally to become individual chromosomes.

<span class="mw-page-title-main">Telophase</span> Final stage of a cell division for eukaryotic cells both in mitosis and meiosis

Telophase is the final stage in both meiosis and mitosis in a eukaryotic cell. During telophase, the effects of prophase and prometaphase are reversed. As chromosomes reach the cell poles, a nuclear envelope is re-assembled around each set of chromatids, the nucleoli reappear, and chromosomes begin to decondense back into the expanded chromatin that is present during interphase. The mitotic spindle is disassembled and remaining spindle microtubules are depolymerized. Telophase accounts for approximately 2% of the cell cycle's duration.

Genotoxicity is the property of chemical agents that damage the genetic information within a cell causing mutations, which may lead to cancer. While genotoxicity is often confused with mutagenicity, all mutagens are genotoxic, but some genotoxic substances are not mutagenic. The alteration can have direct or indirect effects on the DNA: the induction of mutations, mistimed event activation, and direct DNA damage leading to mutations. The permanent, heritable changes can affect either somatic cells of the organism or germ cells to be passed on to future generations. Cells prevent expression of the genotoxic mutation by either DNA repair or apoptosis; however, the damage may not always be fixed leading to mutagenesis.

<span class="mw-page-title-main">Spindle checkpoint</span> Cell cycle checkpoint

The spindle checkpoint, also known as the metaphase-to-anaphase transition, the spindle assembly checkpoint (SAC), the metaphase checkpoint, or the mitotic checkpoint, is a cell cycle checkpoint during metaphase of mitosis or meiosis that prevents the separation of the duplicated chromosomes (anaphase) until each chromosome is properly attached to the spindle. To achieve proper segregation, the two kinetochores on the sister chromatids must be attached to opposite spindle poles. Only this pattern of attachment will ensure that each daughter cell receives one copy of the chromosome. The defining biochemical feature of this checkpoint is the stimulation of the anaphase-promoting complex by M-phase cyclin-CDK complexes, which in turn causes the proteolytic destruction of cyclins and proteins that hold the sister chromatids together.

<span class="mw-page-title-main">Kinetochore</span> Protein complex that allows microtubules to attach to chromosomes during cell division

A kinetochore is a disc-shaped protein structure associated with duplicated chromatids in eukaryotic cells where the spindle fibers attach during cell division to pull sister chromatids apart. The kinetochore assembles on the centromere and links the chromosome to microtubule polymers from the mitotic spindle during mitosis and meiosis. The term kinetochore was first used in a footnote in a 1934 Cytology book by Lester W. Sharp and commonly accepted in 1936. Sharp's footnote reads: "The convenient term kinetochore has been suggested to the author by J. A. Moore", likely referring to John Alexander Moore who had joined Columbia University as a freshman in 1932.

<span class="mw-page-title-main">Cohesin</span> Protein complex that regulates the separation of sister chromatids during cell division

Cohesin is a protein complex that mediates sister chromatid cohesion, homologous recombination, and DNA looping. Cohesin is formed of SMC3, SMC1, SCC1 and SCC3. Cohesin holds sister chromatids together after DNA replication until anaphase when removal of cohesin leads to separation of sister chromatids. The complex forms a ring-like structure and it is believed that sister chromatids are held together by entrapment inside the cohesin ring. Cohesin is a member of the SMC family of protein complexes which includes Condensin, MukBEF and SMC-ScpAB.

A dicentric chromosome is an abnormal chromosome with two centromeres. It is formed through the fusion of two chromosome segments, each with a centromere, resulting in the loss of acentric fragments and the formation of dicentric fragments. The formation of dicentric chromosomes has been attributed to genetic processes, such as Robertsonian translocation and paracentric inversion. Dicentric chromosomes have important roles in the mitotic stability of chromosomes and the formation of pseudodicentric chromosomes. Their existence has been linked to certain natural phenomena such as irradiation and have been documented to underlie certain clinical syndromes, notably Kabuki syndrome. The formation of dicentric chromosomes and their implications on centromere function are studied in certain clinical cytogenetics laboratories.

<span class="mw-page-title-main">Clastogen</span> Substance that can cause breaks in chromosomes

A clastogen is a mutagenic agent that disturbs normal DNA related processes or directly causes DNA strand breakages, thus causing the deletion, insertion, or rearrangement of entire chromosome sections. These processes are a form of mutagenesis which if left unrepaired, or improperly repaired, can lead to cancer. Known clastogens include acridine yellow, benzene, ethylene oxide, arsenic, phosphine, mimosine, actinomycin D, camptothecin, methotrexate, methyl acrylate, resorcinol and 5-fluorodeoxyuridine. Additionally, 1,2-dimethylhydrazine is a known colon carcinogen and shows signs of possessing clastogenic activity. There are many clastogens not listed here and research is ongoing to discover new clastogens. Some known clastogens only exhibit clastogenic activity in certain cell types, such as caffeine which exhibits clastogenic activity in plant cells. Researchers are interested in clastogens for researching cancer, as well as for other human health concerns such as the inheritability of clastogen effected paternal germ cells that lead to fetus developmental defects.

<span class="mw-page-title-main">Micronucleus test</span> Test for potential genotoxic compounds

A micronucleus test is a test used in toxicological screening for potential genotoxic compounds. The assay is now recognized as one of the most successful and reliable assays for genotoxic carcinogens, i.e., carcinogens that act by causing genetic damage and is recommended by the OECD guideline for the testing of chemicals. There are two major versions of this test, one in vivo and the other in vitro.

Anaphase lag is a consequence of an event during cell division where sister chromatids do not properly separate from each other because of improper spindle formation. The chromosome or chromatid does not properly migrate during anaphase and the daughter cells will lose some genetic information. It is one of many causes of aneuploidy. This event can occur during both meiosis and mitosis with unique repercussions. In either case, anaphase lag will cause one daughter cell to receive a complete set of chromosomes while the other lacks one paired set of chromosomes, creating a form of monosomy. Whether the cell survives depends on which sister chromatid was lost and the background genomic state of the cell. The passage of abnormal numbers of chromosomes will have unique consequences with regards to mosaicism and development as well as the progression and heterogeneity of cancers.

<span class="mw-page-title-main">Chromothripsis</span> Massive chromosomal rearrangement process linked to cancer

Chromothripsis is a mutational process by which up to thousands of clustered chromosomal rearrangements occur in a single event in localised and confined genomic regions in one or a few chromosomes, and is known to be involved in both cancer and congenital diseases. It occurs through one massive genomic rearrangement during a single catastrophic event in the cell's history. It is believed that for the cell to be able to withstand such a destructive event, the occurrence of such an event must be the upper limit of what a cell can tolerate and survive. The chromothripsis phenomenon opposes the conventional theory that cancer is the gradual acquisition of genomic rearrangements and somatic mutations over time.

<span class="mw-page-title-main">Chromatin bridge</span> Medical condition

Chromatin bridge is a mitotic occurrence that forms when telomeres of sister chromatids fuse together and fail to completely segregate into their respective daughter cells. Because this event is most prevalent during anaphase, the term anaphase bridge is often used as a substitute. After the formation of individual daughter cells, the DNA bridge connecting homologous chromosomes remains fixed. As the daughter cells exit mitosis and re-enter interphase, the chromatin bridge becomes known as an interphase bridge. These phenomena are usually visualized using the laboratory techniques of staining and fluorescence microscopy.

<span class="mw-page-title-main">Cancerous micronuclei</span>

Cancerous micronuclei is a type of micronucleus that is associated with cancerous cells.

Chromosomal instability (CIN) is a type of genomic instability in which chromosomes are unstable, such that either whole chromosomes or parts of chromosomes are duplicated or deleted. More specifically, CIN refers to the increase in rate of addition or loss of entire chromosomes or sections of them. The unequal distribution of DNA to daughter cells upon mitosis results in a failure to maintain euploidy leading to aneuploidy. In other words, the daughter cells do not have the same number of chromosomes as the cell they originated from. Chromosomal instability is the most common form of genetic instability and cause of aneuploidy.

The association between obesity, as defined by a body mass index of 30 or higher, and risk of a variety of types of cancer has received a considerable amount of attention in recent years. Obesity has been associated with an increased risk of esophageal cancer, pancreatic cancer, colorectal cancer, breast cancer, endometrial cancer, kidney cancer, thyroid cancer, liver cancer and gallbladder cancer. Obesity may also lead to increased cancer-related mortality. Obesity has also been described as the fat tissue disease version of cancer, where common features between the two diseases were suggested for the first time.

References

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