Germline mutation

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Transmittance of a de novo mutation in germ cells to offspring. De novo mutations.png
Transmittance of a de novo mutation in germ cells to offspring.

A germline mutation, or germinal mutation, is any detectable variation within germ cells (cells that, when fully developed, become sperm and ova). [1] Mutations in these cells are the only mutations that can be passed on to offspring, when either a mutated sperm or oocyte come together to form a zygote. [2] After this fertilization event occurs, germ cells divide rapidly to produce all of the cells in the body, causing this mutation to be present in every somatic and germline cell in the offspring; this is also known as a constitutional mutation. [2] Germline mutation is distinct from somatic mutation.

Contents

Germline mutations can be caused by a variety of endogenous (internal) and exogenous (external) factors, and can occur throughout zygote development. [3] A mutation that arises only in germ cells can result in offspring with a genetic condition that is not present in either parent; this is because the mutation is not present in the rest of the parents' body, only the germline. [3]

When mutagenesis occurs

Germline mutations can occur before fertilization and during various stages of zygote development. [3] When the mutation arises will determine the effect it has on offspring. If the mutation arises in either the sperm or the oocyte before development, then the mutation will be present in every cell in the individual's body. [4] A mutation that arises soon after fertilization, but before germline and somatic cells are determined, then the mutation will be present in a large proportion of the individual's cell with no bias towards germline or somatic cells, this is also called a gonosomal mutation. [4] A mutation that arises later in zygote development will be present in a small subset of either somatic or germline cells, but not both. [3] [4]

Causes

Endogenous factors

A germline mutation often arises due to endogenous factors, like errors in cellular replication and oxidative damage. [5] This damage is rarely repaired imperfectly, but due to the high rate of germ cell division, can occur frequently. [5]

Endogenous mutations are more prominent in sperm than in ova. [6] This is because spermatocytes go through a larger number of cell divisions throughout a male's life, resulting in more replication cycles that could result in a DNA mutation. [5] Errors in maternal ovum also occur, but at a lower rate than in paternal sperm. [5] The types of mutations that occur also tend to vary between the sexes. [7] A mother's eggs, after production, remain in stasis until each is utilized in ovulation. This long stasis period has been shown to result in a higher number of chromosomal and large sequence deletions, duplications, insertions, and transversions. [7] The father's sperm, on the other hand, undergoes continuous replication throughout his lifetime, resulting in many small point mutations that result from errors in replication. These mutations commonly include single base pair substitutions, deletions, and insertions. [6]

Oxidative damage is another endogenous factor that can cause germline mutations. This type of damage is caused by reactive oxygen species that build up in the cell as a by-product of cellular respiration. [8] These reactive oxygen species are missing an electron, and because they are highly electronegative (have a strong electron pull) they will rip an electron away from another molecule. [8] This can initiate DNA damage because it causes the nucleic acid guanine to shift to 8-oxoguanine (8-oxoG). This 8-oxoG molecule is then mistaken for a thymine by DNA polymerase during replication, causing a G>T transversion on one DNA strand, and a C>A transversion on the other. [9]

Male germline

In mice and humans the spontaneous mutation rate in the male germ line is significantly lower than in somatic cells. [10] Furthermore, although the spontaneous mutation rate in the male germ line increases with age, the rate of increase is lower than in somatic tissues. Within the testicular spermatogonial stem cell population the integrity of DNA appears to be maintained by highly effective DNA damage surveillance and protective DNA repair processes. [10] The progressive increase in the mutation rate with age in the male germ line may be a result of a decline in the accuracy of the repair of DNA damages, or of an increase in DNA replication errors. Once spermatogenesis is complete, the differentiated spermatozoa that are formed no longer have the capability for DNA repair, and are thus vulnerable to attack by prevalent oxidative free radicals that cause oxidative DNA damage. Such damaged spermatozoa may undergo programmed cell death (apoptosis). [10]

Exogenous factors

A germline mutation can also occur due to exogenous factors. Similar to somatic mutations, germline mutations can be caused by exposure to harmful substances, which damage the DNA of germ cells. This damage can then either be repaired perfectly, and no mutations will be present, or repaired imperfectly, resulting in a variety of mutations. [11] Exogenous mutagens include harmful chemicals and ionizing radiation; the major difference between germline mutations and somatic mutations is that germ cells are not exposed to UV radiation, and thus not often directly mutated in this manner. [12] [13]

Clinical implications

Different germline mutations can affect an individual differently depending on the rest of their genome. A dominant mutation only requires 1 mutated gene to produce the disease phenotype, while a recessive mutation requires both alleles to be mutated to produce the disease phenotype. [14] For example, if the embryo inherits an already mutated allele from the father, and the same allele from the mother underwent an endogenous mutation, then the child will display the disease related to that mutated gene, even though only 1 parent carries the mutant allele. [14] This is only one example of how a child can display a recessive disease while a mutant gene is only carried by one parent. [14] Detection of chromosomal abnormalities can be found in utero for certain diseases by means of blood samples or ultrasound, as well as invasive procedures such as an amniocentesis. Later detection can be found by genome screening.

Cancer

Mutations in tumour suppressor genes or proto-oncogenes can predispose an individual to developing tumours. [15] It is estimated that inherited genetic mutations are involved in 5-10% of cancers. [16] These mutations make a person susceptible to tumour development if the other copy of the oncogene is randomly mutated. These mutations can occur in germ cells, allowing them to be heritable. [15] Individuals who inherit germline mutations in TP53 are predisposed to certain cancer variants because the protein produced by this gene suppresses tumors. Patients with this mutation are also at a risk for Li–Fraumeni syndrome. [16] Other examples include mutations in the BRCA1 and BRCA2 genes which predispose to breast and ovarian cancer, or mutations in MLH1 which predispose to hereditary non-polyposis colorectal cancer.

Huntington's disease

Huntington's disease is an autosomal dominant mutation in the HTT gene. The disorder causes degradation in the brain, resulting in uncontrollable movements and behavior. [17] The mutation involves an expansion of repeats in the Huntington protein, causing it to increase in size. Patients who have more than 40 repeats will most likely be affected. The onset of the disease is determined by the amount of repeats present in the mutation; the greater the number of repeats, the earlier symptoms of the disease will appear. [17] [18] Because of the dominant nature of the mutation, only one mutated allele is needed for the disease to be in effect. This means that if one parent is affected, the child will have a 50% chance of inheriting the disease. [19] This disease does not have carriers because if a patient has one mutation, they will (most likely) be affected. The disease typically has a late onset, so many parents have children before they know they have the mutation. The HTT mutation can be detected through genome screening.

Trisomy 21

Trisomy 21 (also known as Down syndrome) results from a child having 3 copies of chromosome 21. [20] This chromosome duplication occurs during germ cell formation, when both copies of chromosome 21 end up in the same daughter cell in either the mother or father, and this mutant germ cell participates in fertilization of the zygote. [20] Another, more common way this can occur is during the first cell division event after the formation of the zygote. [20] The risk of Trisomy 21 increases with maternal age with the risk being 1/2000 (0.05%) at age 20 increasing to 1/100 (1%) at age 40. [21] This disease can be detected by non-invasive as well as invasive procedures prenatally. Non-invasive procedures include scanning for fetal DNA through maternal plasma via a blood sample. [22]

Cystic fibrosis

Cystic fibrosis is an autosomal recessive disorder that causes a variety of symptoms and complications, the most common of which is a thick mucus lining in lung epithelial tissue due to improper salt exchange, but can also affect the pancreas, intestines, liver, and kidneys. [23] [24] Many bodily processes can be affected due to the hereditary nature of this disease; if the disease is present in the DNA of both the sperm and the egg, then it will be present in essentially every cell and organ in the body; these mutations can occur initially in the germline cells, or be present in all parental cells. [23] The most common mutation seen in this disease is ΔF508, which means a deletion of the amino acid at the 508 position. [25] If both parents have a mutated CFTR (cystic fibrosis transmembrane conductance regulator) protein, then their children have a 25% of inheriting the disease. [23] If a child has 1 mutated copy of CFTR, they will not develop the disease, but will become a carrier of the disease. [23] The mutation can be detected before birth through amniocentesis, or after birth via prenatal genetic screening. [26]

Current therapies

Many Mendelian disorders stem from dominant point mutations within genes, including cystic fibrosis, beta-thalassemia, sickle-cell anemia, and Tay–Sachs disease. [14] By inducing a double stranded break in sequences surrounding the disease-causing point mutation, a dividing cell can use the non-mutated strand as a template to repair the newly broken DNA strand, getting rid of the disease-causing mutation. [27] Many different genome editing techniques have been used for genome editing, and especially germline mutation editing in germ cells and developing zygotes; however, while these therapies have been extensively studied, their use in human germline editing is limited. [28]

CRISPR/Cas9 editing

The CRISPR editing system is able to target specific DNA sequences and, using a donor DNA template, can repair mutations within this gene. DNA Repair after CRISPR-Cas9 cut.svg
The CRISPR editing system is able to target specific DNA sequences and, using a donor DNA template, can repair mutations within this gene.

This editing system induces a double stranded break in the DNA, using a guide RNA and effector protein Cas9 to break the DNA backbones at specific target sequences. [27] This system has shown a higher specificity than TALENs or ZFNs due to the Cas9 protein containing homologous (complementary) sequences to the sections of DNA surrounding the site to be cleaved. [27]  This broken strand can be repaired in 2 main ways: homologous directed repair (HDR) if a DNA strand is present to be used as a template (either homologous or donor), and if one is not, then the sequence will undergo non-homologous end joining (NHEJ). [27] NHEJ often results in insertions or deletions within the gene of interest, due to the processing of the blunt strand ends, and is a way to study gene knockouts in a lab setting. [29] This method can be used to repair a point mutation by using the sister chromosome as a template, or by providing a double stranded DNA template with the CRISPR/Cas9 machinery to be used as the repair template. [27]

This method has been used in both human and animal models ( Drosophila , Mus musculus , and Arabidopsis ), and current research is being focused on making this system more specific to minimize off-target cleavage sites. [30]

TALEN editing

The TALEN (transcription activator-like effector nucleases) genome editing system is used to induce a double-stranded DNA break at a specific locus in the genome, which can then be used to mutate or repair the DNA sequence. [31] It functions by using a specific repeated sequence of an amino acid that is 33-34 amino acids in length. [31] The specificity of the DNA binding site is determined by the specific amino acids at positions 12 and 13 (also called the Repeat Variable Diresidue (RVD)) of this tandem repeat, with some RVDs showing a higher specificity for specific amino acids over others. [32] Once the DNA break is initiated, the ends can either be joined with NHEJ that induces mutations, or by HDR that can fix mutations. [27]

ZFN editing

Similar to TALENs, zinc finger nucleases (ZFNs) are used to create a double stranded break in the DNA at a specific locus in the genome. [31] The ZFN editing complex consists of a zinc finger protein (ZFP) and a restriction enzyme cleavage domain. [33] The ZNP domain can be altered to change the DNA sequence that the restriction enzyme cuts, and this cleavage event initiates cellular repair processes, similar to that of CRISPR/Cas9 DNA editing. [33]

Compared to CRISPR/Cas9, the therapeutic applications of this technology are limited, due to the extensive engineering required to make each ZFN specific to the desired sequence. [33]

See also

Related Research Articles

<span class="mw-page-title-main">Mutation</span> Alteration in the nucleotide sequence of a genome

In biology, a mutation is an alteration in the nucleic acid sequence of the genome of an organism, virus, or extrachromosomal DNA. Viral genomes contain either DNA or RNA. Mutations result from errors during DNA or viral replication, mitosis, or meiosis or other types of damage to DNA, which then may undergo error-prone repair, cause an error during other forms of repair, or cause an error during replication. Mutations may also result from insertion or deletion of segments of DNA due to mobile genetic elements.

Gene knockouts are a widely used genetic engineering technique that involves the targeted removal or inactivation of a specific gene within an organism's genome. This can be done through a variety of methods, including homologous recombination, CRISPR-Cas9, and TALENs.

In cellular biology, a somatic cell, or vegetal cell, is any biological cell forming the body of a multicellular organism other than a gamete, germ cell, gametocyte or undifferentiated stem cell. Such cells compose the body of an organism and divide through the process of binary fission and mitotic division.

In cellular biology, the term somatic is often used to refer to the cells of the body, in contrast to the reproductive (germline) cells, which usually give rise to the egg or sperm. These somatic cells are diploid, containing two copies of each chromosome, whereas germ cells are haploid, as they only contain one copy of each chromosome. Although under normal circumstances all somatic cells in an organism contain identical DNA, they develop a variety of tissue-specific characteristics. This process is called differentiation, through epigenetic and regulatory alterations. The grouping of similar cells and tissues creates the foundation for organs.

A genetic screen or mutagenesis screen is an experimental technique used to identify and select individuals who possess a phenotype of interest in a mutagenized population. Hence a genetic screen is a type of phenotypic screen. Genetic screens can provide important information on gene function as well as the molecular events that underlie a biological process or pathway. While genome projects have identified an extensive inventory of genes in many different organisms, genetic screens can provide valuable insight as to how those genes function.

<span class="mw-page-title-main">Germline</span> Population of a multicellular organisms cells that pass on their genetic material to the progeny

In biology and genetics, the germline is the population of a multicellular organism's cells that pass on their genetic material to the progeny (offspring). In other words, they are the cells that form the egg, sperm and the fertilised egg. They are usually differentiated to perform this function and segregated in a specific place away from other bodily cells.

Gene knockdown is an experimental technique by which the expression of one or more of an organism's genes is reduced. The reduction can occur either through genetic modification or by treatment with a reagent such as a short DNA or RNA oligonucleotide that has a sequence complementary to either gene or an mRNA transcript.

<span class="mw-page-title-main">Mosaic (genetics)</span> Condition in multi-cellular organisms

Mosaicism or genetic mosaicism is a condition in which a multicellular organism possesses more than one genetic line as the result of genetic mutation. This means that various genetic lines resulted from a single fertilized egg. Mosaicism is one of several possible causes of chimerism, wherein a single organism is composed of cells with more than one distinct genotype.

The term Modifications in genetics refers to both naturally occurring and engineered changes in DNA. Incidental, or natural mutations occur through errors during replication and repair, either spontaneously or due to environmental stressors. Intentional modifications are done in a laboratory for various purposes, developing hardier seeds and plants, and increasingly to treat human disease. The use of gene editing technology remains controversial.

<span class="mw-page-title-main">Designer baby</span> Genetically modified human embryo

A designer baby is a baby whose genetic makeup has been selected or altered, often to exclude a particular gene or to remove genes associated with disease. This process usually involves analysing a wide range of human embryos to identify genes associated with particular diseases and characteristics, and selecting embryos that have the desired genetic makeup; a process known as preimplantation genetic diagnosis. Screening for single genes is commonly practiced, and polygenic screening is offered by a few companies. Other methods by which a baby's genetic information can be altered involve directly editing the genome before birth, which is not routinely performed and only one instance of this is known to have occurred as of 2019, where Chinese twins Lulu and Nana were edited as embryos, causing widespread criticism.

A postzygotic mutation is a change in an organism's genome that is acquired during its lifespan, instead of being inherited from its parent(s) through fusion of two haploid gametes. Mutations that occur after the zygote has formed can be caused by a variety of sources that fall under two classes: spontaneous mutations and induced mutations. How detrimental a mutation is to an organism is dependent on what the mutation is, where it occurred in the genome and when it occurred.

<span class="mw-page-title-main">Insert (molecular biology)</span>

In Molecular biology, an insert is a piece of DNA that is inserted into a larger DNA vector by a recombinant DNA technique, such as ligation or recombination. This allows it to be multiplied, selected, further manipulated or expressed in a host organism.

<span class="mw-page-title-main">Transcription activator-like effector nuclease</span>

Transcription activator-like effector nucleases (TALEN) are restriction enzymes that can be engineered to cut specific sequences of DNA. They are made by fusing a TAL effector DNA-binding domain to a DNA cleavage domain. Transcription activator-like effectors (TALEs) can be engineered to bind to practically any desired DNA sequence, so when combined with a nuclease, DNA can be cut at specific locations. The restriction enzymes can be introduced into cells, for use in gene editing or for genome editing in situ, a technique known as genome editing with engineered nucleases. Alongside zinc finger nucleases and CRISPR/Cas9, TALEN is a prominent tool in the field of genome editing.

<span class="mw-page-title-main">Genome editing</span> Type of genetic engineering

Genome editing, or genome engineering, or gene editing, is a type of genetic engineering in which DNA is inserted, deleted, modified or replaced in the genome of a living organism. Unlike early genetic engineering techniques that randomly inserts genetic material into a host genome, genome editing targets the insertions to site-specific locations. The basic mechanism involved in genetic manipulations through programmable nucleases is the recognition of target genomic loci and binding of effector DNA-binding domain (DBD), double-strand breaks (DSBs) in target DNA by the restriction endonucleases, and the repair of DSBs through homology-directed recombination (HDR) or non-homologous end joining (NHEJ).

<span class="mw-page-title-main">Genetic engineering techniques</span> Methods used to change the DNA of organisms

Genetic engineering techniques allow the modification of animal and plant genomes. Techniques have been devised to insert, delete, and modify DNA at multiple levels, ranging from a specific base pair in a specific gene to entire genes. There are a number of steps that are followed before a genetically modified organism (GMO) is created. Genetic engineers must first choose what gene they wish to insert, modify, or delete. The gene must then be isolated and incorporated, along with other genetic elements, into a suitable vector. This vector is then used to insert the gene into the host genome, creating a transgenic or edited organism.

Human germline engineering is the process by which the genome of an individual is edited in such a way that the change is heritable. This is achieved through genetic alterations within the germ cells, or the reproductive cells, such as the egg and sperm. Human germline engineering is a type of genetic modification that directly manipulates the genome using molecular engineering techniques. Aside from germline engineering, genetic modification can be applied in another way, somatic genetic modification. Somatic gene modification consists of altering somatic cells, which are all cells in the body that are not involved in reproduction. While somatic gene therapy does change the genome of the targeted cells, these cells are not within the germline, so the alterations are not heritable and cannot be passed on to the next generation.

Off-target genome editing refers to nonspecific and unintended genetic modifications that can arise through the use of engineered nuclease technologies such as: clustered, regularly interspaced, short palindromic repeats (CRISPR)-Cas9, transcription activator-like effector nucleases (TALEN), meganucleases, and zinc finger nucleases (ZFN). These tools use different mechanisms to bind a predetermined sequence of DNA (“target”), which they cleave, creating a double-stranded chromosomal break (DSB) that summons the cell's DNA repair mechanisms and leads to site-specific modifications. If these complexes do not bind at the target, often a result of homologous sequences and/or mismatch tolerance, they will cleave off-target DSB and cause non-specific genetic modifications. Specifically, off-target effects consist of unintended point mutations, deletions, insertions inversions, and translocations.

<span class="mw-page-title-main">He Jiankui affair</span> 2018 scientific and bioethical controversy

The He Jiankui affair is a scientific and bioethical controversy concerning the use of genome editing following its first use on humans by Chinese scientist He Jiankui, who edited the genomes of human embryos in 2018. He became widely known on 26 November 2018 after he announced that he had created the first human genetically edited babies. He was listed in the Time's 100 most influential people of 2019. The affair led to legal and ethical controversies, resulting in the indictment of He and two of his collaborators, Zhang Renli and Qin Jinzhou. He eventually received widespread international condemnation.

<span class="mw-page-title-main">CRISPR gene editing</span> Gene editing method

CRISPR gene editing is a genetic engineering technique in molecular biology by which the genomes of living organisms may be modified. It is based on a simplified version of the bacterial CRISPR-Cas9 antiviral defense system. By delivering the Cas9 nuclease complexed with a synthetic guide RNA (gRNA) into a cell, the cell's genome can be cut at a desired location, allowing existing genes to be removed and/or new ones added in vivo.

A somatic mutation is a change in the DNA sequence of a somatic cell of a multicellular organism with dedicated reproductive cells; that is, any mutation that occurs in a cell other than a gamete, germ cell, or gametocyte. Unlike germline mutations, which can be passed on to the descendants of an organism, somatic mutations are not usually transmitted to descendants. This distinction is blurred in plants, which lack a dedicated germline, and in those animals that can reproduce asexually through mechanisms such as budding, as in members of the cnidarian genus Hydra.

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