Extrachromosomal circular DNA (eccDNA) is a type of double-stranded circular DNA structure that was first discovered in 1964 by Alix Bassel and Yasuo Hotta. In contrast to previously identified circular DNA structures (e.g., bacterial

plasmid A plasmid is a small, extrachromosomal DNA molecule within a cell that is physically separated from chromosomal DNA and can replicate independently. They are most commonly found as small circular, double-stranded DNA molecules in bacteria and ...

mitochondrial DNA Mitochondrial DNA (mtDNA and mDNA) is the DNA located in the mitochondrion, mitochondria organelles in a eukaryotic cell that converts chemical energy from food into adenosine triphosphate (ATP). Mitochondrial DNA is a small portion of the D ...

circular bacterial chromosome A circular chromosome is a chromosome in bacteria, archaea, mitochondria, and chloroplasts, in the form of a molecule of circular DNA, unlike the linear chromosome of most eukaryotes. Most prokaryote chromosomes contain a circular DNA molecule. ...

s, or

chloroplast DNA Chloroplast DNA (cpDNA), also known as plastid DNA (ptDNA) is the DNA located in chloroplasts, which are photosynthetic organelles located within the cells of some eukaryotic organisms. Chloroplasts, like other types of plastid, contain a genome s ...

), eccDNA are circular DNA found in the

eukaryotic The eukaryotes ( ) constitute the Domain (biology), domain of Eukaryota or Eukarya, organisms whose Cell (biology), cells have a membrane-bound cell nucleus, nucleus. All animals, plants, Fungus, fungi, seaweeds, and many unicellular organisms ...

nuclei of plant and animal (including human) cells. Extrachromosomal circular DNA is derived from chromosomal DNA, can range in size from 50

base pair A base pair (bp) is a fundamental unit of double-stranded nucleic acids consisting of two nucleobases bound to each other by hydrogen bonds. They form the building blocks of the DNA double helix and contribute to the folded structure of both DNA ...

s to several mega-base pairs in length, and can encode

regulatory element A regulatory sequence is a segment of a nucleic acid molecule which is capable of increasing or decreasing the expression of specific genes within an organism. Regulation of gene expression is an essential feature of all living organisms and vir ...

s and full-length

gene In biology, the word gene has two meanings. The Mendelian gene is a basic unit of heredity. The molecular gene is a sequence of nucleotides in DNA that is transcribed to produce a functional RNA. There are two types of molecular genes: protei ...

s. eccDNA has been observed in various eukaryotic species and it is proposed to be a byproduct of programmed DNA recombination events, such as

V(D)J recombination V(D)J recombination (variable–diversity–joining rearrangement) is the mechanism of somatic recombination that occurs only in developing lymphocytes during the early stages of T and B cell maturation. It results in the highly diverse repertoire ...

Historical Background

In 1964, Bassel and Hotta published their initial discovery of eccDNA that they made while researching

Franklin Stahl Franklin (Frank) William Stahl (born October 8, 1929) is an American molecular biologist and geneticist. With Matthew Meselson, Stahl conducted the famous Meselson-Stahl experiment showing that DNA is replicated by a semiconservative mechanism ...

’s chromosomal theory. In their experiments, they visualized isolated wheat nuclei and boar sperm by using

electron microscopy An electron microscope is a microscope that uses a beam of electrons as a source of illumination. It uses electron optics that are analogous to the glass lenses of an optical light microscope to control the electron beam, for instance focusing i ...

. Their research found that boar sperm cells contained eccDNA of various sizes. In 1965, Arthur Spriggs’ research group identified eccDNA in the samples of five pediatric patients’ embryonic tumors and one adult patient’s bronchial

carcinoma Carcinoma is a malignancy that develops from epithelial cells. Specifically, a carcinoma is a cancer that begins in a tissue that lines the inner or outer surfaces of the body, and that arises from cells originating in the endodermal, mesoder ...

. In the following years, additional research led to the discovery of eccDNA in various species listed in Table 1:

21st Century Research

In the 21st century, researchers have focused on better characterizing the specific subtypes of eccDNA, as well as the structure and function of these molecules within biological systems: * In 2012, Shibata et al. discovered a specific type of eccDNA called

microDNA MicroDNA is the most abundant subtype of Extrachromosomal Circular DNA (eccDNA) in humans, typically ranging from 200-400 base pairs in length and enriched in non-repetitive genomic sequences with a high density of exons. Additionally, microDNA h ...

. The researchers found tens of thousands of

s in mouse tissues and cell lines, as well as human cell lines. * In 2017, Turner et al. identified using

whole-genome sequencing Whole genome sequencing (WGS), also known as full genome sequencing or just genome sequencing, is the process of determining the entirety of the DNA sequence of an organism's genome at a single time. This entails sequencing all of an organism's ...

(WGS),

cytogenetic analysis Cytogenetics is essentially a branch of genetics, but is also a part of cell biology/cytology (a subdivision of human anatomy), that is concerned with how the chromosomes relate to cell behaviour, particularly to their behaviour during mitosis an ...

, and structural modeling that extrachromosomal circular DNA is highly amplified and common in various types of

cancer Cancer is a group of diseases involving Cell growth#Disorders, abnormal cell growth with the potential to Invasion (cancer), invade or Metastasis, spread to other parts of the body. These contrast with benign tumors, which do not spread. Po ...

s. They found that eccDNA molecules have significant heterogeneity between different cells even if they are derived from the same individual. Furthermore, these eccDNA molecules contained tumor-driving genes and were reported to be rarely found in non-cancerous tissues. * In 2018, Møller et al. used healthy human muscle and

blood cell A blood cell (also called a hematopoietic cell, hemocyte, or hematocyte) is a cell produced through hematopoiesis and found mainly in the blood. Major types of blood cells include red blood cells (erythrocytes), white blood cells (leukocytes), ...

samples to identify over 100,000 types of eccDNA, which suggested that eccDNA could be found within

somatic cell 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. Somatic cells compose the body of an organism ...

s ubiquitously. * In 2019, Wu et al. found that ecDNA (subtype of eccDNA) associates with

chromatin Chromatin is a complex of DNA and protein found in eukaryote, eukaryotic cells. The primary function is to package long DNA molecules into more compact, denser structures. This prevents the strands from becoming tangled and also plays important r ...

, but unlike

chromosome A chromosome is a package of DNA containing part or all of the genetic material of an organism. In most chromosomes, the very long thin DNA fibers are coated with nucleosome-forming packaging proteins; in eukaryotic cells, the most import ...

s it does not have higher-order compaction, which increases its accessibility. * In 2021, Wang et al. elaborated on the formation of eccDNAs and identified the

immunostimulant Immunostimulants, also known as immunostimulators, are substances (drugs and nutrients) that stimulate the immune system usually in a non-specific manner by inducing activation or increasing activity of any of its components. One notable example i ...

function of eccDNAs. They also developed an improved eccDNA purification protocol that decreases linear DNA contamination within purified samples.

eccDNA Purification

Historically, eccDNA was purified using a two-step procedure that involved first isolating crude extrachromosomal DNA and subsequently digesting linear DNA via

exonuclease Exonucleases are enzymes that work by cleaving nucleotides one at a time from the end (exo) of a polynucleotide chain. A hydrolyzing reaction that breaks phosphodiester bonds at either the 3′ or the 5′ end occurs. Its close relative is th ...

digestion Digestion is the breakdown of large insoluble food compounds into small water-soluble components so that they can be absorbed into the blood plasma. In certain organisms, these smaller substances are absorbed through the small intestine into th ...

. Yet, this technique often results in linear DNA contamination because exonuclease digestion is not sufficient to remove all linear DNA. In 2021, Wang et al. developed a three-step eccDNA enrichment method that improved eccDNA purification: * The cells were first dehydrated in > 90% methanol. To extract crude extrachromosomal DNA, the cells were lysed with a pH 11.8 alkaline lysis buffer, neutralized with a neutralization buffer, and precipitated using a precipitation buffer. A commercial plasmid purification kit's silica column was used to isolate DNA from other cell components. * The eluted DNA was digested with the

restriction enzyme A restriction enzyme, restriction endonuclease, REase, ENase or'' restrictase '' is an enzyme that cleaves DNA into fragments at or near specific recognition sites within molecules known as restriction sites. Restriction enzymes are one class o ...

''PacI'' to linearize

(

mtDNA Mitochondrial DNA (mtDNA and mDNA) is the DNA located in the mitochondria organelles in a eukaryotic cell that converts chemical energy from food into adenosine triphosphate (ATP). Mitochondrial DNA is a small portion of the DNA contained in ...

) and an exonuclease that can digest linear DNA. * Finally, circular DNA was selectively recovered by a commercial solution and silica beads to remove linear DNA that was not removed by exonuclease digestion.

Double minutes (DM) vs. extrachromosomal circular DNA (eccDNA)

Initially, the term

double minute Double minutes (DMs) are small fragments of extrachromosomal DNA, which have been observed in a large number of human tumors including breast, lung, ovary, colon, and most notably, neuroblastoma. They are a manifestation of gene amplification as a ...

s (DM) was commonly used to refer to extrachromosomal circular DNA because it often appeared as a pair in early studies. As research has continued, different subtypes of extrachromosomal circular DNA have been identified that are not double minutes (e.g.,

). In 2014, Barreto et al. identified that double minutes only comprise roughly 30% of extrachromosomal DNA. Thus, the term extrachromosomal circular DNA (eccDNA) is becoming more widely used, while the term double minutes is now reserved for a specific subtype of eccDNA.

Structure

eccDNA are circular DNA that have been found in human, plant, and animal cells and are present in the

cell nucleus The cell nucleus (; : nuclei) is a membrane-bound organelle found in eukaryote, eukaryotic cell (biology), cells. Eukaryotic cells usually have a single nucleus, but a few cell types, such as mammalian red blood cells, have #Anucleated_cells, ...

in addition to the chromosomal

DNA Deoxyribonucleic acid (; DNA) is a polymer composed of two polynucleotide chains that coil around each other to form a double helix. The polymer carries genetic instructions for the development, functioning, growth and reproduction of al ...

. eccDNA is distinguishable from other circular DNA in cells, such as

(mtDNA), because it ranges in size from a few hundred bases to megabases and is derived from genomic DNA. For example, eccDNA can be formed from

exon An exon is any part of a gene that will form a part of the final mature RNA produced by that gene after introns have been removed by RNA splicing. The term ''exon'' refers to both the DNA sequence within a gene and to the corresponding sequence ...

s of protein coding genes, like

mucin Mucins () are a family of high molecular weight, heavily glycosylated proteins ( glycoconjugates) produced by epithelial tissues in most animals. Mucins' key characteristic is their ability to form gels; therefore they are a key component in ...

and

titin Titin (; also called connectin) is a protein that in humans is encoded by the ''TTN'' gene. The protein, which is over 1 μm in length, functions as a molecular spring that is responsible for the passive elasticity of muscle. It comprises 2 ...

. Researchers have hypothesized that eccDNA may contribute to the expression of different

isoforms A protein isoform, or "protein variant", is a member of a set of highly similar proteins that originate from a single gene and are the result of genetic differences. While many perform the same or similar biological roles, some isoforms have uniqu ...

of a gene by interfering with or promoting the

transcription Transcription refers to the process of converting sounds (voice, music etc.) into letters or musical notes, or producing a copy of something in another medium, including: Genetics * Transcription (biology), the copying of DNA into RNA, often th ...

of specific

s. eccDNA has been classified as one of four different categories of circular DNA based on size and sequence, including small polydispersed circular DNA (spcDNA), telomeric circles (t-circles),

(100-400 bp), and extrachromosomal DNA (ecDNA). Each of these types has its own unique biological characteristics (see Table 2):

eccDNA biogenesis

While the exact mechanism for eccDNA generation is still unknown, some studies have suggested that eccDNA generation might be linked to DNA damage repair, hyper-transcription,

homologous recombination Homologous recombination is a type of genetic recombination in which genetic information is exchanged between two similar or identical molecules of double-stranded or single-stranded nucleic acids (usually DNA as in Cell (biology), cellular organi ...

, and

replication stress DNA replication stress refers to the state of a cell whose genome is exposed to various stresses. The events that contribute to replication stress occur during DNA replication, and can result in a stalled replication fork. There are many events t ...

. There are multiple proposed mechanisms for eccDNA formation: (1)

replication slippage Replication may refer to: Science * Replication (scientific method), one of the main principles of the scientific method, a.k.a. reproducibility ** Replication (statistics), the repetition of a test or complete experiment ** Replication crisis * ...

creates a loop on the

template strand Transcription is the process of copying a segment of DNA into RNA for the purpose of gene expression. Some segments of DNA are transcribed into RNA molecules that can encode proteins, called messenger RNA (mRNA). Other segments of DNA are transc ...

that is then excised and ligated into a circle leaving a

microdeletion In genetics, a deletion (also called gene deletion, deficiency, or Deletion anomaly, deletion mutation) (sign: delta (letter), Δ) is a mutation (a genetic aberration) in which a part of a chromosome or a sequence of DNA is left out during DNA rep ...

on the

, (2)

creates a loop in the product strand that is excised and ligated into a circle that does not generate a

in the

, (3) the ODERA mechanism of eccDNA formation, and (4) a double stranded break in a repeat region is repaired by

, during which the fragment forms a circle and the chromosome suffers a

Research conducted in 2021 demonstrated that

apoptotic Apoptosis (from ) is a form of programmed cell death that occurs in multicellular organisms and in some eukaryotic, single-celled microorganisms such as yeast. Biochemical events lead to characteristic cell changes ( morphology) and death. These ...

cells are a source of eccDNAs; this was concluded on account of the study showing that

DNA fragmentation DNA fragmentation is the separation or breaking of DNA strands into pieces. It can be done intentionally by laboratory personnel or by cells, or can occur spontaneously. Spontaneous or accidental DNA fragmentation is fragmentation that gradually a ...

(ADF) is a prerequisite for eccDNA formation through purification methods. eccDNA can be generated as a result of micro-nuclei formation, indicating

chromosomal instability 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 addi ...

. It has been proposed that premature apoptosis and/or errors in chromosomal segregation during mitosis could lead to micro-nuclei formation.

eccDNA in non-cancerous cells

To test whether eccDNAs occur in non-cancer cells, mouse embryonic

stem cell In multicellular organisms, stem cells are undifferentiated or partially differentiated cells that can change into various types of cells and proliferate indefinitely to produce more of the same stem cell. They are the earliest type of cell ...

s and

Southern Blot Southern blot is a method used for detection and quantification of a specific DNA sequence in DNA samples. This method is used in molecular biology. Briefly, purified DNA from a biological sample (such as blood or tissue) is digested with res ...

analysis were used; the results confirmed that eccDNA is found in both cancerous and non-cancerous cells. It is also known that eccDNA is unlikely to be derived from specific genome regions; sequencing data from 2021 reports that the data suggests eccDNAs are widespread across the entirety of the

genome A genome is all the genetic information of an organism. It consists of nucleotide sequences of DNA (or RNA in RNA viruses). The nuclear genome includes protein-coding genes and non-coding genes, other functional regions of the genome such as ...

. Genome mapping of full-length eccDNAs demonstrated their different genomic alignment patterns, which includes at adjacent, overlapped, or nested positions on the same

or across different

s. eccDNAs originate mostly from single, continuous genomic loci, meaning that one single genomic fragment self-circularizes to form the eccDNA, rather than being formed from ligation of different genomic fragments. These two variants can be classified as continuous and non-continuous eccDNAs, respectively. To further understand the reason behind the circularization of fragmented DNA, the three various mammalian

ligase In biochemistry, a ligase is an enzyme that can catalyze the joining ( ligation) of two molecules by forming a new chemical bond. This is typically via hydrolysis of a small pendant chemical group on one of the molecules, typically resulting i ...

enzyme An enzyme () is a protein that acts as a biological catalyst by accelerating chemical reactions. The molecules upon which enzymes may act are called substrate (chemistry), substrates, and the enzyme converts the substrates into different mol ...

s were tested: ''

Lig1 DNA ligase 1 also DNA ligase I, is an enzyme that in humans is encoded by the ''LIG1'' gene. DNA ligase 1 is the only known eukaryote, eukaryotic DNA ligase involved in both DNA replication and DNA repair, repair, making it the most studied of th ...

Lig3 DNA ligase 3 also DNA ligase III, is an enzyme that, in humans, is encoded by the ''LIG3'' gene. ''LIG3'' encodes ATP-dependent DNA ligases that seal interruptions in the phosphodiester backbone of duplex DNA. There are three families of AT ...

,'' and ''

Lig4 DNA ligase 4 also DNA ligase IV, is an enzyme that in humans is encoded by the ''LIG4'' gene. Function DNA ligase 4 is an ATP-dependent DNA ligase that joins double-strand breaks during the non-homologous end joining pathway of double-stran ...

''''.'' Using knockout models in the CH12F3 mouse

B-lymphocyte B cells, also known as B lymphocytes, are a type of the lymphocyte subtype. They function in the humoral immunity component of the adaptive immune system. B cells produce antibody molecules which may be either secreted or inserted into the plasm ...

cell line, research conducted in 2021 identified ''

'' as the main ligase for eccDNA generation in these cells.

Function

The exact function of eccDNA has been debated, but some studies have suggested that eccDNAs might contribute to

gene amplification Gene amplification refers to a number of natural and artificial processes by which the number of copies of a gene is increased "without a proportional increase in other genes". Artificial DNA amplification In research or diagnosis DNA amplificati ...

immune function The immune system is a network of biological systems that protects an organism from diseases. It detects and responds to a wide variety of pathogens, from viruses to bacteria, as well as cancer cells, parasitic worms, and also objects such as ...

, and

aging Ageing (or aging in American English) is the process of becoming Old age, older until death. The term refers mainly to humans, many other animals, and fungi; whereas for example, bacteria, perennial plants and some simple animals are potentiall ...

eccDNA function in immune system

According to research conducted in 2021, another function of eccDNAs is their role as possible

s. eccDNA significantly induces type I interferons (IFNα, IFNβ),

interleukin-6 Interleukin 6 (IL-6) is an interleukin that acts as both a pro-inflammatory cytokine and an anti-inflammatory myokine. In humans, it is encoded by the ''IL6'' gene. In addition, osteoblasts secrete IL-6 to stimulate osteoclast formation. Smoo ...

(IL-6), and

tumor necrosis factor Tumor necrosis factor (TNF), formerly known as TNF-α, is a chemical messenger produced by the immune system that induces inflammation. TNF is produced primarily by activated macrophages, and induces inflammation by binding to its receptors o ...

(TNF), even more so than linear DNA and other generally potent

cytokine Cytokines () are a broad and loose category of small proteins (~5–25 kDa) important in cell signaling. Cytokines are produced by a broad range of cells, including immune cells like macrophages, B cell, B lymphocytes, T cell, T lymphocytes ...

inducers at their highest concentration levels. Similar patterns are observed with

macrophage Macrophages (; abbreviated MPhi, φ, MΦ or MP) are a type of white blood cell of the innate immune system that engulf and digest pathogens, such as cancer cells, microbes, cellular debris and foreign substances, which do not have proteins that ...

s as the data showed that eccDNAs are very potent immunostimulants in activating both

bone marrow Bone marrow is a semi-solid biological tissue, tissue found within the Spongy bone, spongy (also known as cancellous) portions of bones. In birds and mammals, bone marrow is the primary site of new blood cell production (or haematopoiesis). It i ...

-derived

dendritic cell A dendritic cell (DC) is an antigen-presenting cell (also known as an ''accessory cell'') of the mammalian immune system. A DC's main function is to process antigen material and present it on the cell surface to the T cells of the immune system ...

s and bone marrow-derived

s. Additionally, experiments altered the eccDNA structure with one nick per eccDNA segment and subsequently treated with

s to generate linear versions of the eccDNA. In these experiments,

, an important marker for

immune system The immune system is a network of biological systems that protects an organism from diseases. It detects and responds to a wide variety of pathogens, from viruses to bacteria, as well as Tumor immunology, cancer cells, Parasitic worm, parasitic ...

activity, was shown to be much higher in the non-treated eccDNA compared to the linearized treatment, conferring that the circular structure of eccDNA rather than the genetic sequence itself gives the eccDNA its immune function.

eccDNA function in cancer

Some known functions of eccDNA include contributions to intercellular

genetic heterogeneity Genetic heterogeneity refers to different genetic causes for the same disease and can be classified into three types: allelic heterogeneity, locus heterogeneity, and phenotypic heterogeneity. Allelic heterogeneity occurs when different mutations wi ...

tumors A neoplasm () is a type of abnormal and excessive growth of tissue. The process that occurs to form or produce a neoplasm is called neoplasia. The growth of a neoplasm is uncoordinated with that of the normal surrounding tissue, and persists ...

, and more specifically the amplification of

oncogene An oncogene is a gene that has the potential to cause cancer. In tumor cells, these genes are often mutated, or expressed at high levels.

s and drug-resistant

s. This also supports that the

s on eccDNA are expressed. Overall, eccDNA has been linked to

and

drug resistance Drug resistance is the reduction in effectiveness of a medication such as an antimicrobial or an antineoplastic in treating a disease or condition. The term is used in the context of resistance that pathogens or cancers have "acquired", that is ...

, gene compensation, and for this reason it continues to be a significant topic of discussion.

Applications

Role in cancer

A subtype of eccDNA, such as ecDNA,

ribosomal DNA The ribosomal DNA (rDNA) consists of a group of ribosomal RNA encoding genes and related regulatory elements, and is widespread in similar configuration in all domains of life. The ribosomal DNA encodes the non-coding ribosomal RNA, integral struc ...

locus (

Extrachromosomal rDNA circle Extrachromosomal rDNA circles (aka ERC) are pieces of extrachromosomal circular DNA (eccDNA) derived from ribosomal DNA (rDNA). Initially found in baker's yeast, these self-replicating circles are suggested to contribute to their aging and found i ...

), and double minutes have been associated with

genomic instability Genome instability (also genetic instability or genomic instability) refers to a high frequency of mutations within the genome of a cellular lineage. These mutations can include changes in nucleic acid sequences, chromosomal rearrangements or aneu ...

Double minute Double minutes (DMs) are small fragments of extrachromosomal DNA, which have been observed in a large number of human tumors including breast, lung, ovary, colon, and most notably, neuroblastoma. They are a manifestation of gene amplification as a ...

ecDNAs are fragments of

extrachromosomal DNA Extrachromosomal DNA (abbreviated ecDNA) is any DNA that is found off the chromosomes, either inside or outside the nucleus of a cell. Most DNA in an individual genome is found in chromosomes contained in the nucleus. Multiple forms of extrachrom ...

, which were originally observed in a large number of human

tumor A neoplasm () is a type of abnormal and excessive growth of tissue. The process that occurs to form or produce a neoplasm is called neoplasia. The growth of a neoplasm is uncoordinated with that of the normal surrounding tissue, and persists ...

s including breast, lung, ovary, colon, and most notably,

neuroblastoma Neuroblastoma (NB) is a type of cancer that forms in certain types of nerve tissue. It most frequently starts from one of the adrenal glands but can also develop in the head, neck, chest, abdomen, or Vertebral column, spine. Symptoms may include ...

. They are a manifestation of

during the development of tumors, which give the cells selective advantages for growth and survival. Double minutes, like actual

chromosomes A chromosome is a package of DNA containing part or all of the genetic material of an organism. In most chromosomes, the very long thin DNA fibers are coated with nucleosome-forming packaging proteins; in eukaryotic cells, the most importa ...

, are composed of

and replicate in the

nucleus Nucleus (: nuclei) is a Latin word for the seed inside a fruit. It most often refers to: *Atomic nucleus, the very dense central region of an atom *Cell nucleus, a central organelle of a eukaryotic cell, containing most of the cell's DNA Nucleu ...

of the

cell Cell most often refers to: * Cell (biology), the functional basic unit of life * Cellphone, a phone connected to a cellular network * Clandestine cell, a penetration-resistant form of a secret or outlawed organization * Electrochemical cell, a de ...

during

cell division Cell division is the process by which a parent cell (biology), 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 eukar ...

. Unlike typical chromosomes, they are composed of circular fragments of

, up to only a few million

base pairs A base pair (bp) is a fundamental unit of double-stranded nucleic acids consisting of two nucleobases bound to each other by hydrogen bonds. They form the building blocks of the DNA double helix and contribute to the folded structure of both DNA ...

in size and contain no

centromere 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 fiber ...

telomere A telomere (; ) is a region of repetitive nucleotide sequences associated with specialized proteins at the ends of linear chromosomes (see #Sequences, Sequences). Telomeres are a widespread genetic feature most commonly found in eukaryotes. In ...

chromosomes (DMs), which present as paired chromatin bodies under

light microscopy Microscopy is the technical field of using microscopes to view subjects too small to be seen with the naked eye (objects that are not within the resolution range of the normal eye). There are three well-known branches of microscopy: optical, el ...

, have been shown to be a subset of ecDNA. Double minute chromosomes represent about 30% of the cancer-containing spectrum of ecDNA, including single bodies, and have been found to contain identical gene content as single bodies. The ecDNA notation encompasses all forms of the large gene-containing

found in

cancer cell Cancer cells are cells that divide continually, forming solid tumors or flooding the blood or lymph with abnormal cells. Cell division is a normal process used by the body for growth and repair. A parent cell divides to form two daughter cells, an ...

s. This type of ecDNA is commonly seen in

cells of various histologies, but virtually never in normal tissue. ecDNA are thought to be produced through

double-strand break DNA repair is a collection of processes by which a cell identifies and corrects damage to the DNA molecules that encode its genome. A weakened capacity for DNA repair is a risk factor for the development of cancer. DNA is constantly modified ...

s in chromosomes or over replication of DNA in an organism. The circular shape of ecDNA differs from the linear structure of

chromosomal A chromosome is a package of DNA containing part or all of the genetic material of an organism. In most chromosomes, the very long thin DNA fibers are coated with nucleosome-forming packaging proteins; in eukaryotic cells, the most importa ...

in meaningful ways that influence cancer

pathogenesis In pathology, pathogenesis is the process by which a disease or disorder develops. It can include factors which contribute not only to the onset of the disease or disorder, but also to its progression and maintenance. The word comes . Descript ...

Oncogene An oncogene is a gene that has the potential to cause cancer. In tumor cells, these genes are often mutated, or expressed at high levels.

s encoded on ecDNA have massive transcriptional output, ranking in the top 1% of

s in the entire

transcriptome The transcriptome is the set of all RNA transcripts, including coding and non-coding, in an individual or a population of cells. The term can also sometimes be used to refer to all RNAs, or just mRNA, depending on the particular experiment. The ...

. In contrast to bacterial

s or

, ecDNA are chromatinized, containing high levels of active

histone In biology, histones are highly basic proteins abundant in lysine and arginine residues that are found in eukaryotic cell nuclei and in most Archaeal phyla. They act as spools around which DNA winds to create structural units called nucleosomes ...

marks, but a paucity of repressive histone marks. The ecDNA

architecture lacks the higher-order compaction that is present on chromosomal DNA and is among the most accessible DNA in the entire cancer genome. From eccDNA, matrix attachment regions (MARs) were found to activate amplification of

oncogene An oncogene is a gene that has the potential to cause cancer. In tumor cells, these genes are often mutated, or expressed at high levels.

s. Transfection of these MARs into human embryonic kidney 293T cells resulted in an increase in

gene expression Gene expression is the process (including its Regulation of gene expression, regulation) by which information from a gene is used in the synthesis of a functional gene product that enables it to produce end products, proteins or non-coding RNA, ...

, suggesting that these eccDNA-derived MARs are involved in oncogene activation. eccDNA also appears to play a role in other cancers such as

breast cancer Breast cancer is a cancer that develops from breast tissue. Signs of breast cancer may include a Breast lump, lump in the breast, a change in breast shape, dimpling of the skin, Milk-rejection sign, milk rejection, fluid coming from the nipp ...

, where oncogenes in human epidermal growth factor receptor 2 (HER2)-positive breast cancer genes in eccDNA are amplified. This eccDNA has also shown the ability to acquire resistance to therapies for

receptor tyrosine kinase Receptor tyrosine kinases (RTKs) are the high-affinity cell surface receptors for many polypeptide growth factors, cytokines, and hormones. Of the 90 unique tyrosine kinase genes identified in the human genome, 58 encode receptor tyrosine kinas ...

s (RTKs), like HER26.

Role in aging

Yeast Yeasts are eukaryotic, single-celled microorganisms classified as members of the fungus kingdom (biology), kingdom. The first yeast originated hundreds of millions of years ago, and at least 1,500 species are currently recognized. They are est ...

are model organisms for studying

, and eccDNAs have been shown to accumulate in old cells and play a role in causing aging in yeast. Speculation continues on the generality of this concept in higher species, like

mammal A mammal () is a vertebrate animal of the Class (biology), class Mammalia (). Mammals are characterised by the presence of milk-producing mammary glands for feeding their young, a broad neocortex region of the brain, fur or hair, and three ...

References

ref 36 Altungöz; Yüksel (September 2023). "Gene amplifications and extrachromosomal circular DNAs: function and biogenesis". Molecular Biology Reports. 50 (9): 7693–7703. doi:10.1007/s11033-023-08649-1. PMID 37433908 – via Springer.