DNA repair
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Image:Brokechromo.jpg DNA repair is a process constantly operating in cells; it is essential to survival because it protects the genome from damage and harmful mutations. In human cells, both normal metabolic activities and environmental factors (such as UV rays) can cause DNA damage, resulting in as many as 500,000 individual molecular lesions per cell per day. These lesions cause structural damage to the DNA molecule, and can dramatically alter the cell's way of reading the information encoded in its genes. Consequently, the DNA repair process must be constantly operating, to correct rapidly any damage in the DNA structure.
As cells age, however, the rate of DNA repair decreases until it can no longer keep up with ongoing DNA damage. The cell then suffers one of three possible fates:
- an irreversible state of dormancy, known as senescence
- cell suicide, also known as apoptosis or programmed cell death
- carcinogenesis, or the formation of cancer.
Most cells in the body first become senescent. Then, after irreparable DNA damage, apoptosis occurs. In this case, apoptosis functions as a "last resort" mechanism to prevent a cell from becoming carcinogenic (able to form a tumor - see cancer) and endangering the organism.
When cells become senescent, alterations in biosynthesis and turnover cause them to function less efficiently, which inevitably causes disease. The DNA repair ability of a cell is vital to the integrity of its genome and thus to its normal functioning and that of the organism. Many genes that were initially shown to influence lifespan have turned out to be involved in DNA damage repair and protection.
Failure to correct molecular lesions in cells that form gametes leads to mutated progeny and might thus influence the rate of evolution.
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DNA damage
DNA damage, due to normal metabolic processes inside the cell, occurs at a rate of 50,000 to 500,000 molecular lesions per cell per day. However, many more sources of damage can drive this rate even higher. Whilst this constitutes only 0.0002% of the human genome's 3,000,000,000 (3 billion) bases, a single unrepaired lesion to a critical cancer-related gene (such as a tumor suppressor gene) can have catastrophic consequences for an individual.
Nuclear versus mitochondrial DNA damage
In human, and eukaryotic cells in general, DNA is found in two cellular locations - inside the nucleus and inside the mitochondria (mitochondrial genetics). Nuclear DNA (nDNA) exists in large-scale aggregate structures known as chromosomes, which are composed of DNA wound up around bead-like proteins called histones. Whenever a cell needs to express the genetic information encoded in its nDNA the required chromosomal region is unravelled, genes located therein are expressed, and then the region is condensed back to its quiescent conformation. Mitochondrial DNA (mtDNA) is located inside mitochondria organelles, exists in multiple copies, and is also tightly associated with a number of proteins to form a complex known as the nucleoid. Inside mitochondria, reactive oxygen species (ROS), or free radicals, byproducts of the constant production of adenosine triphosphate (ATP) via oxidative phosphorylation, create a highly oxidative environment that is known to damage mtDNA.
Sources of damage
DNA damage can be subdivided into two main types:
- endogenous damage such as attack by reactive oxygen radicals produced from normal metabolic byproducts (spontaneous mutation);
- exogenous damage caused by external agents such as
- ultraviolet [UV 200-300nm] radiation from the sun
- other radiation frequencies, including x-rays and gamma rays
- hydrolysis or thermal disruption
- certain plant toxins
- human-made mutagenic chemicals, such as hydrocarbons from cigarette smoke
- cancer chemotherapy and radiotherapy
Before cell division the replication of damaged DNA can lead to the incorporation of wrong bases opposite damaged ones. After the wrong bases are inherited by daughter cells these become mutated cells (cells that carry mutations), and there is no way back (except through the rare processes of back mutation and gene conversion).
Types of damage
Endogenous damage affects the primary rather than secondary structure of the double helix. Secondary structure does not occur in DNA, but in protein folding. DNA is wrapped tightly around various [Histone] proteins. Endogenous damage can be subdivided into four classes:
- oxidation of bases [e.g. 8-oxo-7,8-dihydroguanine (8-oxoG)] and generation of DNA strand interruptions from reactive oxygen species,
- alkylation of bases (usually methylation), such as formation of 7-methylguanine
- hydrolysis of bases, such as depurination and depyrimidination.
- mismatch of bases, due to DNA replication in which the wrong DNA base is stitched into place in a newly forming DNA strand.
DNA repair mechanisms
Cells cannot tolerate DNA damage that compromises the integrity and accessibility of essential information in the genome (but cells remain superficially functional when so-called "non-essential" genes are missing or damaged). Depending on the type of damage inflicted on the DNA's double helical structure, a variety of repair strategies has evolved to restore lost information. As templates for restoration cells use the unmodified complementary strand of the DNA or the sister chromosome. Without access to template information, DNA repair is error-prone (but this can be the standard pathway, e.g. most double strand-breaks in mammalian cells are repaired without template assistance; see below).
Damage to DNA alters the spatial configuration of the helix and such alterations can be detected by the cell. Once damage is localized, specific DNA repair molecules are summoned to, and bind at or near the site of damage, inducing other molecules to bind and form a complex that enables the actual repair to take place. The types of molecules involved and the mechanism of repair that is mobilized depend on:
- the type of DNA damage at stake
- whether the cell has entered into a state of senescence
- the phase of the cell cycle that the cell is in
Single strand damage
When only one of the two strands of a chromosome has a defect, the other strand can be used as a template to guide the correction of the damaged strand. In order to repair damage to one of the two helical domains of DNA, there are numerous mechanisms by which DNA repair can take place. These include:
- Direct reversal of damage by various mechanisms that specialize in reversing specific types of damage. Examples include methyl guanine methyl transferase (MGMT) which specifically removes methyl groups from guanine, and photolyase in bacteria, which breaks the chemical bond created by UV light between adjacent thymidine bases. No template strand is required for this form of repair.
- Excision repair mechanisms that remove the damaged nucleotide replacing it with an undamaged nucleotide complementary to the nucleotide in the undamaged DNA strand. These include:
- Base excision repair (BER), which repairs damage due to a single nucleotide caused by oxidation, alkylation, hydrolysis, or deamination;
- Nucleotide excision repair (NER), which repairs damage affecting 2−30 nucleotide-length strands. These include bulky, helix distorting damage, such as thymine dimerization caused by UV light as well as single-strand breaks. A specialized form of NER known as Transcription-Coupled Repair (TCR) deploys high-priority NER repair enzymes to genes that are being actively transcribed;
- Mismatch repair (MMR), which corrects errors of DNA replication and recombination that result in mispaired nucleotides following DNA replication.
Double strand breaks
A particularly hazardous type of DNA damage to dividing cells is a break to both strands in the double helix. Two mechanisms exist to repair this damage. They are generally known as Non-Homologous End-Joining and recombinational repair, template-assisted repair, or homologous recombination.
Recombinational repair requires the presence of an identical or nearly identical sequence to be used as a template for repair of the break. The enzymatic machinery responsible for this repair process is nearly identical to the machinery responsible for chromosomal crossover in germ cells during meiosis. The recombinational repair mechanism is predominantly used during the phases of the cell cycle when the DNA is replicating or has completed replicating its DNA. This allows a damaged chromosome to be repaired using the newly created sister chromatid as a template, i.e. an identical copy that is moreover orderly paired to the damaged region. Many genes in the human genome are present in multiple copies providing many possible sources of identical sequences. But recombinational repair that relies on these copies as templates for each other is problematic because it leads to chromosomal translocations and other types of chromosomal rearrangements. Non-Homologous End-Joining (NHEJ) rejoins the two ends of the break in absence of a template sequence. However there is often DNA sequence loss during this process and so this repair can be mutagenic. NHEJ can occur at all stages of the cell cycle but in mammalian cells is the main repair mechanism until DNA replication makes it possible for recombinational repair to use the sister chromatid as a template. Since the vast majority of the genome in humans and other multicellular organisms is made up of DNA that contains no genes, the so-called "junk DNA", mutagenic NHEJ is likely to be less harmful than template-assisted repair would be in presence of multiple template sequences, since in the latter case undesirable chromosomal rearrangements are generated. The enzymatic machinery used for NHEJ is also utilized in B-cells to rejoin breaks created by the RAG proteins during VDJ recombination a crucial step in the generation of antibody diversity by the immune system.
See also Carcinogenesis
DNA repair in disease and aging
Poor DNA repair induces pathology
As cells get older, the amount of DNA damage accumulates overtaking the rate of repair and resulting in a reduction of protein synthesis. As proteins in the cell are used for numerous vital functions, the cell becomes slowly impaired and eventually dies. When enough cells in an organ reach such a state, the organ itself will become compromised and the symptoms of disease begin to manifest. Experimental studies in animals, where genes associated with DNA repair were silenced, resulted in accelerated aging, early manifestation of age related diseases and increased susceptibility to cancer. In studies where the expression of certain DNA repair genes was increased resulted in extended lifespan and resistance to carcinogenic agents in cultured cells.
DNA repair rate is variable
If the rate of DNA damage exceeds the capacity of the cell to repair it, the accumulation of errors can overwhelm the cell and result in senescence, apoptosis or cancer. Inherited diseases associated with faulty DNA repair functioning result in premature aging (e.g. Werner's syndrome) and increased sensitivity to carcinogens (e.g Xeroderma Pigmentosum). Studies in animals, where DNA repair genes are prevented from functioning, show similar disease profiles.
On the other hand, organisms with enhanced DNA repair systems, such as Deinococcus radiodurans (also known as "Conan the bacterium", listed in the Guinness Book of World Records as "the world's toughest bacterium"), exhibit remarkable resistance to radioactivity, because their DNA repair enzymes are able to perform at unusually fast rates to keep up with radiation induced-damage, and because it carries 4–10 copies of the genome. In human studies, Japanese centenarians have been found to have a common mitochondrial genotype, which predisposes them to reduced DNA damage in their mitochondria.
Studies in smokers have found that, for people with a mutation that causes them to express less of the powerful DNA repair gene hOGG1, their vulnerability to lung and other smoking related cancers are increased. Single nucleotide polymorphisms (SNP) associated with this mutation can be clinically detected.
Hereditary DNA repair disorders
Defects in the NER mechanism are responsible for several genetic disorders, including:
- xeroderma pigmentosum: hypersensitivity to sunlight/UV, resulting in increased skin cancer incidence and premature aging
- Cockayne syndrome: hypersensitivity to UV and chemical agents
- trichothiodystrophy: sensitive skin, brittle hair and nails
Mental retardation often accompanies the latter two disorders, suggesting increased vulnerability of developmental neurons.
Other DNA repair disorders include:
- Werner's syndrome: premature aging and retarded growth
- Bloom's syndrome: sunlight hypersensitivity, high incidence of malignancies (especially leukemias).
- ataxia telangiectasia: sensitivity to ionizing radiation and some chemical agents
All of the above diseases are often called "segmental progerias" ("accelerated aging diseases") because their victims appear elderly and suffer from aging-related diseases at an abnormally young age.
Other diseases associated with reduced DNA repair function include Fanconi's anemia, hereditary breast cancer and hereditary colon cancer.
Chronic DNA repair disorders
Chronic disease can be associated with increased DNA damage. For example, smoking cigarettes causes oxidative damage to the DNA and other components of heart and lung cells, resulting in the formation of DNA adducts (molecules that disrupt DNA). DNA damage has now been shown to be a causative factor in diseases from atherosclerosis to Alzheimer's, where patients have a lesser capacity for DNA repair in their brain cells. Mitochondrial DNA damage has also been implicated in numerous disorders.
Longevity genes and DNA repair
Image:Dnadamage.jpg Certain genes are known to influence variation in lifespan within a population of organisms. Studies in model organisms such as yeast, worms, flies and mice have identified single genes, which when modified, can double lifespan (eg. a mutation in the age-1 gene of the nematode Caenorhabditis elegans). These genes are known to be associated specifically with cell functions other than DNA repair, but when the pathways that they influence are followed to their final destination, it was observed that they mediate one of three functions:
- increasing the rate of DNA repair,
- increasing the rate of antioxidant production, or
- decreasing the rate of oxidant production.
Therefore, the common pattern across most lifespan influencing genes is in their downstream effect of altering the rate of DNA damage.
Caloric restriction increases DNA repair
Caloric restriction (CR) has been shown to increase lifespan and decrease age related disease in all organisms where it has been studied, from single celled life such as yeast, to multicellular organisms such as worms, flies, mice and primates. The mechanism by which CR works is associated with a number of genes related to nutrient sensing which signal the cell to alter metabolic activity when there is a shortage of nutrients, particularly carbohydrates. When the cell senses a decrease in carbohydrate availability, activation of the lifespan influencing genes DAF-2, AGE-1 and SIR-2 (see accompanying illustration "Most lifespan influencing genes affect the rate of DNA damage") is triggered.
The reason why a shortage of nutrients will induce a cellular state of increased DNA repair and an increase in lifespan is suggested to be associated with an evolutionarily conserved mechanism of cellular hibernation. Essentially this permits a cell to maintain a dormant state until conditions that are more favorable are met. During this period, the cell must decrease its normal rate of metabolism and one of the ways it can accomplish this is by reducing genomic instability. Thus, the cellular rate of aging is mutable and can be influenced by environmental factors such as nutrient availability, which mediate their effect by altering the rate of DNA repair.
DNA repair and evolution
One form of DNA damage is alteration of a nucleotide (a mutation), altering the information carried in the DNA sequence. Because DNA mutation and recombination are the main means for evolution to occur, the rate of DNA repair influences the rate of evolution. With a very high level of DNA repair rate, the rate of mutation is reduced, resulting in corresponding reduction in the rate of evolution. Conversely, high mutation rates increase the rate of evolution.
DNA repair mechanisms are ancient
From a geologic chronological perspective, DNA repair mechanisms evolved during the Precambrian period not long after the life began to use nucleic acids as a means of encoding genetic information. During this period atmospheric oxygen began to increase steadily and then with the explosion of photosynthetic plants during the Cambrian period the levels approximated those that we have today. The toxicity of oxygen due to the formation of free radicals required the evolution of mechanisms able to reduce and repair such damage. Today, we can see highly conserved mechanisms of DNA repair that humans share with species as diverse as flies and worms.
Disease, death and evolution
DNA repair rates play a vital role at the cellular scale of (non-infectious) disease and aging, and at the population scale of evolution. Two important relationships have been established:
- DNA repair rate and mutation
- DNA repair rate and aging
As mutation is directly related to evolution, a new way of looking at the relationship between evolution and aging emerges. It is apparent that, while the mechanism of mutation provides the genome the plasticity to adapt, it is also responsible for destabilizing it, as well as for rendering it vulnerable to disease and aging. Are organisms subject to disease and aging primarily because mutation is the primary driver of evolution? This remains a contentious issue and numerous theories of aging have been offered.
Medicine & DNA repair modulation
A vast body of evidence correlates DNA damage to death and disease. As indicated by new overexpression studies, increasing the activity of some DNA repair enzymes could decrease the rate of aging and disease. This may result in the development of human interventions that can add many healthy and disease-free years to an aging population. Not all DNA repair enzymes are beneficial when overexpressed, however. Some DNA repair enzymes can introduce new mutations in healthy DNA. Reduced substrate specificity has been implicated in these errors.
Cancer treatment
Procedures such as chemotherapy and radiotherapy work by overwhelming the capacity of the cell to repair DNA damage and resulting in cell death. Cells that are most rapidly dividing such as cancer cells are preferentially affected. The side effect is that other non-cancerous but similarly rapidly dividing cells such as stem cells in the bone marrow are also affected. Modern cancer treatments attempt to localize the DNA damage to cells and tissues only associated with cancer.
Gene therapy
For therapeutic uses of DNA repair, the challenge is to discover which particular DNA repair enzymes exhibit the most precise specificity for damaged sites, so its overexpression will lead to enhanced DNA repair function. Once the appropriate repair factors have been identified, the next step is in selecting the appropriate way to deliver them into cells, to generate viable disease and aging treatments. The development of smart genes, which are able to alter the amount of protein they produce based on changing cellular conditions, stand to increase the efficacy of DNA repair augmentation treatments.
Gene repair
Unlike the multiple mechanisms of endogenous DNA repair, gene repair (or gene correction) refers to a form of gene therapy, which precisely targets and corrects chromosomal mutations responsible for a disorder. It does so by replacing the flawed DNA sequence with the desired sequence, using techniques such as oligonucleotide-directed mutagenesis. Genetic mutations requiring repair are normally inherited, but in some cases they can also be induced or acquired (such as in cancer).
References
- S. Tornaletti and G. P. Pfeiffer (1996) UV damage and repair mechanisms in mammalian cells. Bioessays 18, 221–228.
See also
External links
- A comprehensive list of Human DNA Repair Genes
- 3D structures of some DNA repair enzymes
- Human DNA repair diseases
- Damage-Based Theories of Aging Includes a description of the DNA damage theory of aging.
- DNA repair special interest group
- DNA Repair
- DNA Damage and DNA Repair
- Segmental Progeria
- Vegetables contain chemicals that boost DNA repair, Georgetown University Medical Center, February 9, 2006.
- Vegetables contain chemicals that boost DNA repair, British Journal of Cancer, February 13, 2006.
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