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106-437: H4K16ac is unusual in that it has both transcriptional activation AND repression activities. The loss of H4K20me3 along with a reduction of H4K16ac is a strong indicator of cancer. Proteins are typically acetylated on lysine residues and this reaction relies on acetyl-coenzyme A as the acetyl group donor. In histone acetylation and deacetylation , histone proteins are acetylated and deacetylated on lysine residues in

212-540: A DNA-binding domain that binds to a DNA sequence specific to the activator, and an activation domain that functions to increase gene transcription by interacting with other molecules. Activator DNA-binding domains come in a variety of conformations, including the helix-turn-helix , zinc finger , and leucine zipper among others. These DNA-binding domains are specific to a certain DNA sequence, allowing activators to turn on only certain genes. Activation domains also come in

318-456: A G[8,5-Me]T-modified plasmid in E. coli with specific DNA polymerase knockouts. Viability was very low in a strain lacking pol II, pol IV, and pol V, the three SOS-inducible DNA polymerases, indicating that translesion synthesis is conducted primarily by these specialized DNA polymerases. A bypass platform is provided to these polymerases by Proliferating cell nuclear antigen (PCNA). Under normal circumstances, PCNA bound to polymerases replicates

424-409: A barrier to all DNA-based processes that require recruitment of enzymes to their sites of action. To allow DNA repair, the chromatin must be remodeled . In eukaryotes, ATP dependent chromatin remodeling complexes and histone-modifying enzymes are two predominant factors employed to accomplish this remodeling process. Chromatin relaxation occurs rapidly at the site of a DNA damage. In one of

530-443: A cell leaves it with an important decision: undergo apoptosis and die, or survive at the cost of living with a modified genome. An increase in tolerance to damage can lead to an increased rate of survival that will allow a greater accumulation of mutations. Yeast Rev1 and human polymerase η are members of Y family translesion DNA polymerases present during global response to DNA damage and are responsible for enhanced mutagenesis during

636-455: A cell undergoes division (see Hayflick limit ). In contrast, quiescence is a reversible state of cellular dormancy that is unrelated to genome damage (see cell cycle ). Senescence in cells may serve as a functional alternative to apoptosis in cases where the physical presence of a cell for spatial reasons is required by the organism, which serves as a "last resort" mechanism to prevent a cell with damaged DNA from replicating inappropriately in

742-446: A cell's ability to carry out its function and appreciably increase the likelihood of tumor formation and contribute to tumor heterogeneity . The vast majority of DNA damage affects the primary structure of the double helix; that is, the bases themselves are chemically modified. These modifications can in turn disrupt the molecules' regular helical structure by introducing non-native chemical bonds or bulky adducts that do not fit in

848-487: A cell. Processes such as phosphorylation , acetylation , and ubiquitination , among others, have been seen to regulate the activity of activators. Depending on the chemical group being added, as well as the nature of the activator itself, post-translational modifications can either increase or decrease the activity of an activator. For example, acetylation has been seen to increase the activity of some activators through mechanisms such as increasing DNA-binding affinity. On

954-470: A certain molecule binds to this site, essentially turning the activator on. Post-translational modifications to activators can also regulate activity, increasing or decreasing activity depending on the type of modification and activator being modified. In some cells, usually eukaryotes, multiple activators can bind to the binding-site; these activators tend to bind cooperatively and interact synergistically. Activator proteins consist of two main domains :

1060-599: A common global response. The probable explanation for this difference between yeast and human cells may be in the heterogeneity of mammalian cells. In an animal different types of cells are distributed among different organs that have evolved different sensitivities to DNA damage. In general global response to DNA damage involves expression of multiple genes responsible for postreplication repair , homologous recombination, nucleotide excision repair, DNA damage checkpoint , global transcriptional activation, genes controlling mRNA decay, and many others. A large amount of damage to

1166-690: A conformational change that allows CAP to bind to a DNA site located adjacent to the lac promoter. CAP then makes a direct protein–protein interaction with RNA polymerase that recruits RNA polymerase to the lac promoter. DNA repair DNA repair is a collection of processes by which a cell identifies and corrects damage to the DNA molecules that encode its genome . In human cells, both normal metabolic activities and environmental factors such as radiation can cause DNA damage, resulting in tens of thousands of individual molecular lesions per cell per day. Many of these lesions cause structural damage to

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1272-478: A gene can be prevented, and thus translation into a protein will also be blocked. Replication may also be blocked or the cell may die. In contrast to DNA damage, a mutation is a change in the base sequence of the DNA. A mutation cannot be recognized by enzymes once the base change is present in both DNA strands, and thus a mutation cannot be repaired. At the cellular level, mutations can cause alterations in protein function and regulation. Mutations are replicated when

1378-403: A genome independently of the underlying genome sequence. This independence from the DNA sequence enforces the epigenetic nature of histone modifications. Chromatin states are also useful in identifying regulatory elements that have no defined sequence, such as enhancers . This additional level of annotation allows for a deeper understanding of cell specific gene regulation. Secondly, it can block

1484-417: A global response to DNA damage in eukaryotes. Experimental animals with genetic deficiencies in DNA repair often show decreased life span and increased cancer incidence. For example, mice deficient in the dominant NHEJ pathway and in telomere maintenance mechanisms get lymphoma and infections more often, and, as a consequence, have shorter lifespans than wild-type mice. In similar manner, mice deficient in

1590-547: A heterodimeric complex with DDB1 . This complex further complexes with the ubiquitin ligase protein CUL4A and with PARP1 . This larger complex rapidly associates with UV-induced damage within chromatin, with half-maximum association completed in 40 seconds. The PARP1 protein, attached to both DDB1 and DDB2, then PARylates (creates a poly-ADP ribose chain) on DDB2 that attracts the DNA remodeling protein ALC1 . Action of ALC1 relaxes

1696-626: A highly complex form of DNA damage as clustered damage. It consists of different types of DNA lesions in various locations of the DNA helix. Some of these closely located lesions can probably convert to DSB by exposure to high temperatures. But the exact nature of these lesions and their interactions is not yet known Translesion synthesis (TLS) is a DNA damage tolerance process that allows the DNA replication machinery to replicate past DNA lesions such as thymine dimers or AP sites . It involves switching out regular DNA polymerases for specialized translesion polymerases (i.e. DNA polymerase IV or V, from

1802-514: A key repair and transcription protein that unwinds DNA helices have premature onset of aging-related diseases and consequent shortening of lifespan. However, not every DNA repair deficiency creates exactly the predicted effects; mice deficient in the NER pathway exhibited shortened life span without correspondingly higher rates of mutation. The maximum life spans of mice , naked mole-rats and humans are respectively ~3, ~30 and ~129 years. Of these,

1908-639: A last resort. Once the DNA damage is repaired or bypassed using polymerases or through recombination, the amount of single-stranded DNA in cells is decreased, lowering the amounts of RecA filaments decreases cleavage activity of LexA homodimer, which then binds to the SOS boxes near promoters and restores normal gene expression. Eukaryotic cells exposed to DNA damaging agents also activate important defensive pathways by inducing multiple proteins involved in DNA repair, cell cycle checkpoint control, protein trafficking and degradation. Such genome wide transcriptional response

2014-505: A mutation. Three mechanisms exist to repair double-strand breaks (DSBs): non-homologous end joining (NHEJ), microhomology-mediated end joining (MMEJ), and homologous recombination (HR): In an in vitro system, MMEJ occurred in mammalian cells at the levels of 10–20% of HR when both HR and NHEJ mechanisms were also available. The extremophile Deinococcus radiodurans has a remarkable ability to survive DNA damage from ionizing radiation and other sources. At least two copies of

2120-445: A population of cells composing a tissue with replicating cells, mutant cells will tend to be lost. However, infrequent mutations that provide a survival advantage will tend to clonally expand at the expense of neighboring cells in the tissue. This advantage to the cell is disadvantageous to the whole organism because such mutant cells can give rise to cancer. Thus, DNA damage in frequently dividing cells, because it gives rise to mutations,

2226-415: A second, with half maximum accumulation within 1.6 seconds after the damage occurs. PARP1 synthesizes polymeric adenosine diphosphate ribose (poly (ADP-ribose) or PAR) chains on itself. Next the chromatin remodeler ALC1 quickly attaches to the product of PARP1 action, a poly-ADP ribose chain, and ALC1 completes arrival at the DNA damage within 10 seconds of the occurrence of the damage. About half of

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2332-408: A specialized polymerase is needed to extend it; Pol ζ . Pol ζ is unique in that it can extend terminal mismatches, whereas more processive polymerases cannot. So when a lesion is encountered, the replication fork will stall, PCNA will switch from a processive polymerase to a TLS polymerase such as Pol ι to fix the lesion, then PCNA may switch to Pol ζ to extend the mismatch, and last PCNA will switch to

2438-412: A targeted protein and immunoprecipitated . It results in good optimization and is used in vivo to reveal DNA-protein binding occurring in cells. ChIP-Seq can be used to identify and quantify various DNA fragments for different histone modifications along a genomic region. 2. Micrococcal Nuclease sequencing ( MNase-seq ) is used to investigate regions that are bound by well positioned nucleosomes. Use of

2544-447: A variety of repair strategies have evolved to restore lost information. If possible, cells use the unmodified complementary strand of the DNA or the sister chromatid as a template to recover the original information. Without access to a template, cells use an error-prone recovery mechanism known as translesion synthesis as a last resort. Damage to DNA alters the spatial configuration of the helix, and such alterations can be detected by

2650-427: A variety of types that are categorized based on the domain's amino acid sequence, including alanine -rich, glutamine -rich, and acidic domains. These domains are not as specific, and tend to interact with a variety of target molecules. Activators can also have allosteric sites that are responsible for turning the activators themselves on and off. Within the grooves of the DNA double helix, functional groups of

2756-576: Is p53 , as it is required for inducing apoptosis following DNA damage. The cyclin-dependent kinase inhibitor p21 is induced by both p53-dependent and p53-independent mechanisms and can arrest the cell cycle at the G1/S and G2/M checkpoints by deactivating cyclin / cyclin-dependent kinase complexes. The SOS response is the changes in gene expression in Escherichia coli and other bacteria in response to extensive DNA damage. The prokaryotic SOS system

2862-611: Is a pair of large protein kinases belonging to the first group of PI3K-like protein kinases-the ATM ( Ataxia telangiectasia mutated ) and ATR (Ataxia- and Rad-related) kinases, whose sequence and functions have been well conserved in evolution. All DNA damage response requires either ATM or ATR because they have the ability to bind to the chromosomes at the site of DNA damage, together with accessory proteins that are platforms on which DNA damage response components and DNA repair complexes can be assembled. An important downstream target of ATM and ATR

2968-419: Is a prominent cause of cancer. In contrast, DNA damage in infrequently-dividing cells is likely a prominent cause of aging. Cells cannot function if DNA damage corrupts the integrity and accessibility of essential information in the genome (but cells remain superficially functional when non-essential genes are missing or damaged). Depending on the type of damage inflicted on the DNA's double helical structure,

3074-420: Is a protein ( transcription factor ) that increases transcription of a gene or set of genes. Activators are considered to have positive control over gene expression, as they function to promote gene transcription and, in some cases, are required for the transcription of genes to occur. Most activators are DNA-binding proteins that bind to enhancers or promoter-proximal elements . The DNA site bound by

3180-483: Is a significant post-translational regulatory mechanism These regulatory mechanisms are analogous to phosphorylation and dephosphorylation by the action of kinases and phosphatases . Not only can the acetylation state of a protein modify its activity but there has been recent suggestion that this post-translational modification may also crosstalk with phosphorylation , methylation , ubiquitination , sumoylation, and others for dynamic control of cellular signaling. In

3286-414: Is a special problem in non-dividing or slowly-dividing cells, where unrepaired damage will tend to accumulate over time. On the other hand, in rapidly dividing cells, unrepaired DNA damage that does not kill the cell by blocking replication will tend to cause replication errors and thus mutation. The great majority of mutations that are not neutral in their effect are deleterious to a cell's survival. Thus, in

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3392-420: Is about two million base pairs at the site of a DNA double-strand break. γH2AX does not, itself, cause chromatin decondensation, but within 30 seconds of irradiation, RNF8 protein can be detected in association with γH2AX. RNF8 mediates extensive chromatin decondensation, through its subsequent interaction with CHD4 , a component of the nucleosome remodeling and deacetylase complex NuRD . DDB2 occurs in

3498-414: Is always highly conserved and one of the strongest short signals in the genome. The high information content of SOS boxes permits differential binding of LexA to different promoters and allows for timing of the SOS response. The lesion repair genes are induced at the beginning of SOS response. The error-prone translesion polymerases, for example, UmuCD'2 (also called DNA polymerase V), are induced later on as

3604-446: Is certain methylation of the bases cytosine and adenine. When only one of the two strands of a double helix 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 paired molecules of DNA, there exist a number of excision repair mechanisms that remove the damaged nucleotide and replace it with an undamaged nucleotide complementary to that found in

3710-407: Is controlled by the ability of the activator to bind to its regulatory site along the DNA. The DNA-binding domain of the activator has an active form and an inactive form, which are controlled by the binding of molecules known as allosteric effectors to the allosteric site of the activator. Activators in their inactive form are not bound to any allosteric effectors. When inactive, the activator

3816-484: Is controlled by two master kinases , ATM and ATR . ATM responds to DNA double-strand breaks and disruptions in chromatin structure, whereas ATR primarily responds to stalled replication forks . These kinases phosphorylate downstream targets in a signal transduction cascade, eventually leading to cell cycle arrest. A class of checkpoint mediator proteins including BRCA1 , MDC1 , and 53BP1 has also been identified. These proteins seem to be required for transmitting

3922-557: Is damaged. This is followed by phosphorylation of the cell cycle checkpoint protein Chk1 , initiating its function, about 10 minutes after DNA is damaged. After DNA damage, cell cycle checkpoints are activated. Checkpoint activation pauses the cell cycle and gives the cell time to repair the damage before continuing to divide. DNA damage checkpoints occur at the G1 / S and G2 / M boundaries. An intra- S checkpoint also exists. Checkpoint activation

4028-485: Is done through various mechanisms, such as recruiting transcription machinery to the promoter and triggering RNA polymerase to continue into elongation. Activator-controlled genes require the binding of activators to regulatory sites in order to recruit the necessary transcription machinery to the promoter region. Activator interactions with RNA polymerase are mostly direct in prokaryotes and indirect in eukaryotes. In prokaryotes, activators tend to make contact with

4134-481: Is known to add the first adenine across the T^T photodimer using Watson-Crick base pairing and the second adenine will be added in its syn conformation using Hoogsteen base pairing . From a cellular perspective, risking the introduction of point mutations during translesion synthesis may be preferable to resorting to more drastic mechanisms of DNA repair, which may cause gross chromosomal aberrations or cell death. In short,

4240-452: 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. A critical enzyme in counteracting

4346-435: Is much higher than the additive effects of the activators if they were working individually. The breakdown of maltose in Escherichia coli is controlled by gene activation. The genes that code for the enzymes responsible for maltose catabolism can only be transcribed in the presence of an activator. The activator that controls transcription of the maltose enzymes is "off" in the absence of maltose. In its inactive form,

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4452-477: Is not clear how the loss of a repressive and an activating mark is an indicator of cancer. It is not clear exactly how but this reduction happens at repetitive sequences along with general reduced DNA methylation. The histone mark acetylation can be detected in a variety of ways: 1. Chromatin Immunoprecipitation Sequencing ( ChIP-sequencing ) measures the amount of DNA enrichment once bound to

4558-440: Is obligately dependent on energy absorbed from blue/UV light (300–500 nm wavelength ) to promote catalysis. Photolyase, an old enzyme present in bacteria , fungi , and most animals no longer functions in humans, who instead use nucleotide excision repair to repair damage from UV irradiation. Another type of damage, methylation of guanine bases, is directly reversed by the enzyme methyl guanine methyl transferase (MGMT),

4664-482: Is regulated by two key proteins: LexA and RecA . The LexA homodimer is a transcriptional repressor that binds to operator sequences commonly referred to as SOS boxes. In Escherichia coli it is known that LexA regulates transcription of approximately 48 genes including the lexA and recA genes. The SOS response is known to be widespread in the Bacteria domain, but it is mostly absent in some bacterial phyla, like

4770-516: Is unable to bind to its specific regulatory sequence in the DNA, and thus has no regulatory effect on the transcription of genes. When an allosteric effector binds to the allosteric site of an activator, a conformational change in the DNA-binding domain occurs, which allows the protein to bind to the DNA and increase gene transcription. Some activators are able to undergo post-translational modifications that have an effect on their activity within

4876-473: Is very complex and tightly regulated, thus allowing coordinated global response to damage. Exposure of yeast Saccharomyces cerevisiae to DNA damaging agents results in overlapping but distinct transcriptional profiles. Similarities to environmental shock response indicates that a general global stress response pathway exist at the level of transcriptional activation. In contrast, different human cell types respond to damage differently indicating an absence of

4982-631: The Spirochetes . The most common cellular signals activating the SOS response are regions of single-stranded DNA (ssDNA), arising from stalled replication forks or double-strand breaks, which are processed by DNA helicase to separate the two DNA strands. In the initiation step, RecA protein binds to ssDNA in an ATP hydrolysis driven reaction creating RecA–ssDNA filaments. RecA–ssDNA filaments activate LexA auto protease activity, which ultimately leads to cleavage of LexA dimer and subsequent LexA degradation. The loss of LexA repressor induces transcription of

5088-462: The cell cycle and is condensed into aggregate structures known as chromosomes during cell division . In either state the DNA is highly compacted and wound up around bead-like proteins called histones . Whenever a cell needs to express the genetic information encoded in its n-DNA the required chromosomal region is unraveled, genes located therein are expressed, and then the region is condensed back to its resting conformation. Mitochondrial DNA (mtDNA)

5194-461: The gene dosage of the gene SIR-2, which regulates DNA packaging in the nematode worm Caenorhabditis elegans , can significantly extend lifespan. The mammalian homolog of SIR-2 is known to induce downstream DNA repair factors involved in NHEJ, an activity that is especially promoted under conditions of caloric restriction. Caloric restriction has been closely linked to the rate of base excision repair in

5300-471: The replication forks , are among known stimulation signals for a global response to DNA damage. The global response to damage is an act directed toward the cells' own preservation and triggers multiple pathways of macromolecular repair, lesion bypass, tolerance, or apoptosis . The common features of global response are induction of multiple genes , cell cycle arrest, and inhibition of cell division . The packaging of eukaryotic DNA into chromatin presents

5406-425: The toxicity of these species is superoxide dismutase , which is present in both the mitochondria and cytoplasm of eukaryotic cells. Senescence, an irreversible process in which the cell no longer divides , is a protective response to the shortening of the chromosome ends, called telomeres . The telomeres are long regions of repetitive noncoding DNA that cap chromosomes and undergo partial degradation each time

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5512-650: The two-hit hypothesis . The rate of DNA repair depends on various factors, including the cell type, the age of the cell, and the extracellular environment. A cell that has accumulated a large amount of DNA damage or can no longer effectively repair its DNA may enter one of three possible states: The DNA repair ability of a cell is vital to the integrity of its genome and thus to the normal functionality of that organism. Many genes that were initially shown to influence life span have turned out to be involved in DNA damage repair and protection. The 2015 Nobel Prize in Chemistry

5618-563: The DNA molecule and can alter or eliminate the cell's ability to transcribe the gene that the affected DNA encodes. Other lesions induce potentially harmful mutations in the cell's genome, which affect the survival of its daughter cells after it undergoes mitosis . As a consequence, the DNA repair process is constantly active as it responds to damage in the DNA structure. When normal repair processes fail, and when cellular apoptosis does not occur, irreparable DNA damage may occur. This can eventually lead to malignant tumors, or cancer as per

5724-644: The DNA so any blocks are removed. Activators may also promote the recruitment of elongation factors, which are necessary for the RNA polymerase to continue transcription. There are different ways in which the activity of activators themselves can be regulated, in order to ensure that activators are stimulating gene transcription at appropriate times and levels. Activator activity can increase or decrease in response to environmental stimuli or other intracellular signals. Activators often must be "turned on" before they can promote gene transcription. The activity of activators

5830-400: The DNA, such as single- and double-strand breaks, 8-hydroxydeoxyguanosine residues, and polycyclic aromatic hydrocarbon adducts. DNA damage can be recognized by enzymes, and thus can be correctly repaired if redundant information, such as the undamaged sequence in the complementary DNA strand or in a homologous chromosome, is available for copying. If a cell retains DNA damage, transcription of

5936-502: The DNA. At a site of lesion , PCNA is ubiquitinated, or modified, by the RAD6/ RAD18 proteins to provide a platform for the specialized polymerases to bypass the lesion and resume DNA replication. After translesion synthesis, extension is required. This extension can be carried out by a replicative polymerase if the TLS is error-free, as in the case of Pol η, yet if TLS results in a mismatch,

6042-477: The N-terminal tail as part of gene regulation . Typically, these reactions are catalyzed by enzymes with histone acetyltransferase (HAT) or histone deacetylase (HDAC) activity, although HATs and HDACs can modify the acetylation status of non-histone proteins as well. The regulation of transcription factors, effector proteins, molecular chaperones , and cytoskeletal proteins by acetylation and deacetylation

6148-522: The NoRC complex silences rDNA with HATs and DNMTs. There is also a reduction in the levels of H3K56ac during aging and an increase in the levels of H4K16ac. Increased H4K16ac in old yeast cells is associated with the decline in levels of the HDAC Sir2, which can increase the life span when overexpressed. The loss of the repressive H4K20me3 mark defines cancer along with a reduction of activating H4K16ac mark. It

6254-400: The RNA polymerase directly in order to help bind it to the promoter. In eukaryotes, activators mostly interact with other proteins, and these proteins will then be the ones to interact with the RNA polymerase. In prokaryotes, genes controlled by activators have promoters that are unable to strongly bind to RNA polymerase by themselves. Thus, activator proteins help to promote the binding of

6360-700: The RNA polymerase to the promoter. This is done through various mechanisms. Activators may bend the DNA in order to better expose the promoter so the RNA polymerase can bind more effectively. Activators may make direct contact with the RNA polymerase and secure it to the promoter. In eukaryotes, activators have a variety of different target molecules that they can recruit in order to promote gene transcription. They can recruit other transcription factors and cofactors that are needed in transcription initiation. Activators can recruit molecules known as coactivators . These coactivator molecules can then perform functions necessary for beginning transcription in place of

6466-432: The SOS genes and allows for further signal induction, inhibition of cell division and an increase in levels of proteins responsible for damage processing. In Escherichia coli , SOS boxes are 20-nucleotide long sequences near promoters with palindromic structure and a high degree of sequence conservation. In other classes and phyla, the sequence of SOS boxes varies considerably, with different length and composition, but it

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6572-691: The Y Polymerase family), often with larger active sites that can facilitate the insertion of bases opposite damaged nucleotides. The polymerase switching is thought to be mediated by, among other factors, the post-translational modification of the replication processivity factor PCNA . Translesion synthesis polymerases often have low fidelity (high propensity to insert wrong bases) on undamaged templates relative to regular polymerases. However, many are extremely efficient at inserting correct bases opposite specific types of damage. For example, Pol η mediates error-free bypass of lesions induced by UV irradiation , whereas Pol ι introduces mutations at these sites. Pol η

6678-493: The absence of pro-growth cellular signaling . Unregulated cell division can lead to the formation of a tumor (see cancer ), which is potentially lethal to an organism. Therefore, the induction of senescence and apoptosis is considered to be part of a strategy of protection against cancer. It is important to distinguish between DNA damage and mutation, the two major types of error in DNA. DNA damage and mutation are fundamentally different. Damage results in physical abnormalities in

6784-503: The activator is referred to as an "activator-binding site". The part of the activator that makes protein–protein interactions with the general transcription machinery is referred to as an "activating region" or "activation domain". Most activators function by binding sequence-specifically to a regulatory DNA site located near a promoter and making protein–protein interactions with the general transcription machinery ( RNA polymerase and general transcription factors ), thereby facilitating

6890-464: The activator is unable to bind to DNA and promote transcription of the maltose genes. When maltose is present in the cell, it binds to the allosteric site of the activator protein, causing a conformational change in the DNA-binding domain of the activator. This conformational change "turns on" the activator by allowing it to bind to its specific regulatory DNA sequence. Binding of the activator to its regulatory site promotes RNA polymerase binding to

6996-448: The activators themselves, such as chromatin modifications. DNA is much more condensed in eukaryotes; thus, activators tend to recruit proteins that are able to restructure the chromatin so the promoter is more easily accessible by the transcription machinery. Some proteins will rearrange the layout of nucleosomes along the DNA in order to expose the promoter site ( ATP-dependent chromatin remodeling complexes ). Other proteins affect

7102-463: The bacterial equivalent of which is called ogt . This is an expensive process because each MGMT molecule can be used only once; that is, the reaction is stoichiometric rather than catalytic . A generalized response to methylating agents in bacteria is known as the adaptive response and confers a level of resistance to alkylating agents upon sustained exposure by upregulation of alkylation repair enzymes. The third type of DNA damage reversed by cells

7208-441: The base pairs are exposed. The sequence of the DNA thus creates a unique pattern of surface features, including areas of possible hydrogen bonding , ionic bonding , as well as hydrophobic interactions . Activators also have unique sequences of amino acids with side chains that are able to interact with the functional groups in DNA. Thus, the pattern of amino acid side chains making up an activator protein will be complementary to

7314-494: The beginning of transcription. The RNA polymerase can sometimes pause shortly after beginning transcription, and activators are required to release RNA polymerase from this “stalled” state. Multiple mechanisms exist for releasing these "stalled" RNA polymerases. Activators may act simply as a signal to trigger the continued movement of the RNA polymerase. If the DNA is too condensed to allow RNA polymerase to continue transcription, activators may recruit proteins that can restructure

7420-412: The binding between histones and DNA via post-translational histone modifications , allowing the DNA tightly wrapped into nucleosomes to loosen. All of these recruited molecules work together in order to ultimately recruit the RNA polymerase to the promoter site. Activators can promote gene transcription by signaling the RNA polymerase to move beyond the promoter and proceed along the DNA, initiating

7526-538: The binding location of proteins in the genome. Use of ChIP-sequencing revealed regions in the genome characterised by different banding. Different developmental stages were profiled in Drosophila as well, an emphasis was placed on histone modification relevance. A look in to the data obtained led to the definition of chromatin states based on histone modifications. The human genome was annotated with chromatin states. These annotated states can be used as new ways to annotate

7632-486: The binding of the general transcription machinery to the promoter. Other activators help promote gene transcription by triggering RNA polymerase to release from the promoter and proceed along the DNA. At times, RNA polymerase can pause shortly after leaving the promoter; activators also function to allow these "stalled" RNA polymerases to continue transcription. The activity of activators can be regulated. Some activators have an allosteric site and can only function when

7738-415: The binding-site, meaning that the binding of one activator increases the affinity of the site to bind another activator (or in some cases another transcriptional regulator) thus making it easier for multiple activators to bind at the site. In these cases, the activators interact with each other synergistically , meaning that the rate of transcription that is achieved from multiple activators working together

7844-429: The capacity of the cell to repair it, the accumulation of errors can overwhelm the cell and result in early senescence, apoptosis, or cancer. Inherited diseases associated with faulty DNA repair functioning result in premature aging, increased sensitivity to carcinogens and correspondingly increased cancer risk (see below ). On the other hand, organisms with enhanced DNA repair systems, such as Deinococcus radiodurans ,

7950-492: The cell replicates. In a population of cells, mutant cells will increase or decrease in frequency according to the effects of the mutation on the ability of the cell to survive and reproduce. Although distinctly different from each other, DNA damage and mutation are related because DNA damage often causes errors of DNA synthesis during replication or repair; these errors are a major source of mutation. Given these properties of DNA damage and mutation, it can be seen that DNA damage

8056-404: The cell. Once damage is localized, specific DNA repair molecules 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. Cells are known to eliminate three types of damage to their DNA by chemically reversing it. These mechanisms do not require a template, since the types of damage they counteract can occur in only one of

8162-687: The checkpoint activation signal to downstream proteins. DNA damage checkpoint is a signal transduction pathway that blocks cell cycle progression in G1, G2 and metaphase and slows down the rate of S phase progression when DNA is damaged. It leads to a pause in cell cycle allowing the cell time to repair the damage before continuing to divide. Checkpoint Proteins can be separated into four groups: phosphatidylinositol 3-kinase (PI3K)-like protein kinase , proliferating cell nuclear antigen (PCNA)-like group, two serine/threonine(S/T) kinases and their adaptors. Central to all DNA damage induced checkpoints responses

8268-441: The chromatin at the site of UV damage to DNA. This relaxation allows other proteins in the nucleotide excision repair pathway to enter the chromatin and repair UV-induced cyclobutane pyrimidine dimer damages. After rapid chromatin remodeling , cell cycle checkpoints are activated to allow DNA repair to occur before the cell cycle progresses. First, two kinases , ATM and ATR are activated within 5 or 6 minutes after DNA

8374-560: The course of changing the DNA's state of supercoiling , which is especially common in regions near an open replication fork. Such breaks are not considered DNA damage because they are a natural intermediate in the topoisomerase biochemical mechanism and are immediately repaired by the enzymes that created them. Another type of DNA double-strand breaks originates from the DNA heat-sensitive or heat-labile sites. These DNA sites are not initial DSBs. However, they convert to DSB after treating with elevated temperature. Ionizing irradiation can induces

8480-487: The earliest steps, the stress-activated protein kinase, c-Jun N-terminal kinase (JNK) , phosphorylates SIRT6 on serine 10 in response to double-strand breaks or other DNA damage. This post-translational modification facilitates the mobilization of SIRT6 to DNA damage sites, and is required for efficient recruitment of poly (ADP-ribose) polymerase 1 (PARP1) to DNA break sites and for efficient repair of DSBs. PARP1 protein starts to appear at DNA damage sites in less than

8586-451: The field of epigenetics , histone acetylation (and deacetylation ) have been shown to be important mechanisms in the regulation of gene transcription. Histones, however, are not the only proteins regulated by posttranslational acetylation. H4K16ac indicates acetylation of lysine 16 on histone H4 protein subunit: The genomic DNA of eukaryotic cells is wrapped around special protein molecules known as histones . The complexes formed by

8692-521: The formation of a compact higher-order chromatin structure. Hypoacetylation of H4K16 appears to cause delayed recruitment of DNA repair proteins to sites of DNA damage in a mouse model of the premature aging, such as Hutchinson–Gilford progeria syndrome . H4K16Ac also has roles in transcriptional activation and the maintenance of euchromatin . H4K16ac is unusual in that it is associated with both transcriptional activation and repression. The bromodomain of TIP5, part of NoRC, binds to H4K16ac and then

8798-412: The four bases. Such direct reversal mechanisms are specific to the type of damage incurred and do not involve breakage of the phosphodiester backbone. The formation of pyrimidine dimers upon irradiation with UV light results in an abnormal covalent bond between adjacent pyrimidine bases. The photoreactivation process directly reverses this damage by the action of the enzyme photolyase , whose activation

8904-479: The function of chromatin remodelers. Thirdly, it neutralizes the positive charge on lysines. Acetylation of histone H4 on lysine 16 (H4K16Ac) is especially important for chromatin structure and function in a variety of eukaryotes and is catalyzed by specific histone lysine acetyltransferases (HATs). H4K16 is particularly interesting because this is the only acetylatable site of the H4 N-terminal tail, and can influence

9010-432: The genome, with random DNA breaks, can form DNA fragments through annealing . Partially overlapping fragments are then used for synthesis of homologous regions through a moving D-loop that can continue extension until complementary partner strands are found. In the final step, there is crossover by means of RecA -dependent homologous recombination . Topoisomerases introduce both single- and double-strand breaks in

9116-537: The histones in a particular region. The current understanding and interpretation of histones comes from two large scale projects: ENCODE and the Epigenomic roadmap. The purpose of the epigenomic study was to investigate epigenetic changes across the entire genome. This led to chromatin states which define genomic regions by grouping the interactions of different proteins and/or histone modifications together. Chromatin states were investigated in Drosophila cells by looking at

9222-550: The incorporation of wrong bases opposite damaged ones. Daughter cells that inherit these wrong bases carry mutations from which the original DNA sequence is unrecoverable (except in the rare case of a back mutation , for example, through gene conversion ). There are several types of damage to DNA due to endogenous cellular processes: Damage caused by exogenous agents comes in many forms. Some examples are: UV damage, alkylation/methylation, X-ray damage and oxidative damage are examples of induced damage. Spontaneous damage can include

9328-469: The looping of the DNA are known as chromatin . The basic structural unit of chromatin is the nucleosome : this consists of the core octamer of histones (H2A, H2B, H3 and H4) as well as a linker histone and about 180 base pairs of DNA. These core histones are rich in lysine and arginine residues. The carboxyl (C) terminal end of these histones contribute to histone-histone interactions, as well as histone-DNA interactions. The amino (N) terminal charged tails are

9434-455: The loss of a base, deamination, sugar ring puckering and tautomeric shift. Constitutive (spontaneous) DNA damage caused by endogenous oxidants can be detected as a low level of histone H2AX phosphorylation in untreated cells. In human cells, and eukaryotic cells in general, DNA is found in two cellular locations – inside the nucleus and inside the mitochondria . Nuclear DNA (n-DNA) exists as chromatin during non-replicative stages of

9540-744: The maximum chromatin relaxation, presumably due to action of ALC1, occurs by 10 seconds. This then allows recruitment of the DNA repair enzyme MRE11 , to initiate DNA repair, within 13 seconds. γH2AX, the phosphorylated form of H2AX is also involved in the early steps leading to chromatin decondensation after DNA double-strand breaks. The histone variant H2AX constitutes about 10% of the H2A histones in human chromatin. γH2AX (H2AX phosphorylated on serine 139) can be detected as soon as 20 seconds after irradiation of cells (with DNA double-strand break formation), and half maximum accumulation of γH2AX occurs in one minute. The extent of chromatin with phosphorylated γH2AX

9646-432: The micrococcal nuclease enzyme is employed to identify nucleosome positioning. Well positioned nucleosomes are seen to have enrichment of sequences. 3. Assay for transposase accessible chromatin sequencing ( ATAC-seq ) is used to look in to regions that are nucleosome free (open chromatin). It uses hyperactive Tn5 transposon to highlight nucleosome localisation. Activator (genetics) A transcriptional activator

9752-417: The minor grooves. Activator-binding sites may be located very close to the promoter or numerous base pairs away. If the regulatory sequence is located far away, the DNA will loop over itself (DNA looping) in order for the bound activator to interact with the transcription machinery at the promoter site. In prokaryotes, multiple genes can be transcribed together ( operon ), and are thus controlled under

9858-420: The most radiation-resistant known organism, exhibit remarkable resistance to the double-strand break-inducing effects of radioactivity , likely due to enhanced efficiency of DNA repair and especially NHEJ. A number of individual genes have been identified as influencing variations in life span within a population of organisms. The effects of these genes is strongly dependent on the environment, in particular, on

9964-460: The nuclear DNA of rodents, although similar effects have not been observed in mitochondrial DNA. The C. elegans gene AGE-1, an upstream effector of DNA repair pathways, confers dramatically extended life span under free-feeding conditions but leads to a decrease in reproductive fitness under conditions of caloric restriction. This observation supports the pleiotropy theory of the biological origins of aging , which suggests that genes conferring

10070-756: The organism's diet. Caloric restriction reproducibly results in extended lifespan in a variety of organisms, likely via nutrient sensing pathways and decreased metabolic rate . The molecular mechanisms by which such restriction results in lengthened lifespan are as yet unclear (see for some discussion); however, the behavior of many genes known to be involved in DNA repair is altered under conditions of caloric restriction. Several agents reported to have anti-aging properties have been shown to attenuate constitutive level of mTOR signaling, an evidence of reduction of metabolic activity , and concurrently to reduce constitutive level of DNA damage induced by endogenously generated reactive oxygen species. For example, increasing

10176-421: The other hand, ubiquitination decreases the activity of activators, as ubiquitin marks proteins for degradation after they have performed their respective functions. In prokaryotes, a lone activator protein is able to promote transcription. In eukaryotes, usually more than one activator assembles at the binding-site, forming a complex that acts to promote transcription. These activators bind cooperatively at

10282-496: The process involves specialized polymerases either bypassing or repairing lesions at locations of stalled DNA replication. For example, Human DNA polymerase eta can bypass complex DNA lesions like guanine-thymine intra-strand crosslink, G[8,5-Me]T, although it can cause targeted and semi-targeted mutations. Paromita Raychaudhury and Ashis Basu studied the toxicity and mutagenesis of the same lesion in Escherichia coli by replicating

10388-412: The processive polymerase to continue replication. Cells exposed to ionizing radiation , ultraviolet light or chemicals are prone to acquire multiple sites of bulky DNA lesions and double-strand breaks. Moreover, DNA damaging agents can damage other biomolecules such as proteins , carbohydrates , lipids , and RNA . The accumulation of damage, to be specific, double-strand breaks or adducts stalling

10494-459: The promoter and thus transcription, producing the enzymes that are needed to break down the maltose that has entered the cell. The catabolite activator protein (CAP), otherwise known as cAMP receptor protein (CRP), activates transcription at the lac operon of the bacterium Escherichia coli . Cyclic adenosine monophosphate (cAMP) is produced during glucose starvation; this molecule acts as an allosteric effector that binds to CAP and causes

10600-449: The same regulatory sequence. In eukaryotes, genes tend to be transcribed individually, and each gene is controlled by its own regulatory sequences. Regulatory sequences where activators bind are commonly found upstream from the promoter, but they can also be found downstream or even within introns in eukaryotes. Binding of the activator to its regulatory sequence promotes gene transcription by enabling RNA polymerase activity. This

10706-399: The shortest lived species, mouse, expresses DNA repair genes, including core genes in several DNA repair pathways, at a lower level than do humans and naked mole rats. Furthermore several DNA repair pathways in humans and naked mole-rats are up-regulated compared to mouse. These observations suggest that elevated DNA repair facilitates greater longevity . If the rate of DNA damage exceeds

10812-451: The site of the post-translational modifications, such as the one seen in H3K36me3 . The post-translational modification of histone tails by either histone modifying complexes or chromatin remodeling complexes are interpreted by the cell and lead to complex, combinatorial transcriptional output. It is thought that a histone code dictates the expression of genes by a complex interaction between

10918-452: The standard double helix. Unlike proteins and RNA , DNA usually lacks tertiary structure and therefore damage or disturbance does not occur at that level. DNA is, however, supercoiled and wound around "packaging" proteins called histones (in eukaryotes), and both superstructures are vulnerable to the effects of DNA damage. DNA damage can be subdivided into two main types: The replication of damaged DNA before cell division can lead to

11024-433: The surface features of the specific DNA regulatory sequence it was designed to bind to. The complementary interactions between the amino acids of the activator protein and the functional groups of the DNA create an "exact-fit" specificity between the activator and its regulatory DNA sequence. Most activators bind to the major grooves of the double helix, as these areas tend to be wider, but there are some that will bind to

11130-498: The undamaged DNA strand. Double-strand breaks, in which both strands in the double helix are severed, are particularly hazardous to the cell because they can lead to genome rearrangements . In fact, when a double-strand break is accompanied by a cross-linkage joining the two strands at the same point, neither strand can be used as a template for the repair mechanisms, so that the cell will not be able to complete mitosis when it next divides, and will either die or, in rare cases, undergo

11236-490: Was awarded to Tomas Lindahl , Paul Modrich , and Aziz Sancar for their work on the molecular mechanisms of DNA repair processes. DNA damage, due to environmental factors and normal metabolic processes inside the cell, occurs at a rate of 10,000 to 1,000,000 molecular lesions per cell per day. While this constitutes at most only 0.0003125% of the human genome's approximately 3.2 billion bases, unrepaired lesions in critical genes (such as tumor suppressor genes ) can impede

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