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Double strand breaks (DSBs) are the most dangerous form of Deoxyribonucleic Acid (DNA) damage. They present the greatest threat to cell survival, as well as to DNA integrity which is experienced in the modern day. DSBs are caused by a variety of different agents both of an endogenous and exogeneous nature. These breaks are seen as lethal if they remain unrepaired or if the break is incorrectly repaired, it may result in either chromosomal rearrangements or neoplastic transformations.
The danger caused by the DSB is seen due to the nature of how it can affect both sides of the strands, providing no scope for an intact complimentary strand to act as a template.Two main mechanisms have arrisen in cells to cope with these DSBs, the NonHomologoue End Joining (NHEJ) pathway, and also the Homologous Recombination(HR) pathway. These two pathways at first glance may appear to be quite similiar, however upon further examination it is evident that, both NHEJ and HR differ in several fundamental biochemical and structural ways. ( Lieber, M. et al (2006).(Cromie, G. A.,et al (2001). Cromie, G. A.,et al (2001b).) In higher eukaryotes, DSBs are repaired in an accurate fashion, with high faithfullness, with the effective restoration of genetic integrity. The number of endogeneous DSBs which occur throughout the cell cylce has been reported to somewhere in the region of 10-100 nucleus per division. .(Wang,et al (2008) ( Burma,et al 2006)
The mechanisms by which, these DSBs occur, is highly varied and also consistent of various mechanisms. The DSB introduction can occur by either intrinsic or extrinsic factors, which may be as a result of cellular metabolism or even as the byproduct of reactions within the body. Extrinsic factors include those such as x-rays, gamma-rays and also those caused via chemotherapeutic drugs. Failure for these to be repaired in a suitable fashion, can result in apoptosis. Mutations which occur may result in gross chromosomal rearrangements be they translocations or even deletions, and are highly detrimental to the cell life cycle.
DSBs occur at a rate of approx 10 spontaneous breaks a day, and have been shown to be of vital importance. In mammals some 30-50% of DSBs are repaired by HR, while the remainder are repaired by the NHEJ pathway, which I will compare and contrast in the following pages of this review.
Fig1: NHEJ pathway (http://www.google.ie/imgres?imgurl=http://www.gurdon.cam.ac.uk/~jacksonlab/Images/nhej.jpg&imgrefurl=http://www.gurdon.cam.ac.uk/~jacksonlab/research_interests.htm&usg=__vZwcWafW6z0mJk0sBPq37aKJWr4=&h=407&w=341&sz=23&hl=en&start=0&zoom=1&tbnid=_7I_7aY1iYzu5M:&tbnh=137&tbnw=115&prev=/images%3Fq%3DNHEJ%26um%3D1%26hl%3Den%26client%3Dfirefox-a%26rls%3Dorg.mozilla:en-GB:official%26biw%3D1708%26bih%3D719%26tbs%3Disch:1&um=1&itbs=1&iact=hc&vpx=127&vpy=53&dur=1010&hovh=245&hovw=205&tx=107&ty=173&ei=JJoITYfXKoW6hAf6tvSEDw&oei=JJoITYfXKoW6hAf6tvSEDw&esq=1&page=1&ndsp=38&ved=1t:429,r:0,s:0)
The most commonly observed Non homologous event is that of the imprecise end joining pathways, (Moore, J.K.,and Haber, J.E. (1996) )
What is NHEJ
The NHEJ pathway is reported to be over 3 million years old, as it has been seen in numerious different organisms, such as bacteria, archaea and eukaryotes. The NHEJ pathway is a mechansim to repair the DSB, which involved a group of numerous different proteins, DNA ligase IV, forms a complex with the cofactor XRCC4, which will join the two DNA ends. For accurate repair the NHEJ pathway relies on short homologous sequences, present in the single-stranded tails of the DNA to be joined.
What is HR
This is different from the mechanism of HR as in HR there is a requirement for a second intact copy of the broken chromosome segment. This copy then acts as a template for DNA synthesis across the break. In the case of HR the DSB end is cleaved, resulting in a 3' single strand tail, which invades the sister chromatid, and pairs with the complementary DNA.
FIG??: HR NEED BETTER PIC
HR is widely used for accurate repair of DSBs, and can also produce new DNA combinations. There is a lot of difference across various different organisms, in how they use HR. However the same basic steps are seen across the board. After a DSB has occurred, the DNA sections at the 5' end are removed in a process called resection, which is a medical term for removal of part of an organ. The 3' overhang, then invades into a similar DNA molecule which is not broken. After this a Holliday junction is formed in such case as there is a connection between the two DNA molecules. The means by which these junctions are cut will result in what is seen as crossover or a non crossover event. However when HR is used as a means of DNA repair the majority of the resulting products are of the non-crossover breed, and thus HR is used to restore the damaged DNA molecule to its original appearance.
As I have said previously, HR is conserved across various species, and thus it can be suggested that a similar mechanism will have arrisen across the board. This is seen as HR genes and proteins are strongly associated with an increased susceptibility to cancer, proteins involved in HR as often those which are the topic of recent cancer related research. Even as recent as the year 2007, the Nobel Prize in Medicine was awarded to a scientist, who came up with the idea of targeting HR techniques for use in molecular biology, this technique has aided with the advancement of novel treatments into not only cancer, but also numerous other DNA repair diseases.
Ageing is defined as a progressive functional decline and increasing mortality over time. DNA damage is accumulated with age, and DNA repair defects can result in phenotypes which resemble premature ageing. Damage to nuclear DNA which encode a vast majority of cellular RNA and proteins as well as those involved in mitochondrial DNA have been shown to contribute to the ageing process. (Karanjawala and Lieber, 2004). In addition to both external ageing sources such as ionizing radiation and genotoxic drugs, cell-intrinsic sources of DNA damage are also possible, which can and will ultimately lead to ageing. These may include spontaneous chemical change in the DNA, programmed DSBs, replicaton errors and also the effects of numerous radioactive oxygen species (ROS) such as hydrogen peroxide and nitric oxide to mention a few.
Fig : Pictorial diagrammatic effect of ageing on US President Obama, after his term as president of the USA
In cells the use of programmed DSB followed by repair of the break is used during the arrisal of genetic diversity, Cells express a class of endonucleases which are of the Spoo-11 class, which will induce a DSB at a random location along the chromosome, these are usually repaird by the HR pathway, and this method is used in V(D)J recombination, which will be explored later.