User:Yousefmalnajjar/G-quadruplex

Genome Regulation through formation of G-quadruplex structures (YA)
Many of the genome regulatory processes have been linked to the formation of G-quadruplex structures, attributable to the huge role it plays in DNA repair of apurinic/apyrimidinic sites also known as AP sites.. A new technique to map AP sites has been developed known as AP-seq which utilizes a biotin-labeled aldehyde-reactive probe (ARP) to tag certain regions of the genome where AP site damage occurrence has been significant. Another genome-wide mapping sequencing method known as ChIP-sequencing, was utilized to map both; damage in AP sites, and the enzyme responsible for its repair, AP endonuclease 1 (APE1). Both of these genome-wide mapping sequencing methods, ChIP-sequencing and ARP, have indicated that AP site damage occurrence is nonrandom. AP site damage was also more prevalent in certain regions of the genome that contain specific active promoter and enhancer markers, some of which were linked to regions responsible for lung adenocarcinoma and colon cancer. AP site damage was found to be predominant in PQS regions of the genome, where formation of G-quadruplex structures is regulated and promoted by the DNA repair process, base excision repair (BER). Base excision repair processes in cells have been proved to be reduced with aging as its components in the mitochondria begin to decline, which can lead to the formation of many diseases such as Alzheimer's disease (AD). These G-quadruplex structures are said to be formed in the promoter regions of DNA through superhelicity, which favors the unwinding of the double helical structure of DNA and in turn loops the strands to form G-quadruplex structures in guanine rich regions. The BER pathway is signalled when it indicates an oxidative DNA base damage, where structures like, 8-Oxoguanine-DNA glycosylase 1 (OGG1), APE1 and G-quadruplex play a huge role in its repair. These enzymes participate in BER to repair certain DNA lesions such as 7,8-dihydro-8-oxoguanine (8-oxoG), which forms under oxidative stress to guanine bases.

 Role of Endogenous Oxidize DNA Base Damage on G4 formation (BI)

Guanine (G) bases in G-quadruplex have the lowest redox potential causing it to be more susceptible to the formation of 8-oxoguanine (8-oxoG), an endogenous oxidized DNA base damage in the genome. Due to Guanine having a lower electron reduction potential than the other nucleotides bases, 8-oxo-2'-deoxyguanosine (8-oxo-dG), is a known major product of DNA oxidation. Its concentration is used as a measurement of oxidative stress within a cell. When DNA undergoes oxidative damage, a possible structural change in guanine, after ionizing radiation, gives rise to an enol form, 8-OH-Gua. This oxidative product is formed through a tautomeric shift from the original damage guanine, 8-oxo-Gua, and represents DNA damage that causes changes in the structure. This form allows for the base excision repair (BER) enzyme OGG1 to bind and remove the oxidative damage with the help of APE1, resulting in an AP site. Moreover, an AP site is a location in DNA that has neither a purine or a pyrimidine base due to DNA damage, they are the most prevalent type of endogenous DNA damage in cells. AP sites can be generated spontaneously or after the cleavage of modified bases, like 8-OH-Gua. The generation of an AP site enables the melting of the duplex DNA to unmask the PQS, adopting a G-quadruplex fold. With the use of genome-wide ChIP-sequencing analyses, cell-based assays, and in vitro biochemical analyses, a connection has been made between oxidized DNA base-derived AP sites, and the formation of the G-quadruplex.

DNA Oxidation Contribution to Diseases (BI)

Furthermore, the concentration of 8-oxo-dG is a known biomarker of oxidative stress within a cell, and excessive amount of oxidative stress has been linked to carcinogenesis and other diseases. When produced, 8-oxo-dG, has the ability to inactivate OGG1, thus preventing the repair of DNA damage caused by the oxidation of guanine. The possible inactivation allows for un-repaired DNA damages to gather in non-replicating cells, like muscle, and can cause aging as well. Moreover, oxidative DNA damage like 8-oxo-dG contributes to carcinogenesis through the modulation of gene expression, or the induction of mutations. On the condition that 8-oxo-dG is repaired by BER, parts of the repair protein is left behind which can lead to epigenetic alterations, or the modulation of gene expression. Upon insertion of 8-oxo-dG into thymidine kinase gene of humans, it was determine that if 8-oxo-dG was left unchecked and not repaired by BER, it can lead to frequent mutations and eventually carcinogenesis.

APE1 role in Gene Regulation (YA)
AP endonuclease 1 (APE1) is an enzyme responsible for the promotion and the formation of G-quadruplex structures. APE1 is mainly in charge of repairing damage caused to AP sites through the BER pathway. APE1 is considered to be very crucial as AP site damage is known to be the most recurring type of endogenous damage to DNA. The oxidation of certain purine bases, like guanine, forms oxidized nucleotides that impairs DNA function by mismatching nucleotides in the sequences. This is more common in PQS sequences which form oxidized structures, such as 8-oxoguanine. Once the cell is aware of oxidative stress and damage, it recruits OGG1 to the site, whose main function is to initiate the BER pathway. OGG1 does so by cleaving the oxidized base and thus creating an AP site, primarily through the process of negative superhelicity. This AP site then signals cells to engage APE1 binding, which binds to the open duplex region. The binding of APE1 then plays an important role by stabilizing the formation of G-quadruplex structures in that region. This promotes formation of G-quadruplex structures by the folding of the stand. This looping process brings four bases in close proximity that will be held together by Hoogsteen base pairing. After this stage the APE1 gets acetylated by multiple lysine residues on the chromatin, forming acetylated APE1 (AcAPE1). AcAPE1 is very crucial to the BER pathway as it acts as a transcriptional coactivator or corepressor, functioning to load transcription factors (TF) into the site of damage allowing it to regulate the gene expression. AcAPE1 is also very important since it allows APE1 to bind for longer periods of time by delay of its dissociation from the sequence, allowing the repair process to be more efficient. Deacetylation of AcAPE1 is the driving force behind the loading of these TFs, where APE1 dissociates from the G-quadruplex structures. When a study downregulated the presence of APE1 and AcAPE1 in the cell, the formation of G-quadruplex structures was inhibited, which proves the importance of APE1 for the formation of these structures. However, not all G-quadruplex structures require APE1 for formation, in fact some of them formed greater G-quadruplex structures in its absence. Therefore we can conclude that APE1 has two important roles in genome regulation- Stabilizing the formation of g-quadruplex structures and loading the transcriptional factors onto the AP site.