Quantitative Analysis of ATM, ATR and γ-H2AX as DNA Damage Sensor Proteins through Western Blotting

Living cells sometimes encounter continuous or accidental exposure of genotoxic agents which misbalances the integrity of subcellular components. DNA damage by these genotoxic agents initiates various cellular responses. These cellular responses include sensors, transducers and effectors. Sensor protein act at the frontline of the DNA response mechanism. The transducer amplifies the intracellular signals to the down stream regulatory molecules. Then the effector molecules progresses towards the execution of cell cycle arrest, DNA repair or apoptosis. The most important sensor proteins are ATM and ATR. One of the most imperative substrate that is phosphorylated by these protein kinases is γ-H2AX which is a vital component for nuclear foci formation to execute DNA repair mechanisms. This complete process is under tight regulation and any imbalance in these processes may lead to hypersensitivity to cellular stress and susceptibility to DNA damage, genomic effects and resistance to apoptosis, thus initiating the cascade of carcinogenesis. There are various agents that modifies DNA molecule, for example; environmental isocyanates and their derivatives that have capability to cause toxicogenomic effects. Isocyanates are thus becoming of interest in the field of genetic toxicology as they may react with DNA to produce DNA damage.