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ATM-SPARK: A GFP phase separation-based activity reporter of ATM

The kinase ataxia telangiectasia mutated (ATM) plays a key role in the DNA damage response (DDR). It is thus essential to visualize spatiotemporal dynamics of ATM activity during DDR. Here, we designed a robust ATM activity reporter based on phosphorylation-inducible green fluorescent protein phase separation, dubbed ATM-SPARK (separation of phases-based activity reporter of kinase). Upon ATM activation, it undergoes phase separation via multivalent interactions, forming intensely bright droplets. The reporter visualizes spatiotemporal dynamics of endogenous ATM activity in living cells, and its signal is proportional to the amount of DNA damage. ATM-SPARK also enables high-throughput screening of biological and small-molecule regulators. We identified the protein phosphatase 4 that blocks ATM activity. We also identified BGT226 as a potent ATM inhibitor with a median inhibitory concentration of ~3.8 nanomolars. Furthermore, BGT226 sensitizes cancer cells to the radiomimetic drug neocarzinostatin, suggesting that BGT226 might be combined with radiotherapeutic treatment. ATM-SPARK achieves large dynamic range, bright fluorescence, and simple signal pattern.

 

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The method, called ATM-SPARK, uses a phosphorylation-inducible green fluorescent protein phase separation to form bright droplets in response to ATM activation. This allows for the spatiotemporal dynamics of ATM activity to be observed in living cells, with the signal intensity proportional to the amount of DNA damage.

The ATM-SPARK method also enables high-throughput screening of biological and small-molecule regulators. In fact, the authors of the study were able to identify the protein phosphatase 4 as a blocker of ATM activity, and they discovered the small molecule BGT226 as a potent ATM inhibitor with a median inhibitory concentration of about 3.8 nanomolars. Furthermore, they found that BGT226 can sensitize cancer cells to the radiomimetic drug neocarzinostatin, suggesting that it may be useful in combination with radiotherapeutic treatments.

Overall, ATM-SPARK appears to be a powerful tool for studying ATM activity and identifying potential regulators and inhibitors. Its large dynamic range, bright fluorescence, and simple signal pattern make it an attractive option for high-throughput screening and drug discovery.

 

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