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ATR-mediated mechanotransduction and connections with the actin cytoskeleton

Periodic Reporting for period 1 - MECHANOCHECK (ATR-mediated mechanotransduction and connections with the actin cytoskeleton)

Reporting period: 2016-04-01 to 2018-03-31

ATR is a protein kinase that controls DNA damage response (DDR), together with ATM, Chk1 and Chk2. DDR genes are often mutated in cancer cells and act as an anti-cancer barrier in response to oncogenic stimuli. ATR is essential and protects the integrity of replicating chromosomes, prevents fragile site expression and aberrant condensation events. While the tumour suppressive role of ATR in mediating the DNA damage response has been extensively studied, the recent findings from the Foiani’s laboratory that ATR controls chromatin association to the nuclear envelope and responds to mechanical stress by re-localizing at nuclear membranes, prompted us to propose a novel role for ATR in controlling cellular and nuclear plasticity.
Using multidisciplinary techniques and experimental approaches, including AFM, micropattern and innovated cell stretching and cell compression device, we quantitatively studied how ATR acts on cellular and nuclear plasticity. We found that ATR functions control cell plasticity, which sense mechanical stress. Basically, ATR-defective cells become twice softer than normal cells. At spatio-temporal level, the mechano-response occurs primarily through the actin cytoskeleton, which forms the cellular mechanical system linking the extracellular microenvironment to the nucleus. We investigated the connection between ATR and actin and we surprisingly found out that ATR directly controls nuclear plasticity, which in turn controls cell plasticity, rather than play a role in actin organization and dynamics. ATR defective cells have compromised nuclear morphology and organization; these cells are highly susceptive to mechanical stress, and have a lower survive rate during interstitial migration. These results suggest that ATR could play an active role in influencing the metastatic process, thus implying that ATR may possess a tumorigenic function.
This line of research contributes to open new directions in the ATR field and to link the control of cellular and nuclear plasticity with the capability of cells in interstitial migration. Thus it might lead to novel target for cancer metastasis and new cancer drug discovery.
The results of present project (Mechanocheck) show that despite its well-known function in DNA damage response, ATR play a novel role in controlling cellular and nuclear plasticity and affect interstitial migration. The novel finding has been disseminated in various conferences. Besides that, the project leads to two novel devices: cell stretching and cell compression device. Both of them have been successfully patented. Cell stretching device was reported in an Italian National TV programs (SuperQuark, Tutta Salute). First, a new assay (assembly of perinuclear actin ring during prophase) was established to test the ATR drug effect on perinuclear actin dynamics. The results show that ATR doesn’t affect perinuclear actin assembly in prophase. Further, using the micropattern technique, we confirmed that ATR is not involved in actin organization. Interestingly, using AFM, we quantitively show that ATR influences cell plasticity. Moreover, ATR was found to affect nuclear plasticity directly. By innovated cell compression technique, ATR defective cells have compromised nuclei, which are susceptible to mechanical stress. ATR defective cells exhibit an inadequate in interstitial migration, suggesting a key role in cancer migration and metastasis.
The results and developed novel techniques have been disseminated in a various conferences, including public and scientific communication initiatives. They have been also shared with companies to see the potential of commercialization.
Mechanocheck has achieved most of its objectives. Despite its well-known function in DNA damage response, we found a novel role of ATR in controlling cellular and nuclear plasticity. The project opens new scientific directions in the ATR field and to link the control of cellular and nuclear plasticity with the capability of cells in interstitial migration and cancer metastasis. This could of impact to lead to novel cancer cell target and new drug development in clinical application. Beyond that, the original and innovative techniques designed for the project have been successfully patented. We are currently planning to commercialize the devices through collaboration with companies and start up a new technology company.
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