MYOSIN II ASSEMBLY REGULATION THROUGH THE MYOSIN PHOSPHATASE TARGETING SUBUNIT 1 DETERMINES CORTICAL MECHANICS FUNCTION ACCORDING TO MYOSIN ASSEMBLY AND MECHANORESPONSE POTENTIAL IN PANCREATIC CANCER CELLS

Pancreatic ductal adenocarcinoma (PDAC) remains one of the deadliest cancers as the fourth leading cause of cancer related deaths with a 5-year survival rate of 10.8%. A critical need is to develop new PDAC-specific prognostics and targets. To this end, we are investigating the role of the myosin phosphatase targeting subunit 1 (MYPT1) in PDAC cell mechanics. MYPT1 was originally implicated in PDAC progression through a clinical trial where it was targeted by an antibody response to a whole tumor cell vaccine. Through this trial, MYPT1 expression was also found to be upregulated in patient PDAC cells, compared to normal ductal epithelium, and in established PDAC cell lines. MYPT1 is a component of the myosin phosphatase complex where it binds to the PP1β/δ catalytic phosphatase subunit and the phosphorylated regulatory light chain (RLC) of non-muscle myosin II (NMII). The MYPT1 complex facilitates RLC dephosphorylation, which leads to inactivation of NMII. NMII bipolar thick filaments are largely responsible for cell shape changes and have been linked to the altered cell mechanics associated with cancer transformation and metastasis. As a major regulator of NMII, we hypothesize MYPT1 has implications in cancer cell shape control. To elucidate MYPT1’s function in PDAC, I developed an in vitro model system composed of engineered cell lines using CRISPR-mediated MYPT1 knockout at each stage of PDAC transformation and progression. Each of the cell lines was then characterized for alterations in NMII-mediated cortical mechanics and oncogenic behavior. Additionally, performed an in vivo orthotopic xenograft model to assess the tumor formation, growth, and metastatic abilities of these lines. I found that removing MYPT1 from the cell increase NMIIC assembly and reduced its mechanoresponse ability. At the cellular and tissue levels, the alteration in myosin IIC function causes reduced migration and dissemination in vitro and reduced tumor formation, growth, and metastatic potential in vivo. I conclude that the upregulation of MYPT1 specifically lends cells an increased deformation ability through the regulation of NMIIC dynamic assembly and mechanoresponse potential. My dissertation work demonstrates that targeting NMIIC assembly and mechanoresponse is a viable strategy to reduce PDAC tumor burden and metastasis