Abstract: The Hippo pathway is an evolutionarily conserved signaling cascade regulating numerous biological processes, including cell growth and fate decision, organ size control, and regeneration. The core of the Hippo pathway in mammals consists of a kinase cascade, LATS1/2 and MST1/2 that controls various cellular processes through orchestrating the phosphorylation of downstream substrates including YAP and TAZ. In keeping with a possible tumor suppressive role of the Hippo signaling pathway, it has been found that Hippo pathway dysregulation is common in many human tumors including breast, glioma, lung, colorectal cancer, and endometrial cancer. However, although a few of upstream regulators and downstream substrates were identified, the exact molecular mechanisms underlying how upstream signaling pathways control LATS1/2 kinase activity and its physiological functions in breast cancer have not yet been fully elucidated. Hence, the major goal of this proposal is to explore the upstream regulator of LATS1/2 as well as to uncover a novel tumor suppressor role of LATS1/2 in controlling tumorigenesis in the breast cancer setting. To this end, I have obtained preliminary data showing that the deubiquitinating enzyme OTUD3, but not other OTUD family member, specifically interacts and deubiquitinates LATS1. More importantly, I identified Raptor, one of the core components of mTORC1 which is a central cell growth regulator governing cellular metabolism, as a novel phosphorylation substrate of LATS1/2. In this proposal, I plan to: 1) characterize OTUD3 as a novel upstream regulator that positively regulates LATS1/2 kinase activity largely through deubiquitination of LATS1/2; 2) determine the physiological role of LATS1/2 in suppressing breast tumorigenesis largely through phosphorylating Raptor at Ser606 site, which in turn inhibits the kinase activity of mTORC1; 3) determine whether and how Raptor phosphorylation at Ser606 by LATS1/2 regulates breast cancer development in vivo. The long-term goals of my career are to apply the insights of molecular and cellular biology studies to understand the physiological significance of deregulated Hippo/mTORC1 signaling pathways that are important in the development of human malignancies, especially in breast cancer, and to search for proper druggable targets for better anti-breast cancer treatment. This K99/R00 award will provide protected time for me to pursue the novel hypotheses of this proposal, obtain new skill sets to execute experiments and solve problems. In addition, the K99 award will allow me to focus my efforts on independently conducting basic and translational research, and to train future young scientists in the cancer biology field.

Project narrative: Dysregulation of the tumor suppressors LATS1/2, core components of Hippo pathway, is common in many human tumors including breast cancer, but its upstream regulator and physiological role in breast tumorigenesis remains unclear. This proposal aims to elucidate how LATS1/2 kinase activity and tumor suppressive function is regulated by upstream deubiquitinase OTUD3, as well as to examine how LATS1/2 are critically involved in breast tumorigenesis through phosphorylating Raptor at Ser606, which in turn suppresses mTORC1 kinase activity.