Abstract 219: Identification of the mechanistic target of rapamycin complex 2-associated interactome involved in brain cancer cell motility by affinity purification-mass spectrometry
Authors: Chantaravisoot Naphat , Wongkongkathep Piriya , Tamanoi Fuyuhiko , Pisitkun Trairak 
Published: July 2017

Abstract
The Mechanistic Target of Rapamycin Complex 2 (mTORC2), a multiprotein complex with serine-threonine kinase activity, drives several cellular processes of normal cells to promote proper metabolic activity, survival, proliferation, differentiation and movement. On the other hand, mTORC2 has been implicated as a critical player in tumorigenesis, stimulation of cell growth, cancer metabolic reprogramming, and development of highly migratory and invasive capabilities of cancers. Therefore, characterization of mTORC2 is necessary not only for better understandings of known mTORC2-mediated functions, but also for the discovery of new roles and more interacting partners of mTORC2 signaling network in cancers, particularly ones with hyperactivated mTORC2 such as glioblastoma. However, until now mTORC2 is still partially characterized. Previously, mTORC2 was reported to be associated with Filamin A and Myosin-9 which have been known to play important roles in cellular activities requiring locomotion, including cancer migration and invasion. The complex participates in regulating glioblastoma migration and invasion, and Filamin A acts as a physiological downstream target of mTORC2. Currently, proteomics has emerged as a powerful tool to identify, quantify, and examine a large number of proteins. Therefore, in order to extensively unravel mTORC2 signaling pathway, the use of proteomics is necessary.
We characterized mTORC2 by proteomic analysis based on affinity purification coupled to mass spectrometry (AP-MS) using antibodies against RICTOR, a specific component of mTORC2. The protein-protein interactions between RICTOR and other associated proteins were assessed. In addition to canonical components of mTORC2, the study has revealed more insights into a number of mTORC2 interacting partners which can be categorized and related to the main functions previously identified such as actin cytoskeleton reorganization, fatty acid metabolism, protein translation. Interestingly, we also found other types of cytoskeletal proteins, nucleotide metabolism, less-characterized and low abundance proteins associated with mTORC2 such as Hornerin and Desmoplakin. In addition, we have carried out the identification of condition-specific interactions for a dynamic view of mTORC2 interactome and found changes in the mTORC2-associated proteins from different conditions. Furthermore, proteomic approaches can also be relevant to investigation and prediction of mTORC2 upstream regulators, downstream targets, and complex localization. Ultimately, this project will uncover some unknown molecular mechanisms, and result in a significant impact on the field of signal transduction machinery in cancers.
Citation Format: Naphat Chantaravisoot, Piriya Wongkongkathep, Fuyuhiko Tamanoi, Trairak Pisitkun. Identification of the mechanistic target of rapamycin complex 2-associated interactome involved in brain cancer cell motility by affinity purification-mass spectrometry [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 219. doi:10.1158/1538-7445.AM2017-219