Target of rapamycin (TOR) is the main controller of cell growth and metabolism in response to nutrients, growth factors and the cellular energy status. TOR is a serine/threonine kinase conserved from yeast to mammals and is found in two functionally and structurally distinct multi-protein complexes named TOR complex 1 (TORC1) and TORC2. Mammalian TORC1 (mTORC1) contains mTOR, mLST8, raptor and PRAS40, while mTORC2 contains mTOR, mLST8, rictor, mSin1, and PRR5. mTORC2 is activated in response to growth factors, such as insulin and insulin-like growth factor 1 (IGF1), and its main functions involve the regulation of actin cytoskeleton dynamics and phosphorylation of several AGC kinases in their hydrophobic motif. TORC1 is directly inhibited by the immunosuppressant and anti-cancer drug rapamycin, whereas TORC2 is not. Thus, use of rapamycin provides a simple and straightforward method to specifically study the TORC1 signaling branch. There is no known TORC2-specific inhibitor, so genetic manipulation is required to study its biological function(s).
This thesis describes new in vivo and in vitro functions of mTORC2. The first part deals with the in vivo function of mTORC2 in adipose tissue. The adipose tissue, in addition to its function as a long-term fat storage depot, also has endocrine functions, plays an important role in the regulation of whole body glucose and lipid metabolism and is one of the most insulin-responsive tissues in the body. To study mTORC2 function in adipose tissue, we have generated mice that lack the mTORC2-essential component rictor specifically in adipose tissue. Phenotypic characterization revealed the unexpected finding that these mice were larger due to an increase in lean tissue mass and that they had elevated serum IGF1 levels. Furthermore, the knockout mice were hyperinsulinemic, but glucose tolerant. Overall, these findings suggest an important role for adipose mTORC2 in controlling full body growth and whole body glucose metabolism.
The second part of this thesis describes a new in vitro function of mTORC2 in fibroblasts. We have taken advantage of the raptor and rictor floxed mice to isolate mouse embryonic fibroblasts (MEFs), which were then used to establish inducible raptor and rictor knockout MEF cell lines. After initial characterization of these two cell lines, a deeper analysis of the role of mTORC2 in the actin-mediated process of cell migration was performed. We have found that mTORC2 is required for cell migration and for regulating the activity of the Rho GTPases Rac1, Cdc42, and RhoA. We have extended this study by showing that mTORC2-dependent cell migration is also required in oncogenic cells, which suggests that mTORC2 could have an important function in the development of cancer and metastasis
