Regulation of mTOR Complexes in Long-Lived Growth Hormone Receptor Knockout and Snell Dwarf Mice

mTOR is a serine/threonine kinase that binds with multiple proteins to form two complexes, mTORC1 and mTORC2. mTORC1 receives upstream signals including amino acids, growth factors, energy and oxygen levels to regulate cell growth and metabolism. mTORC2 is regulated by insulin/IGF-1 signaling to modulate cell survival and proliferation. mTOR is well known to play a critical role in aging. A reduction in mTORC1 activity extends lifespan in multiple species, including yeast, worms, flies and mice. Growth hormone receptor knockout (GHRKO) mice and Snell dwarf mice are two long-lived mouse models with lower body weight and significantly extended lifespan. Tissues of these mice have lower mTORC1 activity, but the mechanisms are not clear. Here, two ideas were tested about the regulation of mTOR in GHRKO and Snell mice, one involving alteration in the components of the mTOR complexes, and the other focused on the upstream regulation of mTORC1 function. GHRKO and Snell dwarf liver, kidney and skeletal muscle were used to study the protein expression of mTORC components. Only two proteins, DEPTOR and PRAS40 which inhibit mTOR activity, were reduced in the liver, opposite in the direction expected to cause lower mTORC1 function. In addition, DEPTOR or PRAS40 knockdown in GHRKO fibroblasts did not alter mTORC substrate phosphorylation, indicating that the deficit in mTORC1 activity in GHRKO or Snell dwarf mice is not likely to reflect differences in levels of DEPTOR or PRAS40. TSC is a master regulator of mTORC1 activity that integrates upstream signals from growth factors, energy and oxygen levels. TSC components TSC1 and TSC2 were tested in GHRKO and Snell dwarf liver, kidney and muscle. The results showed increased TSC1 and TSC2 protein expression in all tissues tested. Relative phosphorylation of TSC2 at Ser 939 and Thr 1462, which inhibits activity of TSC2, were decreased in these tissues as well, suggesting increased TSC inhibition of mTORC1. TSC2 knockdown in GHRKO fibroblasts augmented the phosphorylation of mTORC1 substrates, consistent with the model in which mTORC1 activity was regulated by TSC. In summary, the results showed that the lower mTORC1 activity in GHRKO and Snell dwarf mice is not regulated by the levels of mTORC components. Instead, the increased amounts of unphosphorylated, active, inhibitory TSC might contribute to lower mTORC1 function in these long-lived mice. This study highlights the potential role of TSC in regulating mTORC1 activity in long-lived mutant mice. Future studies on enhancing TSC function by overexpressing TSC2 or using TSC2 activating molecules may generate additional mouse models that can be used as comparisons with GHRKO and Snell dwarf mice, which would provide more evidence about the role of mTOR in regulating the pace of aging.PhDMolecular & Cellular PathologyUniversity of Michigan, Horace H. Rackham School of Graduate Studieshttp://deepblue.lib.umich.edu/bitstream/2027.42/172734/1/xfshi_1.pd