Laura R. Pearce

The mTOR (mammalian target of rapamycin) protein kinase is an important regulator of cell growth. Two complexes of mTOR have been identified: complex 1, consisting of mTOR-Raptor (regulatory associated protein of mTOR)-mLST8 (termed mTORC1), and complex 2, comprising mTOR-Rictor (rapamycininsensitive companion of mTOR)-mLST8-Sin1 (termed mTORC2). mTORC1 phosphorylates the p70 ribosomal S6K (S6 kinase) at its hydrophobic motif (Thr389), whereas mTORC2 phosphorylates PKB (protein kinase B) at its hydrophobic motif (Ser473). In the present study, we report that widely expressed isoforms of unstudied proteins termed Protor-1 (protein observed with Rictor-1) and Protor-2 interact with Rictor and are components of mTORC2. We demonstrate that immunoprecipitation of Protor-1 or Protor-2 results in the co-immunoprecipitation of other mTORC2 subunits, but not Raptor, a specific component of mTORC1. We show that detergents such as Triton X-100 or n-octylglucoside dissociate mTOR and mLST8 from a complex of Protor-1, Sin1 and Rictor. We also provide evidence that Rictor regulates the expression of Protor-1, and that Protor-1 is not required for the assembly of other mTORC2 subunits into a complex. Protor-1 is a novel Rictor-binding subunit of mTORC2, but further work is required to establish its role.

S6K1 (p70 ribosomal S6 kinase 1) is activated by insulin and growth factors via the PI3K (phosphoinositide 3-kinase) and mTOR (mammalian target of rapamycin) signalling pathways. S6K1 regulates numerous processes, such as protein synthesis, growth, proliferation and longevity, and its inhibition has been proposed as a strategy for the treatment of cancer and insulin resistance. In the present paper we describe a novel cell-permeable inhibitor of S6K1, PF-4708671, which specifically inhibits the S6K1 isoform with a Ki of 20 nM and IC50 of 160 nM. PF-4708671 prevents the S6K1-mediated phosphorylation of S6 protein in response to IGF-1 (insulin-like growth factor 1), while having no effect upon the PMA-induced phosphorylation of substrates of the highly related RSK (p90 ribosomal S6 kinase) and MSK (mitogen- and stress-activated kinase) kinases. PF-4708671 was also found to induce phosphorylation of the T-loop and hydrophobic motif of S6K1, an effect that is dependent upon mTORC1 (mTOR complex 1). PF-4708671 is the first S6K1-specific inhibitor to be reported and will be a useful tool for delineating S6K1-specific roles downstream of mTOR.

Kinase suppressor of Ras 2 (KSR2) is an intracellular scaffolding protein involved in multiple signaling pathways. Targeted deletion of Ksr2 leads to obesity in mice, suggesting a role in energy homeostasis. We explored the role of KSR2 in humans by sequencing 2,101 individuals with severe early-onset obesity and 1,536 controls. We identified multiple rare variants in KSR2 that disrupt signaling through the Raf-MEK-ERK pathway and impair cellular fatty acid oxidation and glucose oxidation in transfected cells; effects that can be ameliorated by the commonly prescribed antidiabetic drug, metformin. Mutation carriers exhibit hyperphagia in childhood, low heart rate, reduced basal metabolic rate and severe insulin resistance. These data establish KSR2 as an important regulator of energy intake, energy expenditure, and substrate utilization in humans. Modulation of KSR2-mediated effects may represent a novel therapeutic strategy for obesity and type 2 diabetes.PaperFlickeyJraWQiOiI4ZjUxYWNhY2IzYjhiNjNlNzFlYmIzYWFmYTU5NmZmYyIsImFsZyI6IlJTMjU2In0.eyJzdWIiOiIxM2ZkMGVjMGEwODY1YzEyM2U0MGRhMTQ1ZjYzZjgxYiIsImtpZCI6IjhmNTFhY2FjYjNiOGI2M2U3MWViYjNhYWZhNTk2ZmZjIiwiZXhwIjoxNjc4NTE4ODkzfQ.ebCaKXCW1UeAldICT0NS9vU7ygd7nXXkUCWgpBUORRwMIWQcffyUi9TjJzPxUP3iWYFTjfzKxX6G_vO5XV0dkunKbLKrQVElOe9j3KSH0QUXO9-sfbDSbvwMoXd2IsOgJbl5lMc4qCKRpF915CT_nHltuNb1kY-uU71BXgDjjTMnTBv8qNHVk4gzUPj81_X8K965fhLHckx6U_QSDreSIvmTYWxC5TDxe3FYQW90h94RCtWjw0CA-WOAqZ4MiIUipTAp_lstwruI1nYkm9Ao4xHlB9xGiE6NSECjEJqPLBob2mCn0SGkK81_EcSuGeK-9G1Wd-7RdAgMztzISI0T1g(mp4, (45.2 MB) Download video

Src homology 2 B adapter protein 1 (SH2B1) modulates signaling by a variety of ligands that bind to receptor tyrosine kinases or JAK-associated cytokine receptors, including leptin, insulin, growth hormone (GH), and nerve growth factor (NGF). Targeted deletion of Sh2b1 in mice results in increased food intake, obesity, and insulin resistance, with an intermediate phenotype seen in heterozygous null mice on a high-fat diet. We identified SH2B1 loss-of-function mutations in a large cohort of patients with severe early-onset obesity. Mutation carriers exhibited hyperphagia, childhood-onset obesity, disproportionate insulin resistance, and reduced final height as adults. Unexpectedly, mutation carriers exhibited a spectrum of behavioral abnormalities that were not reported in controls, including social isolation and aggression. We conclude that SH2B1 plays a critical role in the control of human food intake and body weight and is implicated in maladaptive human behavior.