Genome-wide shRNA screen reveals increased mitochondrial dependence upon mTORC2 addiction

Colombi, Marco; Molle, Klaus D.; Benjamin, Don; Rattenbacher-Kiser, Karin F.; Schaefer, Christiane; Betz, Charles; Thiemeyer, Anke; Regenass, Urs; Hall, Michael N.; Moroni, Christoph

doi:10.1038/onc.2010.539

Cited by 48 publications

(52 citation statements)

References 44 publications

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“…Indeed, when cultured in the absence of IL-3, 6.5 cells undergo apoptosis in the presence of nanomolar amounts of rapamycin. 21 In contrast, it is impossible to induce apoptosis with rapamycin in human cancer cell lines, and micromolar concentrations of rapamycin are required to see (partial) growth inhibition. We therefore assume that despite PTEN silencing and Ras activation, additional mutations in human cancer cells might overrun components of the Akt-mTOR pathway and therefore render the cells less addicted.…”

Section: Discussionmentioning

confidence: 99%

“…Indeed, mTOR and mTORC2 were under the genes identified in the shRNA screen, confirming the specificity of the system. 21 Thus, we used the same system for a chemical HTS. We could identify compounds that fall into two main mechanistic classes.…”

Section: Discussionmentioning

confidence: 99%

“…To further validate this system, the parental clone 15v4 was stably transfected with an shRNA against PTEN. 21 Indeed, also the resulting clone shPTEN was sensitive to rapamycin, and its effect could be abrogated by the addition of exogenous IL-3 ( Fig. 1C).…”

Section: Isogenic Cell Systemmentioning

confidence: 99%

“…Many of these IL-3-independent clones were shown to have an activated mTOR pathway. 18,21 Several clones had mutations or deletions of the tumor suppressor PTEN, such as clone 6.5. The clone designated D2c harbors a mutation in the tsc2 gene, leading to complete absence of TSC2 protein (Fig.…”

Section: Isogenic Cell Systemmentioning

confidence: 99%

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An Isogenic Cell Panel Identifies Compounds That Inhibit Proliferation of mTOR-Pathway Addicted Cells by Different Mechanisms

et al. 2014

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The mTOR pathway is a critical integrator of nutrient and growth factor signaling. Once activated, mTOR promotes cell growth and proliferation. Several components of the mTOR pathway are frequently deregulated in tumors, leading to constitutive activation of the pathway and thus contribute to uncontrolled cell growth. We performed a high-throughput screen with an isogenic cell line system to identify compounds specifically inhibiting proliferation of PTEN/mTOR-pathway addicted cells. We show here the characterization and mode of action of two such compound classes. One compound class inhibits components of the PTEN/mTOR signaling pathway, such as S6 ribosomal protein phosphorylation, and leads to cyclin D3 downregulation. These compounds are not adenosine triphosphate competitive inhibitors for kinases in the pathway, nor do they require FKBP12 for activity like rapamycin. The other compound class turned out to be a farnesylation inhibitor, blocking the activity of GTPases, as well as an inducer of oxidative stress. Our results demonstrate that an isogenic cell system with few specific mutations in oncogenes and tumor suppressor genes can identify different classes of compounds selectively inhibiting proliferation of PTEN/mTOR pathway-addicted isogenic clones. The identified mechanisms are in line with the known cellular signaling networks activated by the altered oncogenes and suppressor genes in the isogenic system.

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Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Section: Isogenic Cell Systemmentioning

confidence: 99%

Section: Isogenic Cell Systemmentioning

confidence: 99%

See 2 more Smart Citations

An Isogenic Cell Panel Identifies Compounds That Inhibit Proliferation of mTOR-Pathway Addicted Cells by Different Mechanisms

et al. 2014

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“…Furthermore, deficiency of rictor in Jurkat cells, a leukemic T cell line, increased oxygen consumption (Schieke et al 2006). However, mTORC2 could play a more complex function in mitochondrial metabolism since a PTEN-deficient cell line that is mTORC2 addicted/IL3-independent was shown to require a number of genes involved in mitochondrial functions (Colombi et al 2011). Adipose-specific knockout of rictor in mice revealed that mTORC2 can function to control whole body growth (Cybulski et al 2009).…”

Section: Metabolismmentioning

confidence: 99%

The Target of Rapamycin: Structure and Functions

Wu¹,

Chou²,

Jacinto³

2012

Protein Kinases

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mTORC1 and mTORC2 as regulators of cell metabolism in immunity

et al. 2017

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The mechanistic target of rapamycin (mTOR) pathway is an evolutionarily conserved signaling pathway that senses intra-and extracellular nutrients, growth factors, and pathogen-associated molecular patterns to regulate the function of innate and adaptive immune cell populations. In this review, we focus on the role of the mTOR complex 1 (mTORC1) and mTORC2 in the regulation of the cellular energy metabolism of these immune cells to regulate and support immune responses. In this regard, mTORC1 and mTORC2 generally promote an anabolic response by stimulating protein synthesis, glycolysis, mitochondrial functions, and lipid synthesis to influence proliferation and survival, effector and memory responses, innate training and tolerance as well as hematopoietic stem cell maintenance and differentiation. Deactivation of mTOR restores cell homeostasis after immune activation and optimizes antigen presentation and memory T-cell generation. These findings show that the mTOR pathway integrates spatiotemporal information of the environmental and cellular energy status by regulating cellular metabolic responses to guide immune cell activation. Elucidation of the metabolic control mechanisms of immune responses will help to generate a systemic understanding of the immune system.

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Genome-wide shRNA screen reveals increased mitochondrial dependence upon mTORC2 addiction

Cited by 48 publications

References 44 publications

An Isogenic Cell Panel Identifies Compounds That Inhibit Proliferation of mTOR-Pathway Addicted Cells by Different Mechanisms

An Isogenic Cell Panel Identifies Compounds That Inhibit Proliferation of mTOR-Pathway Addicted Cells by Different Mechanisms

The Target of Rapamycin: Structure and Functions

mTORC1 and mTORC2 as regulators of cell metabolism in immunity

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