mTOR plays important roles in a variety of cellular processes including; cell growth, cell survival, motility, transcription and cell proliferation. It acts a central hub, integrating signals from a number of upstream pathways including Ras, PI3K and IRS-1 and therefore plays an important role in responding to signals from growth factors, amino acids and insulin. It also plays a role in cellular nutrient, oxygen and energy sensing.
mTOR related conditions and inhibitors
These broad activities mean that its dysregulation is implicated in a number of conditions including diabetes, obesity, cancer and Alzheimer’s disease.
It is also of great interest in the aging process and organ transplants, inhibitors such as rapamycin are used to prevent transplant rejection.
mTOR achieves its diverse roles through participating in two protein complexes referred to as mTOR complex 1 (mTORC1) and mTOR complex 2 (mTORC2) which differ in subcellular localization and binding partners.
While mTORC1 regulates translation through 4EBP1 and p70S6K, mTORC2 activates AKT through phosphorylation at Ser473.
AKT phosphorylation is lost in RICTOR Knockout Cell Lines. Cells starved in 0.5% fetal calf serum over night and subsequently stimulated with 10% FCS for 0,1,2 or 3h (as indicated).
Cells were washed with PBS, lysed in Frackelton buffer (10 mM Tris/HCl pH 7.5, 50 mM NaCl, 30 mM sodium pyrophosphate, 1 % Triton X-100, 50 mM NaF and protease inhibitors) and analyzed by SDS-PAGE and Western blotting using anti-phospho AKT (Ser 473; #4058 from Cell Signaling) or anti-AKT (#4691 from Cell Signaling). HAP1 wild-type cells were compared to cells bearing a frameshift mutation in RICTOR.
Read our application note on screening mTOR targeted compounds using Horizon Cell Lines.
Examples of popular knockout cell lines for mTOR Pathway
Human knockout HAP-1 cells
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Cancer-related cell lines
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Cas9 Stable Cell Lines
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CRISPRmod CRISPRa dCas9-VPR Stable Cell Lines
Streamline CRISPR activation experiments with stably expressing dCas9-VPR cell lines
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The PI3K pathway is aberrantly activated in numerous cancers, either through oncogenic activation of upstream effectors or through mutation of PI3K itself.
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