The biology of fasting
mTOR
Also called: mTORC1, Mechanistic target of rapamycin
Written by Aaron CuhaReviewed Sep 2026
The kinase that decides whether a cell should be building or conserving, and the reason protein specifically, rather than calories generally, is what ends the fasted state.
What it is
mTOR is a large kinase that exists in two different multiprotein assemblies. The one this page is about is mTORC1, defined by a scaffold protein called raptor, and it is the nutrient sensitive one. The other, mTORC2, has different partners and different jobs and is not covered here.
mTORC1's function is to make a single decision on the cell's behalf: build, or conserve. When it is active, the cell makes protein, makes lipid, makes nucleotides and holds recycling in check. When it is off, protein synthesis falls, autophagy is released, and the cell scales up its lysosomal capacity.
It takes at least four independent classes of input: amino acids, cellular energy status, growth factor and insulin signalling, and stress. It needs several of them to be favourable at once. That is why insulin alone does not fully switch it on when amino acids are absent, and it is the reason a fast suppresses it on more than one arm simultaneously.
The machinery
The central architectural fact is that activating mTORC1 is a question of location, not just of kinase chemistry. With no amino acids around, mTORC1 floats in the cytosol. With amino acids present it is recruited to the surface of the lysosome, and its direct activator, a small GTPase called Rheb, lives on that surface. If mTORC1 never gets to the lysosome, Rheb cannot switch it on no matter what else is true.
The recruitment is done by the Rag GTPases, a family of four that work as heterodimers, bind raptor in an amino acid sensitive way, and are required for amino acids to activate mTORC1 at all. A Rag mutant locked in its GTP-bound state binds mTORC1 strongly and makes the pathway resistant to amino acid withdrawal; a GDP-locked mutant abolishes amino acid stimulation. The Rags do not make mTOR a better kinase. They move it to where Rheb is.
Three specific amino acid sensors feed into this, and all three work by the same inverted logic. Sestrin2 binds and inhibits the GATOR2 complex when leucine is scarce; leucine binds directly into a single pocket in Sestrin2 and that binding is what breaks the Sestrin2 to GATOR2 interaction. CASTOR1 does the same job for arginine. SAMTOR senses methionine indirectly, through S-adenosylmethionine, whose concentration tracks methionine availability, and acts on GATOR1.
Follow that backwards and the reason protein is privileged becomes obvious. Every dedicated, structurally solved chemical sensor in this pathway is a sensor for leucine, arginine or SAM. A meal of pure fat presents a ligand to none of them. A meal containing protein presents leucine and arginine within minutes of digestion.
Insulin acts on a separate arm. It activates PI3K and AKT, AKT inhibits the TSC1/TSC2 complex, TSC2 is the switch-off enzyme for Rheb, so inhibiting TSC raises Rheb-GTP and therefore mTORC1 activity at the lysosome. A fast closes both arms at once and AMPK adds a third inhibitory input by phosphorylating TSC2 and raptor directly.
Downstream, mTORC1 phosphorylates S6K1 at threonine 389 and 4E-BP1, which together control cap dependent translation initiation; phosphorylates ULK1 at serine 757 to hold autophagy off; and phosphorylates the transcription factor TFEB to keep it out of the nucleus, which is how lysosomal capacity, not just autophagy initiation, is held down.
What switches it on
Falling leucine, arginine and methionine, falling insulin, and rising AMPK activity. Any one of those pushes mTORC1 down; a fast supplies all three. Protein restriction without calorie restriction reaches this node through the amino acid sensors alone.
On the timing
There is no continuous human time course of mTORC1 activity across a multi-day fast in multiple tissues, because measuring it means taking a muscle biopsy and running a Western blot. What exists is a small number of biopsy timepoints. At 72 hours, mTOR phosphorylation in human skeletal muscle is down by roughly half. In the first 8 hours of a fast, the mTORC1 related readouts do not move substantially. Between those two anchors, nobody has looked.
What has been measured
In people
mTORC1 activity in a living person is measured as the phosphorylation state of the kinase and its targets in a needle biopsy of skeletal muscle. There is no imaging method and no blood test for tissue mTORC1 activity. Within that limit, the 72 hour result is clear and consistent, and there is a separate line of human evidence from drugs that inhibit the pathway on purpose.
- 72 hour fast, 8 healthy men, forearm amino acid tracer kinetics plus vastus lateralis biopsies. mTOR phosphorylation at serine 2448 fell by approximately 50%, with reduced phosphorylation of the downstream targets 4EBP1, rpS6 and ULK1. In the same biopsies LC3B-II rose about 30% and p62 rose about 10%, which is a genuinely ambiguous combination and is reported here as it was measured (PLoS One, 2014).
- 72 hour fast, obese versus lean subjects, with the same signalling readouts. mTOR and ULK1 serine 757 signalling in skeletal muscle was measured before and after the fast in both groups (American Journal of Physiology: Endocrinology and Metabolism, 2016).
- The first 8 hours, 10 healthy men, biopsies at 4 and 8 hours, with and without roughly 400 kcal of added arm ergometer exercise. The mTORC1 related readouts did not move substantially, and adding whole body energetic stress did not change that (Pflugers Archiv, 2021).
- Pharmacological inhibition, 2014. Elderly volunteers given the rapalog RAD001 showed an approximately 20% enhanced response to influenza vaccination at doses that were relatively well tolerated, along with a reduced percentage of CD4 and CD8 T lymphocytes expressing PD-1 (Science Translational Medicine, 2014).
- Pharmacological inhibition, 2018, and this is the larger trial. A phase 2a randomised placebo controlled trial in 264 elderly subjects used a low dose combination of a catalytic and an allosteric inhibitor selective for TORC1 for 6 weeks. It was associated with a significant decrease in the rate of infections reported over the following year (P = 0.001), upregulation of antiviral gene expression, and improved influenza vaccine response (Science Translational Medicine, 2018).
- The caveat that undercuts the whole readout, and it belongs here rather than in a footnote. In six young men who performed resistance exercise and consumed 0, 5, 10, 20 or 40 g of whole egg protein, muscle protein synthesis measured by stable isotope infusion showed a clear dose response and was maximally stimulated at 20 g. Phosphorylation of S6K1 threonine 389, ribosomal protein S6 serine 240/244 and eIF2B epsilon serine 539 was unaffected by protein ingestion. In the tissue and the timeframe where the synthetic output moved substantially, the canonical mTORC1 phospho-markers did not move at all (American Journal of Clinical Nutrition, 2009).
In other species and in cell culture
The entire sensing architecture was worked out by biochemistry, structural biology and cell culture, largely in immortalised human derived cell lines with purified proteins. The lifespan evidence for inhibiting this pathway is in mice, and it is unusually well controlled mouse evidence.
- Cultured cell lines and purified proteins: the Rag GTPases binding raptor and mediating amino acid signalling to mTORC1 (Science, 2008); Sestrin2 as the leucine sensor and the structure showing leucine in a single pocket (Science, 2016, two papers); CASTOR1 as the arginine sensor and the structural mechanism of arginine sensing (Cell, 2016; Nature, 2016); SAMTOR as the S-adenosylmethionine sensor (Science, 2017). These are human proteins in human derived cells. That is human molecular biology, and it is not measurement in a living person.
- Cultured cells: mTORC1 phosphorylating ULK1 at serine 757 to hold autophagy initiation off, and AMPK phosphorylating other sites to switch it on (Nature Cell Biology, 2011).
- Mouse lifespan, three independent sites, genetically heterogeneous UM-HET3 mice. Rapamycin fed from 600 days of age, which is late in a mouse's life, extended median and maximal lifespan in both sexes. On the basis of age at 90% mortality the increase was 14% for females and 9% for males. The effect appeared at all three sites and disease patterns in treated mice did not differ from controls. The authors state the effect could be postponement of cancer death, retardation of ageing, or both (Nature, 2009).
- Mouse lifespan, dose escalation. At a threefold higher dose, rapamycin increased median lifespan by 23% in males and 26% in females, with maximal longevity increased in both sexes, and the effect was larger in females at every dose tested. The same paper reports that several endocrine and metabolic changes seen in diet restricted mice are not seen in rapamycin treated mice, and that hepatic xenobiotic metabolism gene expression differs markedly between the two, concluding the interventions are distinct (Aging Cell, 2014).
Why it matters
mTORC1 is where the specific claim that fasting is about more than calories can be made precisely rather than vaguely. The sensors are for leucine, arginine and methionine. Nothing in the pathway binds a fatty acid or a sugar. That is the mechanistic reason a protein containing meal ends the fasted signalling state more decisively than the same energy as carbohydrate or fat, and it is why protein restriction and energy restriction are not the same intervention at this node.
It is also the node with the strongest pharmacological longevity evidence in a mammal. Rapamycin extending lifespan in genetically heterogeneous mice at three independent sites is one of the most robust results in the biology of ageing. It is a mouse lifespan result. No human lifespan trial of rapamycin or any rapalog exists. What does exist in humans is immune function and infection rate over one year in older adults, which is a real and useful endpoint and a different one.
A claim you will see repeated
Rapamycin is often described as a fasting mimetic, and fasting is often described as natural rapamycin. Neither is accurate. Rapamycin chronically inhibits one arm of one node with none of the fuel switching, hormonal or autophagic context of a fast, and the dose escalation paper found the metabolic signature of rapamycin treated mice to be clearly distinct from that of diet restricted mice. The second thing worth holding is the measurement caveat above. A study reporting that a fast lowered mTORC1 phospho-markers in human muscle has shown that the phospho-markers fell. In the one human experiment where synthetic output and phospho-markers were measured side by side, the output moved and the markers did not.
Citations
- Human2014Fasting increases human skeletal muscle net phenylalanine release and this is associated with decreased mTOR signaling
PLoS One
8 healthy men studied postabsorptive and after a 72 hour fast, with forearm amino acid tracer kinetics and vastus lateralis biopsies. mTOR phosphorylation at serine 2448 fell approximately 50%, with reduced phosphorylation of 4EBP1 threonine 46, rpS6 serine 235/236 and ULK1 serine 757. LC3B-II rose approximately 30% and p62 rose approximately 10%.
- Human2016Differential regulation of lipid and protein metabolism in obese vs. lean subjects before and after a 72-h fast
American Journal of Physiology: Endocrinology and Metabolism
Skeletal muscle mTOR and ULK1 serine 757 signalling measured in obese and lean subjects before and after a 72 hour fast, alongside lipid and protein metabolism.
- Human2021Increasing whole-body energetic stress does not augment fasting-induced changes in human skeletal muscle
Pflugers Archiv, European Journal of Physiology
10 healthy males, two supervised fasts with and without 2 hours of arm ergometer exercise adding roughly 400 kcal, vastus lateralis biopsies before, at 4 hours and at 8 hours. Signalling, mRNA and protein expression and substrate storage appeared unaffected by whole body energetic stress during the initial hours of fasting.
- Human2014mTOR inhibition improves immune function in the elderly
Science Translational Medicine
Elderly volunteers given the rapalog RAD001. The vaccine response to influenza was enhanced by about 20% at doses that were relatively well tolerated, and the percentage of CD4 and CD8 T lymphocytes expressing PD-1 fell.
- Human2018TORC1 inhibition enhances immune function and reduces infections in the elderly
Science Translational Medicine
Phase 2a randomised placebo controlled trial in 264 elderly subjects, low dose combination of a catalytic and an allosteric TORC1 selective inhibitor for 6 weeks. Significant decrease in the rate of infections reported over the following year (P = 0.001), upregulation of antiviral gene expression, and improved influenza vaccine response.
- Human2009Ingested protein dose response of muscle and albumin protein synthesis after resistance exercise in young men
American Journal of Clinical Nutrition
6 healthy young men, five trials, leg resistance exercise then 0, 5, 10, 20 or 40 g whole egg protein, synthesis measured over 4 hours by primed constant leucine infusion. Muscle protein synthesis was maximally stimulated at 20 g and leucine oxidation rose significantly at 20 and 40 g. Phosphorylation of S6K1 threonine 389, rpS6 serine 240/244 and eIF2B epsilon serine 539 was unaffected by protein ingestion.
- In vitro2008The Rag GTPases bind raptor and mediate amino acid signaling to mTORC1
Science
Cultured cells. The Rag GTPases bind raptor in an amino acid sensitive manner and are required for amino acid activation of mTORC1. They do not stimulate mTOR kinase activity directly; they relocalise mTORC1 to the lysosomal surface where its activator Rheb resides.
- In vitro2016Sestrin2 is a leucine sensor for the mTORC1 pathway
Science
Cultured cells and purified proteins. Sestrin2 binds and inhibits GATOR2 when leucine is scarce; leucine binds Sestrin2 directly with a dissociation constant in the physiologically relevant range and releases GATOR2, permitting mTORC1 activation.
- In vitro2016Structural basis for leucine sensing by the Sestrin2-mTORC1 pathway
Science
Structural work showing leucine bound in a single pocket in Sestrin2, and that this binding is what breaks the Sestrin2 to GATOR2 interaction.
- In vitro2016The CASTOR Proteins Are Arginine Sensors for the mTORC1 Pathway
Cell
Cultured cells and purified proteins. CASTOR1 binds and inhibits GATOR2; arginine binds CASTOR1 directly and disrupts the CASTOR1 to GATOR2 interaction.
- In vitro2016Mechanism of arginine sensing by CASTOR1 upstream of mTORC1
Nature
Structure of arginine bound CASTOR1, establishing the chemical basis of arginine sensing upstream of mTORC1.
- In vitro2017SAMTOR is an S-adenosylmethionine sensor for the mTORC1 pathway
Science
Cultured cells and purified proteins. Methionine is sensed indirectly through S-adenosylmethionine. SAMTOR binds GATOR1 and inhibits mTORC1; SAM binds SAMTOR and disrupts that interaction.
- In vitro2011AMPK and mTOR regulate autophagy through direct phosphorylation of Ulk1
Nature Cell Biology
Cultured cells. mTORC1 phosphorylates ULK1 at serine 757 to hold autophagy initiation off when nutrients are plentiful, and AMPK phosphorylates other sites on the same kinase to switch it on when energy is short.
- Animal2009Rapamycin fed late in life extends lifespan in genetically heterogeneous mice
Nature
Genetically heterogeneous UM-HET3 mice at three independent sites, rapamycin started at 600 days of age. Median and maximal lifespan increased in both sexes; on the basis of age at 90% mortality the increase was 14% for females and 9% for males. Disease patterns in treated mice did not differ from controls. A separate cohort started at 270 days also showed increased survival at interim analysis.
- Animal2014Rapamycin-mediated lifespan increase in mice is dose and sex dependent and metabolically distinct from dietary restriction
Aging Cell
At a threefold higher dose than the original studies, rapamycin increased median lifespan 23% in males and 26% in females, with maximal longevity increased in both sexes and larger effects in females at every dose. Several endocrine and metabolic changes seen in diet restricted mice were not seen in rapamycin treated mice, and hepatic xenobiotic metabolism gene expression was quite distinct between the two.