Exercise has beneficial effects on human health, including protection against metabolic disorders such as diabetes1. However, the cellular mechanisms underlying these effects are incompletely understood. The lysosomal degradation pathway, autophagy, is an intracellular recycling system that functions during basal conditions in organelle and protein quality control2. During stress, increased levels of autophagy permit cells to adapt to changing nutritional and energy demands through protein catabolism3. Moreover, in animal models, autophagy protects against diseases such as cancer, neuro-degenerative disorders, infections, inflammatory diseases, ageing and insulin resistance4-6. Here we show that acute exercise induces autophagy in skeletal and cardiac muscle of fed mice. To investigate the role of exercise-mediated autophagy in vivo, we generated mutant mice that show normal levels of basal autophagy but are deficient in stimulus (exercise- or starvation)-induced autophagy. These mice (termed BCL2 AAA mice) contain knock-in mutations in BCL2 phosphorylation sites (Thr69Ala, Ser70Ala and Ser84Ala) that prevent stimulus-induced disruption of the BCL2-beclin-1 complex and autophagy activation. BCL2 AAA mice show decreased endurance and altered glucose metabolism during acute exercise, as well as impaired chronic exercise-mediated protection against high-fat-diet-induced glucose intolerance. Thus, exercise induces autophagy, BCL2 is a crucial regulator of exercise- (and starvation)- induced autophagy in vivo, and autophagy induction may contribute to the beneficial metabolic effects of exercise.

The lysosomal degradation pathway of autophagy has a crucial role in defence against infection, neurodegenerative disorders, cancer and ageing. Accordingly, agents that induce autophagy may have broad therapeutic applications. One approach to developing such agents is to exploit autophagy manipulation strategies used by microbial virulence factors. Here we show that a peptide, Tat–beclin 1—derived from a region of the autophagy protein, beclin 1, which binds human immunodeficiency virus (HIV)-1 Nef—is a potent inducer of autophagy, and interacts with a newly identified negative regulator of autophagy, GAPR-1 (also called GLIPR2). Tat–beclin 1 decreases the accumulation of polyglutamine expansion protein aggregates and the replication of several pathogens (including HIV-1) in vitro, and reduces mortality in mice infected with chikungunya or West Nile virus. Thus, through the characterization of a domain of beclin 1 that interacts with HIV-1 Nef, we have developed an autophagy-inducing peptide that has potential efficacy in the treatment of human diseases.

Aberrant signaling through the class I phosphatidylinositol 3-kinase (PI3K)-Akt axis is frequent in human cancer. Here we show that Beclin 1, an essential autophagy and tumor suppressor protein, is a target of the protein kinase Akt. Expression of a Beclin 1 mutant resistant to Akt-mediated phosphorylation increased autophagy, reduced anchorage-independent growth, and inhibited Akt-driven tumorigenesis. Akt-mediated phosphorylation of Beclin 1 enhanced its interactions with 14-3-3 and vimentin intermediate filament proteins, and vimentin depletion increased autophagy and inhibited Akt-driven transformation. Thus, Akt-mediated phosphorylation of Beclin 1 functions in autophagy inhibition, oncogenesis, and the formation of an autophagy-inhibitory Beclin 1/14-3-3/vimentin intermediate filament complex. These findings have broad implications for understanding the role of Akt signaling and intermediate filament proteins in autophagy and cancer.

Summary Cell surface growth factor receptors couple environmental cues to the regulation of cytoplasmic homeostatic process including autophagy, and aberrant activation of such receptors is a common feature of human malignancies. Here, we defined the molecular basis by which the epidermal growth factor receptor (EGFR) tyrosine kinase regulates autophagy. Active EGFR binds to the autophagy protein Beclin 1, leading to its multisite tyrosine phosphorylation, enhanced binding to inhibitors, and decreased Beclin 1-associated Class III phosphatidylinositol-3 kinase activity. EGFR tyrosine kinase inhibitor (TKI) therapy disrupts Beclin 1 tyrosine phosphorylation and binding to its inhibitors, and restores autophagy in non-small cell lung carcinoma (NSCLC) cells with a TKI-sensitive EGFR mutation. In NSCLC tumor xenografts, the expression of a tyrosine phosphomimetic Beclin 1 mutant leads to reduced autophagy, enhanced tumor growth, tumor dedifferentiation, and resistance to TKI therapy. Thus, oncogenic receptor tyrosine kinases directly regulate the core autophagy machinery, which may contribute to tumor progression and chemoresistance.

Autophagy functions in antiviral immunity. However, it is not yet known whether endogenous autophagy genes protect against viral disease in vertebrates. Using three different approaches to inactivate the autophagy gene Atg5 in virally-infected neurons, we found that loss of Atg5 function increases mouse susceptibility to lethal Sindbis virus CNS infection. This phenotype is associated with delayed clearance of viral proteins, increased accumulation of the cellular p62 adaptor protein, and increased cell death in neurons, but not with altered levels of CNS viral replication. In vitro, p62 interacts with Sindbis virus capsid protein and genetic knockdown of p62 blocks the targeting of viral capsid to autophagosomes. Moreover, p62 or autophagy gene knockdown increases viral capsid accumulation and accelerates virus-induced cell death without affecting virus replication. These results suggest a novel function for autophagy in mammalian antiviral defense: a cell-autonomous mechanism in which p62 adaptor-mediated autophagic viral protein clearance promotes cell survival.

Autophagy is commonly observed in metazoan organisms during programmed cell death (PCD), but its function in dying cells has been unclear. We studied the role of autophagy in embryonic cavitation, the earliest PCD process in mammalian development. Embryoid bodies (EBs) derived from cells lacking the autophagy genes, atg5 or beclin 1, fail to cavitate. This defect is due to persistence of cell corpses, rather than impairment of PCD. Dying cells in autophagy gene null EBs fail to express the "eat-me" signal, phosphatidylserine exposure, and secrete lower levels of the "come-get-me" signal, lysophosphatidylcholine. These defects are associated with low levels of cellular ATP and are reversed by treatment with the metabolic substrate, methylpyruvate. Moreover, mice lacking atg5 display a defect in apoptotic corpse engulfment during embryonic development. We conclude that autophagy contributes to dead-cell clearance during PCD by a mechanism that likely involves the generation of energy-dependent engulfment signals.

Autophagy increases lifespan of model organisms; however, its role in promoting mammalian longevity is less well-established1,2. Here, we report lifespan and healthspan extension in a mouse model with increased basal autophagy. To determine the effects of constitutively increased autophagy on mammalian health, we generated targeted mutant mice with a F121A (Becn1F121A/F121A) mutation in beclin 1 that decreases its interaction with the negative regulator, Bcl-2. We demonstrate that beclin 1/Bcl-2 interaction is disrupted in multiple tissues in Becn1F121A/F121A knock-in (KI) mice in association with higher levels of basal autophagic flux. Compared to wild-type (WT) littermates, the lifespan of both male and female KI mice is significantly increased. The healthspan of the KI mice also improves as aging-related phenotypes are diminished, including age-related renal and cardiac pathological changes and spontaneous tumorigenesis. Moreover, mice deficient in the anti-aging protein, Klotho3, have increased beclin 1/Bcl-2 interaction, decreased autophagy, premature lethality and infertility which are rescued by the beclin 1 F121A mutation. Taken together, our data demonstrate that disruption of the beclin 1/Bcl-2 complex is an effective mechanism to increase autophagy, prevent premature aging, improve healthspan and promote longevity in mammals.

Selective autophagy involves the recognition and targeting of specific cargo, such as damaged organelles, misfolded proteins, or invading pathogens for lysosomal destruction1–4. Yeast genetic screens have identified proteins required for different forms of selective autophagy, including cytoplasm-to-vacuole targeting, pexophagy, and mitophagy, and mammalian genetic screens have identified proteins required for autophagy regulation5. However, there have been no systematic approaches to identify molecular determinants of selective autophagy in mammalian cells. To identify mammalian genes required for selective autophagy, we performed a high-content, image-based, genome-wide siRNA screen to detect genes required for the colocalization of Sindbis virus capsid protein with autophagolysosomes. We identified 141 candidate genes required for viral autophagy, which were enriched for cellular pathways related to mRNA processing, interferon signaling, vesicle trafficking, cytoskeletal motor function, and metabolism. Ninety-six of these genes were also required for Parkin-mediated mitophagy, indicating that common molecular determinants may be involved in autophagic targeting of viral nucleocapsids and autophagic targeting of damaged mitochondria. Murine embryonic fibroblasts lacking one of these gene products, the C2-domain containing protein, Smurf1, are deficient in the autophagosomal targeting of Sindbis and herpes simplex viruses and in the clearance of damaged mitochondria. Moreover, Smurf1-deficient mice display an accumulation of damaged mitochondria in heart, brain, and liver. Thus, our study identifies candidate determinants of selective autophagy, and defines Smurf1 as a newly recognized mediator of both viral autophagy and mitophagy.